Grasasyaceites

Grasasyaceites

International Journal of Fats and Oils

International Journal of Fats and Oils Volumen 62

abril-junio 2011

N.º 2

116 págs.

ISSN: 0017-3495

Volumen 62

N.º 2

abril-junio 2011

Sevilla (España)

ISSN: 0017-3495

Sumario

Editorial / Editorial



Revisión / REview

A. Valenzuela, B. Delplanque y M. Tavella—El ácido esteárico: un posible sustituto para los ácidos grasos trans de origen industrial. / Stearic acid: a possible substitute for trans fatty acids from industrial origin.

131

Investigación / RESEARCH

H.M. Ibraim, A.A. Abou-Arab y F.M. Abu Salem—Efecto antioxidante y antimicrobiano de algunos extractos de plantas naturales añadidos a pastel de cordero durante el almacenamiento. / Antioxidant and antimicrobial effects of some natural plant extracts added to lamb patties during storage.

139

L. Huo, R. Lu, P. Li, Y. Liao, R. Chen, Ch. Deng, Ch. Lu, X. Wei y Y. Li—Actividad antioxidante, fenoles totales y flavonoides totales en extractos de tallos de Jasminum nervosum Lour. / Antioxidant activity, total phenolic, and total flavonoid of extracts from the stems of Jasminum nervosum Lour.

149

G. Budryn, E. Nebesny y D. ¶y‡elewicz—Estabilidad oxidativa de manteca y aceite de girasol suplementados con extractos de café bajo condiciones de almacenamiento. / Oxidative stability of lard and sunflower oil supplemented with coffee extracts under storage conditions.

155

M. Osman, G.I. Mahmoud, R.M. Romeilah y S.A. Sayed—Semillas de altramuces bajan la concentración de lípidos plasmáticos y normaliza los parámetros antioxidantes en ratas. / Lupin seeds lower plasma lipid concentrations and normalize antioxidant parameters in rats.

162

A.L.M.T. Pighinelli, R.A. Ferrari, A.M.R.O. Miguel y K.J. Park—Biodiesel de girasol alto oleico: calidad y diferentes métodos de purificación. / High oleic sunflower biodiesel: quality control and different purification methods.

171

S.T. Jiang y L.Y. Niu—Optimización y evaluación del aceite de germen de trigo extraído por CO2 supercrítico. / Optimization and evaluation of wheat germ oil extracted by supercritical CO2.

181

S.A.S. Chatha, A.I. Hussain, J.R. Bajwa, S.T.H. Sherazi y Aiza Shaukat—Extracto de salvado de trigo: una potente fuente de antioxidantes naturales para la estabilización del aceite de canola. / Wheat bran extracts: a potent source of natural antioxidants for the stabilization of canola oil.

190

D.K. Saxena, S.K. Sharma y S.S. Sambi—Cinética y termodinámica de la extracción del aceite de algodón. / Kinetics and thermodynamics of cottonseed oil extraction.

198

B. Hussain, S. Mahboob, M. Hassan, S. Nadeem y T. Sultana—Efecto del grado de maduración en el perfil de ácidos grasos de diferentes tejidos de rohu (Labeo rohita) salvaje y criado. / Effect of maturation degree on fatty acid profile of different tissues in wild and farmed rohu (Labeu rohita).

206

B. Krishna De y S. Verma—Caracterización de lípidos y ácidos grasos de hongos derivados del suelo Cladosporium sp. / Characterization of lipids and fatty acids of the soil derived fungus Cladosporium sp.

213

información / technical information J. Vilar Hernández, M.M. Velasco Gámez, R. Puentes Poyatos y A.M. Martínez Rodríguez—El olivar tradicional: alternativas estratégicas de competitividad. / Traditional olive growing system: Strategic alternatives to improve competitiveness.

Documentación / documentation

http://grasasyaceites.revistas.csic.es

221

Grasas y aceites Volumen 62 | N.º 2 | 2011 | Sevilla



Instituto

www.publicaciones.csic.es

CONSEJO SUPERIOR DE INVESTIGACIONES CIENTÍFICAS

de la

Grasa

Grasasyaceites International Journal of Fats and Oils

Volumen 62 • N.° 2

abril-junio 2011 Sevilla (España)

ISSN: 0017-3495

CONSEJO SUPERIOR DE INVESTIGACIONES CIENTÍFICAS

Volumen 62 • N.° 2

abril-junio 2011 Sevilla (España)

ISSN: 0017-3495

grasas y aceites

revista publicada por el instituto de la grasa (csic)

La revista Grasas y Aceites (Grasas Aceites), de periodicidad trimestral, es una publicación dedicada a la información científica y técnica sobre grasas comestibles y sus derivados. Publica trabajos de investigación originales, artículos de información, notas de laboratorio, trabajos de revisión así como bibliografía sobre revistas, patentes o libros. El campo que cubre se refiere fundamentalmente a frutos y semillas oleaginosas, materias grasas, productos afines o derivados y aceitunas de mesa. Igualmente, incluye trabajos relacionados con subproductos de todas las materias anteriores y el tratamiento de las aguas residuales de las industrias correspondientes. Los originales recibidos son evaluados por el Consejo de Redacción y por evaluadores externos. Se publica en edición electrónica, modelo acceso abierto en: http://grasasyaceites.revistas.csic.es.

Grasas y Aceites is published by Instituto de la Grasa (CSIC) Grasas y Aceites (Grasas Aceites) is a quarterly published journal devoted to scientific and technological information on the field of edible fat nd oils and their derivatives. Grasas y Aceites publishes full research articles, research notes, reviews as well as information on references, patents, and books. Grasas y Aceites covers the following fields: oleaginous fruit and seeds, edible fatty materials as well as related products and their derivatives, including table olives. It also accept works related to by-products from all the previous materials and the handling and treatment of the wastewaters from the corresponding industries. Originals are always reviewed by the Editorial Board and by qualified experts. There is an electronic edition, open access model, at: http://grasasyaceites.revistas.csic.es

Directora: M.ª Carmen Pérez Camino (Instituto de la Grasa) Secretario: José M.ª García Martos (Instituto de la Grasa) R. Abia González (Instituto de la Grasa) J. A. Cayuela Sánchez (Instituto de la Grasa) José M.ª García Martos (Instituto de la Grasa) M. León Camacho (Instituto de la Grasa) G. Márquez Ruiz (Instituto del Frío) A. Montaño Asquerino (Instituto de la Grasa) M.ª T. Morales Millán (Universidad de Sevilla)

Consejo de Redacción: M.ª Carmen Pérez Camino (Instituto de la Grasa) J. J. Ruiz Barba (Instituto de la Grasa) M.ª V. Ruiz Méndez (Instituto de la Grasa) J. J. Salas Liñán (Instituto de la Grasa) J. Sánchez Perona (Instituto de la Grasa) J. Velasco Jiménez (Instituto de la Grasa) I. Vicario Romero (Universidad de Sevilla) J. Vioque Peña (Instituto de la Grasa)

Consejo Asesor Científico: D. Barrera Arellano (Unicamp, Brasil) A. Jiménez Márquez (CIFA, Mengíbar, Jaén) P. Belton (IFR, U.K.) A. P. Kiritsakis (Technological Educational Institution, Greece) W. W. Christie (SCRI, Scotland, U.K.) G. Lecker (Istituto di Industrie Agrarie, Italy) P. Dais (University of Crete, Greece) J. Linares Fernández (Grupo SOS, Jaén) M. J. Dennis (CSL, Cork, U.K.) C. Mariani (Stazione Sperimentale per le Industrie degli Oli e E. Dubinsky (Eduardo Dubinsky y Asociados, Argentina) dei Grassi, Italy) C. Gallegos (Universidad de Huelva) J. M. Ordovás (Tufs University, USA) P. P. García de Luna (Hospital Univ. Virgen del Rocio, Sevilla) R. Przybylski (University of Manitoba, Canadit) H. Glaeser (European Commission Directorate, Bruxelles) J. L. Sébedio (Institut National de la Recherche Agronomique, C. Gómez Herrera (Instituto de la Grasa, Sevilla) France) J. L. Hanwood (University of Wales, U.K.) M. Tsimidou (Aristotle University of Thessaloniki, Greece) J. R. Izquierdo Álvarez-Buylla (Ministerio de Medio Ambiente P.J. White (Iowa State University, USA) y Medio Rural y Marino, Madrid) Secretaría Técnica: María Teresa Sánchez REDACCIÓN E INTERCAMBIO Revista Grasas y Aceites Instituto de la Grasa Avda. Padre García Tejero, 4 41012 Sevilla (Spain) Tel.: +34 954 611 550 Fax: +34 954 616 790 e-mail: [email protected] http://www.ig.csic.es/revis.html

DISTRIBUCIÓN, SUSCRIPCIÓN Y VENTA Departamento de Publicaciones Librería Científica Medinaceli Sección de revistas Duque de Medinaceli, 6 Vitruvio, 8 28014 Madrid 28006 Madrid (Spain) Tel.: +34 914 295 684 Tels.: +34 915 612 833, e-mail: [email protected] Tels.: +34 915 681 619/620/640 Fax: +34 915 629 634 e-mail: [email protected]

SERVICIO DE INFORMACIÓN Los artículos publicados por Grasas y Aceites son recogidos, entre otras, por las siguientes bases de datos: C.A.S. Chemical Abstracts Services (USA), I.A.L.I.N.E. (Francia), C.A.B.S. Current Awareness in Biological Sciences (Gran Bretaña), F.S.T.A. Food Science and Technology Abstracts (USA-Gran Bretaña), BIOSIS Biological Abstracts (USA), SCISEARCH Science Citation Index Search (USA), SWETS (Holanda), CAB Commowealth Agricultural Bureaux (Gran Bretaña), OSTI Office of Scientific and Technological Information (USA), FOODS-ADLIBRA (USA), PASCAL (Francia), A.A. Analytical Abstract (Gran Bretaña), Cambridge Scientific Abstracts (USA), Current Contents (USA), Deutsche Gessellscharft für Fettiwissenschaft (Alemania), ICYT Índice Español de Ciencia y Tecnología España) y en el CINDOC. © CSIC 2011 Cada autor es responsable del contenido de sus respectivos trabajos. El Consejo Superior de Investigaciones Científicas no se hace responsable, en ningún caso, de la credibilidad y autenticidad de los trabajos. Los originales de la revista Grasas y Aceites, publicados en papel y en versión electrónica, son de propiedad del Consejo Superior de Investigaciones Científicas, siendo necesario citar la procedencia en cualquier reproducción parcial o total. ISSN: 0017-3495 eISSN 1988-4214 NIPO (en papel): 472-11-043-X NIPO (en línea): 472-11-044-5 Depósito legal: M-862, 1958 Impreso en España/Printed in Spain Realización: DiScript Preimpresión, S. L.

Authors take full responsibility for all statements or opinions included in their papers. CSIC does not assume any liability with respect to the credibility or authenticity of the contributions. Articles appeared in Grasas y Aceites, both in the printed and the electronic versions, are property of the CSIC and it is needed to cite the origin of the article in any total or partial reproduction.

GRASAS Y ACEITES

Volumen 62 • N.° 2

abril-junio 2011 Sevilla (España)

ISSN: 0017-3495

Grasasyaceites International Journal of Fats and Oils

CONTENIDO

CONTENTS

Editorial

Editorial

Revisión

Review

El ácido esteárico: un posible sustituto para los ácidos grasos trans de origen industrial.- A. Valenzuela, B. Delplanque y M. Tavella . . . . . . . . . . . . . . . . . . . . . . . . .

Stearic acid: a possible substitute for trans fatty acids from industrial origin.A. Valenzuela, B. Delplanque and M. Tavella . . . . . . . . . . . . . . . . . . . . . . . . . . .

131

131

Investigación

Research

Efecto antioxidante y antimicrobiano de algunos extractos de plantas naturales añadidos a pastel de cordero durante el almacenamiento.- H.M. Ibraim, A.A. Abou-Arab y F.M. Abu Salem . . . . . . . . . .

139

Antioxidant and antimicrobial effects of some natural plant extracts added to lamb patties during storage.- H.M. Ibraim, A.A. Abou-Arab and F.M. Abu Salem . . . . . . . .................................

139

149

Antioxidant activity, total phenolic, and total flavonoid of extracts from the stems of Jasminum nervosum Lour. - L. Huo, R. Lu, P. Li, Y. Liao, R. Chen, Ch. Deng, Ch. Lu, X. Wei and Y. Li . . . . . . . . . . . . . . . . .

149

155

Oxidative stability of lard and sunflower oil supplemented with coffee extracts under storage conditions.- G. Budryn, E. Nebesny and D. ¶y‡elewicz . . . . . . . . . . . .................................

155

162

Lupin seeds lower plasma lipid concentrations and normalize antioxidant parameters in rats.- M. Osman, G.I. Mahmoud, R.M. Romeilah and S.A. Fayed . . . . . . . . . . . . . . . . . . . . . . . . . . . .

162

171

High oleic sunflower biodiesel: quality control and different purification methods.A.L.M.T. Pighinelli, R.A. Ferrari, A.M.R.O. Miguel and K.J. Park . . . . . . . . . . . . . . . .

171

Actividad antioxidante, fenoles totales y flavonoides totales en extractos de tallos de Jasminum nervosum Lour.- L. Huo, R. Lu, P. Li, Y. Liao, R. Chen, Ch. Deng, Ch. Lu, X. Wei y Y. Li . . . . . . . . . . . . . . . . Estabilidad oxidativa de manteca y aceite de girasol suplementados con extractos de café bajo condiciones de almacenamiento.- G. Budryn, E. Nebesny y D. ¶y‡elewicz . . . . . . . . . . . . . . . . . . . . . . Semillas de altramuces bajan la concentración de lípidos plasmáticos y normaliza los parámetros antioxidantes en ratas.- M. Osman, G.I. Mahmoud, R.M. Romeilah y S.A. Sayed . . . . . . . . . . Biodiesel de girasol alto oleico: calidad y diferentes métodos de purificación.A.L.M.T. Pighinelli, R.A. Ferrari, A.M.R.O. Miguel y K.J. Park . . . . . . . . . . . . . . . . . .



127

Optimización y evaluación del aceite de germen de trigo extraído por CO2 supercrítico.- S.T. Jiang y L.Y. Niu . . . . . . . . Extracto de salvado de trigo: una potente fuente de antioxidantes naturales para la estabilización del aceite de canola.S.A.S. Chatha, A.I. Hussain, J.R. Bajwa, S.T.H. Sherazi y Aiza Shaukat . . . . . . . . . Cinética y termodinámica de la extracción del aceite de algodón.- D.K. Saxena, S.K. Sharma y S.S. Sambi . . . . . . . . . . . . Efecto del grado de maduración en el perfil de ácidos grasos de diferentes tejidos de rohu (Labeo rohita) salvaje y criado.B. Hussain, S. Mahboob, M. Hassan, S. Nadeem y T. Sultana . . . . . . . . . . . . . . Caracterización de lípidos y ácidos grasos de hongos derivados del suelo Cladosporium sp.- B. Krishna De y S. Verma

181

Optimization and evaluation of wheat germ oil extracted by supercritical CO2..S.T. Jiang y L.Y. Niu . . . . . . . . . . . . . . . . .

181

190

Wheat bran extracts: a potent source of natural antioxidants for the stabilization of canola oil.- S.A.S. Chatha, A.I. Hussain, J.R. Bajwa, S.T.H. Sherazi and Aiza Shaukat . . . . . . . . . . . . . . . . . . . . . . . . . .

190

198

Kinetics and thermodynamics of cottonseed oil extraction.- D.K. Saxena, S.K. Sharma and S.S. Sambi . . . . . . . . . .

198

206

Effect of maturation degree on fatty acid profile of different tissues in wild and farmed rohu (Labeu rohita).- B. Hussain, S. Mahboob, M. Hassan, S. Nadeem and T. Sultana . . . . . . . . . . . . . . . . . . . . . . . . .

206

213

Characterization of lipids and fatty acids of the soil derived fungus Cladosporium sp.- B. Krishna De and S. Verma . . . . . . .

213

Información

Technical information

El olivar tradicional: alternativas estratégicas de competitividad.- J. Vilar Hernández, M.M. Velasco Gámez, R. Puentes Poyatos y A.M. Martínez Rodríguez . . . . .................................

Traditional olive growing system: Strategic alternatives to improve competitiveness.J. Vilar Hernández, M.M. Velasco Gámez, R. Puentes Poyatos and A.M. Martínez Rodríguez . . . . . . . . . . . . . . . . . . . . . . . .

221

Documentación Libros . . . . . . . . . . . . . . . . . . . . . . . . . . . .

128

221

Documentation 230

Reviews of new books . . . . . . . . . . . . . . .

230

grasas y aceites,

62 (2), 129, 2011, issn: 0017-3495

abril-junio,

Editorial Relevo en la dirección de Grasas y Aceites El Consejo de redacción de la revista Grasas y Aceites dirigido por la Prof. Dra. Rosario Zamora Corchero desde el año 2007, ha pasado el relevo a un nuevo equipo de trabajo. Desde su primera editorial, la Directora se propuso como objetivo fundamental el mejorar sustancialmente la posición de la revista en el conjunto de la literatura científica. Este objetivo se ha visto cumplido con creces. Con una eficaz estrategia de actuación, ha logrado que GRASAS Y ACEITES alcance en estos años una posición hasta ahora sin precedentes en su prolongada vida, al situarse en un puesto avanzado dentro del tercer cuartel en el campo de la Ciencia y Tecnología de los alimentos. A ello ha contribuido sin lugar a dudas la calidad de los trabajos seleccionados para su publicación, la inmediatez con que la revista ve la luz en el momento actual así como la iniciativa del Consejo Superior de Investigaciones Científicas (CSIC) de facilitar el acceso al texto completo de los trabajos desde la dirección: http://grasasyaceites.revistas.csic.es/index.php/grasasyaceites Todos estos logros han sido posible gracias, además de a la eficaz labor de la Directora de la Revista, a los miembros del Consejo de Redacción, y a los revisores, quienes de manera desinteresada han colaborado durante estos años a dar forma a esta revista, a ellos nuestra enhorabuena y gracias porque Grasas y Aceites es de todos. Para este nuevo Consejo de Redacción constituye un reto el recoger este testigo y es nuestra intención continuar, en la medida en que nos sea posible, la línea de actuación trazada por el equipo saliente, para seguir esta trayectoria ascendente en el índice de impacto de nuestra revista. Se continuará la edición de números especiales que contemplan distintos aspectos específicos dentro del mundo de los lípidos, nuestra temática general. Esta iniciativa ha tenido una interesante acogida como lo demuestra de manera objetiva el aumento de citas que ha experimentado la revista, debido en gran medida a estos artículos. Por último, exhortamos a los autores para que contribuyan a este empeño, con el envío de originales cada vez con mayor calidad científica y a nuestros revisores para que continúen con el riguroso examen de los manuscritos. Esto junto con la puntual salida de los números garantizarán a nuestra Revista un prometedor futuro. María del Carmen Pérez Camino

129

grasas y aceites,

62 (2), 131-138, 2011, issn: 0017-3495 doi: 10.3989/gya.033910

abril-junio,

REVISIÓN Stearic acid: a possible substitute for trans fatty acids from industrial origin By Alfonso Valenzuela,1* Bernadette Delplanque2 and Marcelo Tavella3 1

Instituto de Nutrición y Tecnología de los Alimentos (INTA), Universidad de Chile, Santiago, Chile. Facultad de Medicina, Universidad de los Andes, Santiago, Chile 2 Laboratoire de Neuroendocrinologie Moléculaire de la Prise Alimentaire (NMPA) UMR 1197, Université Paris Sud, Orsay, France. 3 Programa de Prevención del Infarto en Argentina - INIBIOLP. Universidad Nacional de La Plata, La Plata, Argentina. (*Corresponding author: [email protected])

RESUMEN El acido esteárico: un posible sustituto para los ácidos grasos trans de origen industrial. Los isómeros trans que contienen los aceites parcialmente hidrogenados de origen industrial, han sido cuestionados y la recomendación es reducir su consumo. La industria de alimentos se enfrenta a un dilema, ya que para disminuir los isómeros trans debe reducir los aceites parcialmente hidrogenados y reemplazarlos por grasas ricas en ácidos grasos saturados. La investigación ha demostrado que los ácidos grasos saturados tienen efecto negativo en los lípidos plasmáticos y su consumo se asocia con un alto riesgo cardiovascular, por lo cual la recomendación es reducir el consumo de estos ácidos grasos. Sin embargo, no todos los ácidos grasos saturados se comportan de la misma forma, siendo el ácido esteárico (18:0) la excepción. El ácido esteárico presenta bajo nivel de absorción intestinal y no modifica negativamente los lípidos plasmáticos, por lo cual se considera como “neutro” para la salud cardiovascular. Los niveles plasmáticos de la apoproteína B-100, que determina las concentraciones de las VLDL y LDL (transportadoras de triglicéridos y colesterol, respectivamente) no son modificados por dietas que aportan hasta 7% de la energía como ácido esteárico. Marcadores de riesgo cardiovascular, como la activación de factores de agregación plaquetaria o los niveles de la proteína C reactiva, no son modificados por dietas que aportan ácido esteárico, como ocurre con otros ácidos grasos saturados. La confirmación del efecto “neutro” del ácido esteárico es una perspectiva para el desarrollo de grasas con alto contenido de este ácido graso para reemplazar las grasas hidrogenadas que contienen isómeros trans. Esta revisión discute estos aspectos. PALABRAS-CLAVE: Ácido esteárico – Alternativa a las grasas hidrogenadas – Efecto metabólico neutro – Isómeros trans – Salud cardiovascular. SUMMARY Stearic acid: a possible substitute for trans fatty acids from industrial origin. Trans isomers, contained in partially hydrogenated oils, which are used in the food industry, have been questioned



and nowadays trends are heading towards reducing their consumption. The food industry is facing a dilemma, since in order to remove trans fatty acids, hydrogenated fats should be eliminated and replaced by fats rich in saturated fatty acids. Scientific research has shown that saturated fatty acids have negative effects on the lipid profile and its consumption is associated with a higher cardiovascular risk. Therefore it is recommended to avoid their consumption. Nevertheless, not all fatty acids behave in the same way, with stearic acid (18:0) the exception. Stearic acid has a low level of intestinal absorption and its intake does not negatively modify the lipid profile. For this reason, it is considered a “neutral” fatty acid with regard to cardiovascular health. B-100 apolipoprotein, whose levels determine plasma VLDL and LDL concentration (triglycerides and cholesterol carriers, respectively), is not modified by diets which provide up to 7% of the energy as stearic acid. Markers of cardiovascular risk, such as activation of platelet aggregation factors or C-reactive protein levels, are not modified by diets providing stearic acid, as occurs with other saturated fatty acids. The confirmation of the “neutral” effect of stearic acid represents a perspective for the development of fats with high contents of this fatty acid to replace hydrogenated fats containing trans isomers. The present review discusses these aspects. KEY-WORDS: Alternatives to hydrogenated fats – Cardiovascular health – “Neutral” metabolic effect – Stearic acid – trans Isomers.

1.  INTRODUCTION The dietary consumption of fatty acids with trans isomerism (TFA) has been called into question by health and food authorities due to their adverse effects on overall health and mainly on cardiovascular health derived from their consumption (Ascherio et al., 1996; Hu et al., 1997; Hunter, 2006; Mozaffarian et al., 2009). Several epidemiological and clinical studies have unmistakably demonstrated that TFA acids produce an increase in plasmatic total cholesterol levels, in LDL-cholesterol levels (bad cholesterol) and in total triglycerides (Kris-Etherton et al., 2005; Mozaffarian et al., 2006; Hunter, 2006). 131

a. valenzuela, b. delplanque and m. tavella

It has been also demonstrated that TFA reduce HDL-cholesterol levels (good cholesterol) (Hu et al., 1997). There is also evidence that TFA can increase type 2 diabetes complications (Salmeron et al., 2001; Tanasescu et al., 2004; Saravanan et al., 2005). These effects have a negative incidence on health, for they increase in the risk of cardiovascular diseases, which are the main reason for death in the population of the western world. Nowadays, it is considered that the TFA negative effect on health can be at least compared with the effect of saturated fatty acids (SAFA) (Sundran et al., 1997; Hu et al., 1997; Valenzuela & Morgado., 1999; Judd et al., 2002; Hunter, 2006). According to epidemiological studies and in order to avoid a possible incidence of cardiovascular risk, health authorities have proposed to limit TFA consumption to 2% of the total energy supply (US FDA, 2003; Uauy et al., 2009). Different countries have adopted policies aiming at a drastic reduction in TFA consumption in their populations (Stender et al., 2006). Since June 1st 2003, Denmark has forbidden any content higher than 2% of TFA in both locally produced and imported fats hence the use of partially hydrogenated fats has been essentially eliminated from this country (Leth et al., 2005). Countries such as Norway, Finland, and the Netherlands have similar initiatives (Aro, 2005). Since 2003, Canada has become the first country in America to regulate the compulsory labelling of TFA and in 2006 it was proposed that TFA should not exceed 2% of the total fat content in vegetable oils and soft margarines for spread, and 5% of the total fat content in other foods. The United States introduced the compulsory labelling of TFA in 2006 (Eckel et al., 2007), an initiative that was followed by many other countries in Latin America, mainly MERCOSUR countries (Argentine, Brazil, Paraguay and Uruguay), and Chile (Valenzuela, 2008). Important fast food chains have withdrawn TFA from their products in different Latin American countries (Argentina, Brazil, Chile, Uruguay) (Valenzuela, 2008a). Most recently, the state of New York, in the United States, banned the use of hydrogenated products containing TFA in fast food restaurants. In response to these requirements, different industrial and fast food companies have announced the removal or future elimination of fats containing TFA in their products (Korver & Katan, 2006; Mozaffarian & Clarke, 2009). In recent years, The World Health Organization (WHO) has held three meetings of Scientific Update on the elimination of TFA: PAHO (Pan American Health Organization) meeting, Washington DC, USA, August 2007; PAHO meeting, Washington, DC, USA, November 2007; and WHO meeting, in Geneva, Italy, February 2008. Their findings indicate that the replacement of TFA in partially hydrogenated vegetable oil with alternative fats and oils would substantially lower cardiovascular disease risk through multiple mechanisms beyond those on cholesterol-lipoprotein fractions (Kris132

grasas y aceites,

Etherton et al., 2005), thus explaining in part the difference derived from estimates based on controlled dietary interventions focusing mainly on serum cholesterol fractions versus prospective cohort studies having cardiovascular disease events as their main outcome (Hunter et al., 2010). It is important to highlight that in Europe, the starting point was in the early 1990s, when the TFA reduction in certain margarines was implemented by means of suppressing partially hydrogenated fats (Morin, 2007). This situation has been envied by Americans and in 1994 Harvard researchers, lead by epidemiologist Walter Willett, encouraged people to join the European initiative (Ascherio et al., 1996). Willett´s group urged Americans to stop eating TFA and encouraged the Food and Drug Administration (FDA) to add them to food labels, a step the agency considered (Ascherio et al., 1996). The same researchers also urged companies to follow Europe’s lead in improving hydrogenation and producing margarine without TFA (Hu et al., 1997). Most recent food surveys pointed out that TFA consumption had effectively decreased in several European countries mainly due to the reformulation of several food products, for example spreads (Korver & Katan, 2006). 2. TFA ISOMERS ORIGIN AND CONSUMPTION TFA isomers have two main origins: biological and technological (Valenzuela & Morgado, 1999). Biological ones come from products derived from ruminant animals (beef, beef tallow, milk and its derived products), and they do not involve more than 5-10% of the total consumption of TFA isomers in European and American countries (Larqué et al., 2001). Therefore, in these countries TFA intake comes essentially from technological sources, (9095%) mainly hydrogenated fats, frying processes and to a lesser degree from edible oils which are treated with a deodorization process (Larqué et al., 2001; Craig-Schmidt, 2006). Current guidelines recommend an intake lower than 1% of the energy as TFA (Eckel et al., 2007). In the United States current consumption is around 2.5-3% of the total energy, meaning 5.8-6.0g/day (Allison et al., 1999), although it might reach 10g/day or more in some segments of the population. Consumption in Latin America varies remarkably from country to country, but the average is around 4.5-5.0 g/day (Valenzuela, 2008a). Consumption in European countries varies depending on the country as well, but it is generally lower than in American countries, ranging from 1.4 to 5.0g/day in a remarkable decreasing rate from North to South (higher in the North than in the South, eg: Netherlands vs Spain) (Kromhout et al., 1995; Hutshof et al., 1999). The TFA source is completely different in Europe. A great deal of TFA fat has animal origin (40 to 60% vs. 5 to 10% in the US), also with a significant decreasing rate from North to South. Animal products are the main

62 (2),

abril-junio,

131-138, 2011,

issn:

0017-3495,

doi:

10.3989/gya.033910

stearic acid: a possible substitute for trans fatty acids from industrial origin

source with 60%; dairy products around 50% (butter 35% and cheese 17%) and ruminants beef 10%. Biscuits, pastry, industrial bakery and cooked dishes come next with 30-40% (Husthof et al., 1999). An important issue to highlight is that the information gathered from countries that recently joined the European Union shows that Eastern European countries seem to have a much higher consumption than in Western Europe. From the food industry point of view, it is very difficult to reduce the use of hydrogenated fats, the main source of TFA isomers, since these fats are essential for the manufacturing of several food products. Hydrogenated fats work as a base for adding other nutrients, they have thermal stability, they provide palatability and crispy characteristics to products, etc (Valenzuela & Morgado, 1999; Korver & Katan, 2006). In this way, it is currently a challenge for the food industry to substitute TFA present in their products without altering organoleptic characteristics such as appearance and stability. Within the few available alternatives to this possible replacement, the ones most commonly found are mainly oils with low polyunsaturated fatty acid contents, or with high SAFA contents (palm oil, high oleic sunflower oil, low linoleic soybean oil, etc.) (Tarragó-Trani et al., 2006). However, nowadays, stearic acid (C18:0) (STA) turns out to be a recent alternative with great significance (Hunter et al., 2010). How could an SAFA replace TFA isomers? The purpose of this review is to analyze the evidence of the neutral effects of STA in lipid and vascular parameters which constitute the markers of cardiovascular disease risk, and to state the reason why this fatty acid contained in fats can constitute a reasonable alternative for the substitution of TFA in our diet.

3.  STA, A “DIFFERENT” FATTY ACID STA is a saturated fatty acid present in fats of both animal and vegetal origin. Following palmitic acid (C16:0), it is the most widely consumed fatty acid in the United States as well as in the Western population in general (Ervin et al., 2004). Palmitic acid (16:0) amounts to 56% of the total consumption and stearic acid 26%, approximately. The rest of the SFA consumption consists of myristic (C14:0), and lauric acid (C12:0), and in lower quantities butyric (C4:0), capric (C10:0), caproic (C6:0), and caprylic acid (C8:0). STA is taken from animal fat (bovine, porcine, ovine and marine fish), and in lower amounts from vegetable fats (coconut oil, soybean oil, corn oil, cocoa butter, etc). Figure 1 shows the STA contents in different fats of regular consumption. Fatty acid absorption into the human digestive system essentially depends on the position that fatty acids have in dietary triglycerides (Mu & Hø´y, 2004), which amount to 90-95% of our fat consumption. The rest of our intake consists of phospholipids (3.5-4%), and different sterols (1.5-2%), such as cholesterol, phytosterols, phytostanols, etc. (Carroll, 1958). In animal fats, SAFAs usually occupy sn-1 and sn-3 positions of triglycerides, and in a lower proportion they are found in the sn-2 position (Bracco, 1994). In bovine, porcine and ovine fats, STA is much more frequently found in the sn-1 and sn-3, than in the sn-2 position. In vegetable fats, SAFAs, especially STAs, mainly occupy the sn-1 and sn-3 positions of triglycerides as well (Mattson & Volpenhein, 1964). The sn-2 Position of these fats and oils is frequently occupied by unsaturated fatty acids, such as oleic acid (C18:1) and less frequently by linoleic (C18:2),

Figure 1 Common fatty acids in edible fats and oils. MUFA: monounsaturated fatty acids. Modified from Kris-Etherton et al., 2005.



grasas y aceites,

62 (2),

abril-junio,

131-138, 2011,

issn:

0017-3495,

doi:

10.3989/gya.033910

133

a. valenzuela, b. delplanque and m. tavella

and alpha linolenic (C18:3) acids (Christie & Moore, 1972). This special stereoisomerism has an important impact on the absorption degree of each fatty acid in the human small intestine (Kubow, 1996; Hunter, 2001). Human digestive lipases, both lingual, gastric, as well as pancreatic, have special specificity to hydrolyze the sn-1 and sn-3 positions of dietary triglycerides (Bracco, 1994; Hunter, 2001), in such a way that less than 20% of the triglycerides are totally hydrolyzed into free fatty acids and glycerol (Mu & Hø´y, 2004). In this way, the main products of triglyceride intestinal hydrolysis are monoglycerides with sn-2 sterified fatty acids, and fatty acids released from the sn-1 and sn-3 positions. This is how STA, released from both animal and vegetal fats, will be present in the small intestine lumen, after the digestive process, primarily in a free form and secondarily as part of sn-2 monoglycerides (Kritchevsky, 1994; Hunter, 2001). When STA is located in the sn-2 position in the triglycerides it is well absorbed, but when found in the sn-1 or sn-3 positions it is only partially absorbed, ranging from 37 to 55% (Mattson et al., 1979). 4.  STA Digestion and Absorption Studies have demonstrated that, comparatively, STA is absorbed in a lower proportion than other SAFAs such as lauric, myristic and palmitic, and even MUFA, as oleic, or type C18:1 TFA isomers (Baer, 2003). The general conclusion is that STA is less absorbed than other dietary SAFAs, therefore its plasmatic concentration would be lower when compared with the ones obtained with similar amounts of other SAFAs (Kris-Etherton et al., 1997; Kris-Etherton et al., 2005). The reason for the reduction in the presence of STA at plasmatic level, compared with equivalent amounts of other SAFAs, is not yet fully known (German & Dillard, 2004). It has been postulated that due to its relatively high melting point (70°C), when released into the intestinal lumen under the effect of enzymatic hydrolysis, it would form calcium and/or magnesium insoluble salts, which would be eliminated through depositions (Mattson et al., 1979). Other researchers have also postulated that a certain percentage of absorbed STA would be turned into oleic acid through intestinal cell desaturation (Garg, 1992). This transformation has been estimated to be around 9% and 14% (Rhee et al., 1997). Whatever the reason might be, the point is that STA absorption is lower than that of SAFA with fewer carbon atoms (Kris-Etherton et al., 2005), particularly when found in sn-1 and sn-3 positions. Nevertheless, the lower absorption is not exclusive when STA takes up these triglyceride positions. SALATRIM is a structured lipid which contains long chain SAFAs, mainly STA, in sn-1 and sn-3 positions, and short chain fatty acids (acetic, propionic and/or butyric) in the sn-2 position. For this reason, SALATRIM is a proved 134

grasas y aceites,

low digestibility fat and its consumption produces the elimination of high quantities of STA through depositions (Finley et al., 1994). This characteristic has made it a popular low calorie fat. However, when SALATRIM is previously randomized through interesterification, the molecule loses symmetry and thus leaves a significant proportion of STA in the s-2 position. When the resulting products effect is tested, the fatty acid also shows low absorption (Berry & Sanders, 2005). Hence, it is suggested that regardless of the position STA occupies in triglycerides (sn-1, sn-2 or sn-3), its intestinal absorption is always low (Berry & Sanders, 2005; Berry et al., 2007). It has been proposed that STA would be a bad substrate for acyl-cholesterol-acyl-transferase enzyme (ACAT) which is responsible for cholesterol re-esterification in the intestine (Daumerie et al., 1992). If cholesterol is not re-esterified into intestine cells, it is again transported to the intestinal lumen and eliminated with faeces (Rodríguez-Meléndez et al., 2006). It has also been proposed that STA would enhance cholesterol hepatic excretion through bile (Imaizumi et al., 1993), although this increase does not occur experimentally in hamsters (Hassel et al., 1997). Moreover, it has been demonstrated that STA inhibits the expression of the Nieman-Pick C1 L1 transporter (NPC1L1) in the FH 74 cell line of intestinal cells (Hunter 2001). This transporter is responsible for carrying cholesterol from intestinal lumen to the enterocyte (Rodríguez-Meléndez et al., 2006). As a result of both mechanisms, lower cholesterol absorption at the intestinal level would be produced due to the effect of STA. 4.1.  Effects on plasmatic lipids Predictive equations for the differential effects of fatty acids on plasmatic lipids developed independently by Keys et al., (1965) and Hegsted et al., (1965) several decades ago, demonstrated the hypercholesterolemic effect of SAFA and its consequent increase in cardiovascular risk. However, when these equations were applied to STA, the fatty acid appeared with a “neutral” effect on plasmatic cholesterol levels. More complex mathematical equations, like the one developed by Mensink y Katan (1992), demonstrated the neutral effect of the fatty acid, not only on total cholesterol, but also on LDL- and HDL- cholesterol levels. Similar results were obtained by Yu et al. (1995), who developed new predictive formulae starting from regression studies, concluding that STA has a neutral effect on the lipid profile in men as well as in women. Moreover, when comparing its effect with lauric, palmitic or myristic acid, STA shows a neutral or slightly positive effect on plasmatic lipids (Hunter, 2001). A meta-analysis carried out by Mensink et al. (2003) on 60 controlled clinical trials compared the effect of SAFA on the modification of plasmatic lipids with respect to their substitution by carbohydrates in the diet. It was demonstrated that STA reduces cholesterol plasmatic levels and

62 (2),

abril-junio,

131-138, 2011,

issn:

0017-3495,

doi:

10.3989/gya.033910

stearic acid: a possible substitute for trans fatty acids from industrial origin

the relationship between total cholesterol and HDLcholesterol. The modification of both parameters is a positive marker for reduced cardiovascular risk. Similar studies have demonstrated that dietary STA, contributing from 9% to 40% of the total energy, does not modify total plasmatic cholesterol, and even in some cases it is reduced (Aro et al., 1997; Nestel et al., 1998; Snook et al., 1999). A more recent study by Thijssen & Mensink (2005), showed that diets with 7% of STA produce changes in the lipid profile similar to those yielded by a diet having an equal amount of oleic or linoleic acid. In this work, a detailed evaluation on the lipoprotein profile was carried out by Nuclear Magnetic Resonance (NMR), showing that the three fatty acids have similar effects on total and LDLcholesterol reduction. Surprisingly for researchers, STA also increased HDL-cholesterol, an effect which has been previously observed only with oleic and linoleic acids. Another remarkable observation on Thijssen & Mensink’s work (2005) is that STA does not modify LDL size, that is, it does not increase smaller sized LDLs, which are considered more atherogenic than larger sized LDLs (Gardner et al., 1996). The experimental protocol used by Thijssen & Mensink involved fats only from natural origin, where STA mainly occupies sn-1 and sn3 positions. A more recent work by Sundran et al (2007), demonstrates that an interesterified fat, with a high proportion of STA (40%), negatively modifies the lipid profile, also increasing the plasmatic glucose levels. However, an analysis on the fatty acid triglyceride sterochemistry, shows that 15% of STA is in the sn-2 position. This result emphasizes the concept that the beneficial effects derived from STA are obtained when it takes up sn-1 and sn-3 of triglyceride positions. An equal effect to other saturated fatty acids is observed when it is located in the sn-2 position (Sundran et al., 2007). Apolipoproteins make up the protein part of lipoproteins and enable the selective recognition of these proteins by different tissues. B-100 apolipoprotein (ApoB-100) is only present in VLDL and LDL which derive from VLDL. In this way, high levels of ApoB-100 are indicative of an active triglyceride (by VLDL) and cholesterol (by LDL) transport. On the contrary, low ApoB-100 plasmatic levels mean low VLDL and LDL concentrations. Two studies demonstrated that when STA provides either 9.3% or 36% of the energy, a reduction in ApoB-100 from 10% to 18% is produced (Tholstrup et al., 1994; Aro et al., 1997). However, the latter study also demonstrated that STA increases lipoprotein (a) plasmatic levels. This type of lipoprotein has similar characteristics to LDL, which is related to a higher atherogenicity and considered an emerging marker of cardiovascular disease (Clevidence et al., 1997). This effect has also been shown by a study on fasting (Tholstrup et al., 1995), and in post-prandial conditions (Tholstrup & Samman, 2004). The available data concerning the changes in lipoprotein (a) concentration with dietary TFA are limited, and further studies concerning a

grasas y aceites,

62 (2),

abril-junio,

131-138, 2011,

issn:

0017-3495,

doi:

relationship between TFA and lipoprotein (a) are needed. 4.2. STA, activation of thrombogenic factors and blood pressure The activation of thrombogenic factors due to post-prandial lipemia effect has been associated with an increase in cardiovascular risk. A meal rich in STA increases lipemia, but to lower levels than lipemia resulting from taking oleic and elaidic acids (Sanders et al., 2000), or palmitic acid (Mennen et al., 1998), probably due to the low absorption effect of STA already discussed. The increase in postprandial lipemia raised the concentration of Factor VIIc activated form, a coagulation factor dependent on vitamin K. Fatty acids in general increase the activation of this Factor (Mitropoulos et al., 1994; Mennen et al., 1998). However, comparatively, STA produces a lower activation of this factor than oleic acid, as observed by Tholstrup et al. (1994) and later confirmed by Sanders et al. (2000); also lower than palmitic acid (Mennen et al., 1998). The effect of STA on hemodynamics is still not clear and in some cases is controversial. Multiple intervention studies have demonstrated an inverse correlation between cholesterol plasmatic levels and STA, and the diastolic pressure measured in middle-aged individuals with high cardiovascular disease risk (Simon et al., 1996). However, when this measurement is made on healthy individuals, both plasmatic cholesterol as well as STA levels significantly increases left ventricular diastolic pressure (Steer et al., 2002). Controlled clinical trials demonstrate that diets with 8 to 13% of the total energy as STA, have no effect on vascular pressure measured in diabetic patients (Storm et al., 1997), or in men and women with vascular pressure at normal ranges (Zock et al., 1993). This information is important because it corresponds to studies with standard STA intakes. Finally, there is little evidence on the effect of STA on molecular markers of inflammation, such as cytokines (interleukine-6, for example), adhesion molecules (such as selectins), or acute-phase expression proteins (such as C-reactive protein), which are important to predict cardiovascular disease (Blake & Ridker, 2002). STA does not modify plasmatic levels of these molecular markers if the diet imparts 11% of its energy as STA (Baer et al., 2004), although it does increase fibrinogen levels when taking in 11% of the energy as STA (Baer et al, 2004). Nevertheless, Kris-Eherton et al (2005) do not consider this last point relevant due to the fact that the average consumption STA is lower than 3% of the energy (Ervin et al., 2004). Thijssen et al. (2005) demonstrated that diets with 7% of the energy as STA, which is significantly higher than average consumption (3%), do not affect blood platelet aggregation. Therefore, researchers conclude that STA would not have thrombogenic effects, making it comparable to oleic or linoleic acid.

10.3989/gya.033910

135

a. valenzuela, b. delplanque and m. tavella

5.  FINAL CONSIDERATIONS There is a clear need to alert oil seed producers that there will likely be a requirement for an increased supply of substitute oils in order to replace TFA and that this represents an opportunity to expand or develop new oil seed varieties. The results of scientific updates should provide the evidence and scientific bases to promote discussions between the international scientific community related to nutrition and health as well as to agriculturalists and the food production industry, relevant health professionals, national and international food regulatory agencies, civil society, and the private sector in order to achieve this goal. Available information, derived from experimental, clinical and epidemiological research, enables us to confirm with reasonable evidence, that when STA is consumed in less than 7% of the total energy, lipid profile, thrombotic factors, hemodynamic and cardiovascular risk molecular markers are not modified (Kris-Etherton et al., 2005; Hunter et al., 2010). This aspect distinguishes STA from other SAFAs present in diets, such as palmitic, lauric and myristic, while it ranks among MUFAs, such as oleic acid, or polyunsaturated acids, such as linoleic acid. The reason for this “neutral” effect is still unclear, and could be assigned to different factors: a lower absorption and higher excretion in the intestinal lumen when located in the sn-1 and sn-3 positions of dietary triglycerides; a partial conversion into oleic acid through intestine cell desaturation; to an inhibitory effect on ApoB-100 synthesis; or to other factors not yet identified. Everything focuses on the fact that fats with a high proportion of STA in the sn-1 and sn-3 positions could be very good substitutes for hydrogenated fats with high contents of TFA isomers. It should be pointed out that the neutral effect on cardiovascular diseases attributed to chocolate consumption would be due, in some degree, to a high proportion of STA in the sn-1 and sn-3 positions found in cocoa butter, together with its high cytoprotective flavonoids content (Ding et al., 2006). Oils with high STA content in the sn1 and sn-3 triglyceride position for frying use, are high temperature resistant due to their saturated characteristics. These could be an adequate substitute for partially hydrogenated fats with high contents of TFA isomers, which are nowadays used in the food industry and mainly for frying processes in fast food chains (DiRienzo et al., 2008). In summary, we conclude that STA may be a reasonable substitute for TFA and also for cholesterol-raising SAFAs for solid fat applications, such as baked goods, shortenings, spreads, and margarines. REFERENCES Allison DB, Egan SK, Barraj LM, Caughman C, Infante M, Heimbach JT. 1999. Estimated intakes of trans fatty acids and other fatty acids in the US population. J. Am. Diet. Assoc. 99, 166-174.

136

grasas y aceites,

Aro A, Jauhiainen M, Partenen R, Salminen L, Mutanen M. 1997. Stearic acid, trans fatty acids, and dairy fat: effects on serum and lipoprotein lipids, apolipoproteins, lipoprotein(a), and lipid transfer protein in healthy subjects. Am. J. Clin. Nutr. 65, 1419-1426. Aro A. 2005. The scientific basis of TFA regulationsis it sufficient? A personal view. First International Symposium on Trans fatty Acids and Health. Rungstedgaard, Denmark, September 11-13, Abstract. Ascherio A, Rimm EB, Giovannucci EL, Spiegelman D, Stampfer M, Willett WC. 1996. Dietary fats and risk of coronary heart disease in men: cohort follow up study in the United States. Brit. Med. J. 313, 84-90. Baer DJ, Judd JT, Kris-Etherton PM, Zhao G, Emken EA. 2003. Stearic acid absorption and its metabolizable energy value are minimally lower than those of other fatty acids in healthy men fed mixed diets. J. Nutr. 133, 4129-4134. Baer DJ, Judd J, Clevidence B, Tracy R. 2004. Dietary fatty acids affect plasma markers of inflammation in healthy men fed controlled diets: a randomized crossover study. Am. J. Clin. Nutr. 79, 969-973. Berry SE, Sanders TA. 2005. Influence of triacylglyceride structure of stearic acid-rich fats on postprandial lipaemia. Proc. Nutr. Soc. 64, 205-212. Berry SE, Miller GJ, Sanders TA. 2007. The solid fat content of stearic acid-rich fats determines their postprandial effects. Am. J. Clin. Nutr. 85, 1486-1494. Blake GJ, Ridker PM. 2002. Inflammatory biomarkers and cardiovascular risk prediction. J. Intern. Med. 252, 283-294. Bracco U. 1994. Effect of triglyceride structure on fat absorption. Am. J. Clin. Nutr. 60, 1002S-1009S. Carroll KK. 1958. Digestibility of individual fatty acids in the rat. J. Nutr. 64, 399-410. Christie WW, Moore JH. 1972. The structure of adipose tissue and heart muscle triglycerides in the domestic chicken (Gallus gallus). J. Sci. Food Agric. 23, 73-77. Clevidence BA, Judd JT, Schaefer EJ. 1997. Plasma lipoprotein (a) levels in men and women consuming diets enriched in saturated cis or transmonounsaturated fatty acids. Arterioescler. Thromb. Vasc. Biol. 17, 1657-1661. Craig-Schmidt M. 2006. World-wide Consumption of trans fatty acids. Atherosclerosis (suppl) 7, 1-4. Daumerie CM, Woollett LA, Dietschy JM. 1992. Fatty acids regulate hepatic low density lipoprotein receptor activity through redistribution of intracellular cholesterol pool. Proc. Natl. Acad. Sci. USA. 89, 10797-10801. Ding E, Hutfless S, Ding X, Girotra S. 2006. Chocolate and prevention of cardiovascular disease: a systematic review. Nutrition & Metabolism, 3, 2-14. Di Rienzo MA, Lemke SL, Petersen BJ, Smith KM. 2008. Effect of substitution of high stearic acid low linolenic acid soybean oil for hydrogenated soybean oil on fatty acid intake. Lipids 43, 451-456. Eckel RH, Borra S, Lichtenstein AH, Yin-Piazza SY. 2007. Understanding the complexity of trans fatty acid reduction in the American diet. Circulation 115, 2231-2246. Ervin RB, Wright JD, Wang CY, Kennedy-Stephenson J. 2004. Dietary intake of fats and fatty acids for the United States population: 1999-2000. Adv. Data (November 8), 1-6. Finley J, Klemann L, Levielle G, Otterburn M, Walchak C. 1994. Caloric availability of SALATRIM in rats and humans. J. Agric. Food Chem. 42, 495-499.

62 (2),

abril-junio,

131-138, 2011,

issn:

0017-3495,

doi:

10.3989/gya.033910

stearic acid: a possible substitute for trans fatty acids from industrial origin

Gardner CD, Fortmann SP, Krauss RM. 1996. Association of small low density lipoproteins particles with the incidence of coronary artery disease in men and women. Jama 276, 875-881. Garg ML. 1992. Stearic Acid Desaturation and incorporation into murine peritoneal macrophage lipids. J. Clin. Biochem. Nutr. 13, 169-178. German JB, Dillard CJ. 2004. Saturated fats: what dietary intake?. Am. J. Clin. Nutr. 80, 550-559. Hassel CA, Mensing E, Gallaher D. 1997. Dietary stearic acid reduces plasma and hepatic cholesterol concentration without increasing bile acid excretion in cholesterol-fed hamsters. J. Nutr. 127, 1148-1155. Hegsted DM, McGandy RB, Myers ML, Stare FJ. 1965. Quantitative effects of dietary fat on serum cholesterol in man. Am. J. Clin. Nutr. 17, 281-295. Hu F, Stampfer M, Manson J, Rimm E, Colditz G, Rosner B, Hennekens C, Willett W. 1997. Dietary fat intake and the risk of coronary heart disease in women. N. Engl. J. Med. 337, 1491-1499. Hunter JE. 2001. Studies on effects of dietary fatty acids as related to their position on triglycerides. Lipids 36, 655-668. Hunter JE. 2006. Dietary trans fatty acids: a review of recent human studies and food industry responses. Lipids 41, 967-992. Hunter JE, Zhang J, Kris-Etherton P. 2010. Cardiovascular disease risk of dietary stearic acid compared with trans, other saturated, and unsaturated fatty acids: a systemic review. Am. J. Clin. Nutr. 91, 46-63. Hutshof K, Van Erp-Baart MA, Anttolainen M. 1999. Intake of fatty acids in western Europe with emphasis on trans fatty acids: the TRANSFAIR study. Eur. J. Clin. Nutr. 53, 143-57. Imaizumi K, Abe K, Kuroiwa C, Sugano M. 1993. Fat containing stearic acid increases fecal neutral steroid excretion and catabolism without affecting plasma cholesterol concentration in hamsters fed a cholesterol-containing diet. J. Nutr. 123, 1683-1702. Judd JT, Clevidence DJ, Kris-Etherton P, Muesing R A, Iwane N. 2002. Dietary cis and trans monounsaturated and saturated fatty acids and plasma lipids and lipoprotein in men. Lipids, 37, 123-131. Keys A, Anderson JT, Grande F. 1965. Serum cholesterol response to changes in the diet IV. Particular saturated fatty acids in the diet. Metabolism 14, 776-787. Korver O, Katan M. 2006. The elimination of trans fats from spreads: how science helped to turn an industry around. Nutr. Rev. 64, 275-279. Kris-Etherton P, Yu S. 1997. Individual fatty acid effects on plasma lipids and lipoproteins: human studies. Am. J. Clin. Nutr. 65, 1628S-1644S. Kris-Etherton PM, Griel AE, Psota TL, Gebauer S K, Zhang J, Etherton TD. 2005. Dietary stearic acid and risk of cardiovascular disease: intake, sources, digestion, and absorption. Lipids, 40, 1193-1200. Kritchevsky D. 1994. Stearic acid and atherogenesis: history. Am. J. Clin. Nutr. 60, 997S-1001S. Kromhout D, Menotti A, Bloemberg B. 1995. Dietary saturated and trans fatty acids and cholesterol and 25-year mortality from coronary heart disease: The Seven Countries Study. Prev. Med. 24, 308-315. Kubow S. 1996. The influence of positional distribution of fatty acids in native, interesterified and structurespecific lipids on lipoprotein metabolism and atherogenesis. J. Nutr. Biochem. 7, 530-541. Larqué E, Zamora S, Gil A. 2001. Dietary trans fatty acids in early life: a review. Early Human Develop. 65, S31-S41.



grasas y aceites,

62 (2),

abril-junio,

131-138, 2011,

issn:

0017-3495,

doi:

Leth T, Bysted A, Erendah-Mikkelsen A. 2005. The effect of the regulation on trans fatty acid content in danish food. First International Symposium on trans fatty acids and health. Rungstedgaard, Denmark, September 11-13, Abstract. Mattson FH, Volpenhein RA. 1964. The digestion and absorption of triglycerides. J. Biol. Chem. 239, 27722777. Mattson FH, Nolen GA, Webb MR. 1979. The absorbability by rats of various triglycerides of stearic and oleic acid and the effect of dietary calcium and magnesium. J. Nutr. 109, 1682-1687. Mennen L, de Maat M, Meijer G, Zock P, Grobbee D, Kok F, Kluft C, Schouten E. 1998. Factor VIIa response to a fat-rich meal does not depend on fatty acid composition: a randomized trial. Arterioscler. Thromb. Vasc. Biol. 18, 599-603. Mensink RP, Katan MB. 1992. Effect of dietary fatty acids on serum lipids and lipoproteins. A Meta-analysis of 27 Trials. Arterioscler. Thromb. 12, 911-919. Mensink RP, Zock PL, Kester AD, Katan MB. 2003. Effects of dietary fatty acids and carbohydrates on the ratio of serum total to HDL-cholesterol and on serum lipids and apolipoproteins: a meta-analysis of 60 controlled trials. Am. J. Clin. Nutr. 77, 1146-1155. Mitropoulos K, Miller G, Martin J, Reeves B, Cooper J. 1994. Dietary fat induces changes in factor VII coagulant activity through effects on plasma free stearic acid concentrations. Atheroscler. Thromb. 14, 214-222. Morin O. 2007. Huiles végétales et margarines : évolution de la qualité - Les solutions technologiques à la réduction des acides gras trans. Cah. Nutr. Diét., 42, 5, A paraître. Mozaffarian D, Katan M, Ascherio A, Stampfer M, Willett W. 2006. Trans fatty acids and cardiovascular disease. New Engl. J. Med. 354, 1601-1613. Mozaffarian D, Aro A, Willett W. 2009. Health effects of trans-fatty acids: experimental and observational evidence. Europ. J. Clin. Nutr. 63, S5-S21. Mozaffarian D, Clarke R. 2009. Quantitative effects on cardiovascular risk factors and coronary heart disease risk of replacing partially hydrogenated vegetable oils with other fats and oils. Eur. J. Clin. Nutr. 63, S22-S33. Mu H, Hø´y C. 2004. The digestion of dietary triacylglycerols. Prog. Lipid. Res. 43, 105-133. Nestel PJ, Pomeroy S, Kay S, Sasahara T, Yamashita T. 1998. Effect of a stearic acid-rich, structured triacylglycerol on plasma lipids concentrations. Am. J. Clin. Nutr. 68, 1196-1201. Rhee SK, Adlof RO, Ciszek A, Brenna JT. 1997. Desaturation and interconversion of dietary stearic and palmitic acid in human plasma and lipoproteins. Am. J. Clin. Nutr. 65, 451-458. Rodríguez-Meléndez R, Rasmussen HE, Lee JY, Carr TP. 2006. NPC1L1 gene expression is downregulated by stearic acid in CCL-241 cells. Faseb J. 20, 138-145. Sanders T, de Grassi T, Miller G, Morrissey J. 2000. Influence of fatty acid chain-lenght and cis/trans isomerization on postprandial lipemia and Factor VII in healthy subjects. Atherosclerosis 149, 413-420. Salmeron J, Hu FB, Manson JE, Stampfer MJ, Colditz GA, Rimm E B, Willett W. 2001. Dietary fat intake and risk of type 2 diabetes in women. Am. J. Clin. Nutr. 73, 1019-1026. Saravanan N, Haseeb A, Ehteham NZ, Ghafoorunissa X. 2005. Differential effects of dietary saturated and

10.3989/gya.033910

137

a. valenzuela, b. delplanque and m. tavella

trans fatty acids on expression of genes associated with insulin sensitivity in rat adipose tissue. Eur. J. Endocrinol. 153, 159-165. Simon J, Fong J, Bernert J. 1996. Serum fatty acids and blood pressure. Hypertension 27, 303-307. Snook J-T, Park S, Williams G, Tsai YH, Lee, N. 1999. Effect of synthetic triglycerides of myristic, palmitic, and stearic acid on serum lipoprotein metabolism. Eur. J. Clin. Nutr. 53, 597-605. Steer P, Millgard J, Sarabi D, Basu S, Vessby B, Kahan T, Edner M, Lind L. 2002. Cardiac and vascular structure and function are related to lipid peroxidation and metabolism. Lipids 37, 231-236. Stender S, Dyerberg J, Astrup A. 2006. High levels of trans fat in popular fast foods. N. Engl. J. Med. 354, 1650-1652. Storm H, Thomsen C, Pedersen E, Rassmussen O, Christiansen C, Hermansen K. 1997. Comparison of a carbohydrate-rich diet and diets rich in stearic or palmitic acid in NIDDM patients. Effects on lipids, glycemic control, and diurnal blood pressure. Diabetes Care 20, 1807-1813. Sundran K, Ismail A, Hayes K, Jeyemalar R, Pathmanathan R. 1997. Trans (elaidic) fatty acids adversely affect the lipoprotein profile relative to specific saturated fatty acids in humans. J. Nutr. 127, 514S-520S. Sundran K, Karupaih T, Hayes K. 2007. Stearic acid-rich interesterified fat and trans-rich fat raise the LDL/HDL ratio and plasma glucose relative to palm olein in humans. Nutrition & Metabolism 4, 3-15. Tanasescu M, Cho E, Manson JE, Hu FB. 2004. Dietary fat and colesterol and the risk of cardiovascular disease among women with type 2 diabetes. Am. J. Clin. Nutr. 79, 999-1005. Tarragó-Trani T, Phillips K, Lemar L, Holden J. 2006. New existing oils and fats used in products with reduced trans-fatty acid content. J. Am. Diet Assoc. 106, 867-880. Thijssen MA, Mensink R P. 2005. Small differences in the effects of stearic acid, oleic acid, and linoleic acid on the serum lipoprotein profile of humans. Am. J. Clin. Nutr. 82, 510-516. Thijssen MA, Hornstra G, Mensink R. 2005. Stearic, oleic, and linoleic acids have comparable effects on markers of trombotic tendency in healthy human subjects. J. Nutr. 135, 2805-2811.

138

grasas y aceites,

Tholstrup T, Marckmann P, Jespersen J, Sandtrom B. 1994. Fat high in stearic acid favorably affects blood lipids and Factor II coagulant activity in comparison with fats high in palmitic acid or high in myristic and lauric acids. Am. J. Clin. Nutr. 59, 371-377. Tholstrup T, Marckmann P, Vessby B, Sandstrom B. 1995. Effects of fats high in individual saturated fatty acids on plasma lipoprotein(a) levels in young healthy men. J. Lipid Res. 36, 1447-1452. Tholstrup T, Samman S. 2004. Postprandial lipoprotein(a) is affected differently by specific individual dietary fatty acids in healthy young men. J. Nutr. 134, 25502555. Uauy R, Aro A, Clarke R, Ghafoorunissa X, L’Abbé M, Mozaffarian D, Skeaff CM, Stender S, Tavella A. 2009. WHO Scientific update on trans fatty acids: summary and conclusions. Eur. J. Clin. Nutr. 63, S68-S75. U.S. Food and Drug Administration and Center for Food Safety and Applied Nutrition 2003. Food Labelling: Trans fatty acids in nutrition labeling, nutrient content claims, and health claims. Federal Register 68, 41434-41506. Valenzuela A, Morgado N. 1999. Trans fatty acid isomers in human health and in the food industry. Biol. Res. 32, 273-287. Valenzuela A. 2008. Trans fatty acid consumption in Latin America. In “Healthy oils and the elimination of industrially produced trans fatty acids in the Americas” Pan American Health Organization (PAHO) Document, Washington DC, pp 15-27. Valenzuela A. 2008a. Ácidos grasos con isomería trans II. Situación de consumo en Latinoamérica y alternativas para su sustitución. Rev. Chil. Nutr. 35, 172-180. Yu S, Derr J, Etherton T, Kris-Etherton P. 1995. Plasma cholesterol-predictive equations demonstrate that stearic acid is neutral and monounsaturated fatty acids are hypocholesterolemics. Am. J. Clin. Nutr. 61, 1129-1139. Zock P, Blijlevens R, de Vries J, Katan M. 1993. Effects of stearic acid and trans fatty acids versus linoleic acid on blood pressure in normotensive women and men. Eur. J. Clin. Nutr. 47, 437-444. Recibido: 16/3/10 Aceptado: 25/5/10

62 (2),

abril-junio,

131-138, 2011,

issn:

0017-3495,

doi:

10.3989/gya.033910

grasas y aceites,

62 (2), 139-148, 2011, issn: 0017-3495 doi: 10.3989/gya.066510

abril-junio,

INVESTIGACIÓN Antioxidant and antimicrobial effects of some natural plant extracts added to lamb patties during storage By Hayam M. Ibrahim,* Azza A. Abou-Arab and Ferial M. Abu Salem Food Technology Dept., National Research Center, Dokki, Cairo, Egypt (*Corresponding author: [email protected])

RESUMEN Efecto antioxidante y antimicrobiano de algunos extractos de plantas naturales añadidos a pastel de cordero durante el almacenamiento. Las plantas naturales están consideradas como un importante producto donde buscar y encontrar nuevas fuentes de antioxidantes naturales y/o agentes antimicrobianos. La concentración óptima de algunos extractos de plantas naturales (jojoba, jatropha, ginseng y jengibre) fueron determinado y añadidas a pasteles de cordero. Algunas características químicas y microbiológicas de los pasteles preparados y almacenados durante 9 días a 4°C fueron evaluados. Tanto la adición de estos extractos como el tiempo de almacenamiento tuvieron un efecto significativo en los pasteles en el periodo de almacenamiento. La efectividad de los extractos naturales ensayados puede ser enumerada en el siguiente orden decreciente de valores de substancias reactivas con el ácido tiobarbitúrico (TBARS): ginseng . jatropha . jojoba . jengibre. El recuento de microorganismos decreció significativamente con la adición de los extractos durante el periodo de almacenamiento. Comparado con los pasteles control, la adición de estos extractos naturales fue efectiva como agente antioxidante y antimicrobiano en la mejora de las propiedades de los pasteles de cordero. PALABRAS-CLAVE: Aminas biogénicas – Antimicrobiano – Antioxidante –Extracto de plantas naturales – Oxidación lipídica – Pastel de cordero – Recuento microbiano. SUMMARY Antioxidant and antimicrobial effect of some natural plant extracts added to lamb patties during storage. Natural plants are considered an important target to investigate in order to provide a new source of natural antioxidants and/or antimicrobial agents. The optimum concentrations of some natural plant (jojoba, jatropha, ginseng and ginger) extracts were determined and added to lamb patties. Some chemical and microbial characteristics of the prepared patties during storage for 9 days at 4°C were evaluated. Both the addition of these extracts and storage time had a significant effect on the patties throughout the storage period. The effectiveness of the tested natural extracts can be listed in the following order of decreasing Thiobarbituric acid reactive substance (TBARS) values: ginseng . jatropha . jojoba . ginger. Aerobic plate count, mould and yeast counts decreased significantly with addition



of the extracts during the storage period. Also, the addition of the extracts was significantly effective in reducing histamine, tyramine and putrescine formation during the storage period. Compared to control patties, the addition of these natural extracts was effective as antioxidant and antimicrobial agents for improving the properties of lamb patties. KEY-WORDS: Antimicrobial – Antioxidant – Biogenic amines – Lamb patties – Lipid oxidation – Microbial counts – Natural plant extracts.

1.  INTRODUCTION Lipid oxidation and the growth of undesirable microorganisms in food products results in the development of spoilage, off flavor, rancidity, and deterioration, rendering such products unacceptable for human consumption (Bozin et al., 2007) and yielding many compounds that contribute to the pathogenesis of cancer, atherosclerosis, heart and allergic diseases (Mielnik et al., 2008; Tang et al., 2001). The presence of biogenic amines in food constitutes a potential public health concern due to their physiological and toxicological effects. It is important to monitor biogenic amine levels in fresh and processed food not only due to their toxicity but also because they can be a useful index of spoilage (Önal, 2007). Lipid oxidation and microbial growth in meat products may be controlled or at least minimized by using either synthetic or natural food additives commonly used in the meat industry (Gray et al., 1996; Lee et al., 1997; Mielnik et al., 2003; Sallam et al., 2004; Estevez and Cava, 2006). The natural antioxidants found in plants have gained considerable interest for their role in preventing the auto-oxidation of fats, oils and fat containing food products (Reddy et al., 2005). The antioxidant properties of herbs, spices, plant and other food extracts are apparently related to their phenolic content, suggesting that antioxidant action is similar to that of synthetic phenolic antioxidants (Lai et al., 1991). 139

h. m. ibraim, a. a. abou-arab and f. m. abu salem

Since the worldwide trend towards the use of natural additives in food (Yanishlieva et al., 2006); natural plants are considered an important target to investigate in order to provide a new source of natural antioxidants and/or antimicrobial agents from a safety view point. Consequently, there is a practical need for the screening and selection of natural antioxidants as effective alternatives in the prevention of food deterioration (Kikuzaki and Nakatani, 1993). Several plants with very high nutritive values exist and yet remain unexploited for human and animal benefits (Oladele and Oshodi, 2007). Therefore, the search for, and development of other antioxidants and antimicrobials of natural origin are highly desirable. Jatropha curcas is a nut belonging to the Euphobiaceae family. Recently, the tree of this plant has been successfully cultivated in Upper Egypt (Hawash et al., 2009) and all parts of it have specific uses as determined by Gubitz et al., (1999) and Makkar et al., (1997). El Diwani et al., (2009) reported that the residue of the methanolic extract of Egyptian jatropha curcas contains bioactive substances such phenolic compounds, which were successful as natural antioxidants against oxidative deterioration. The flavonoid profile of the fruits of the Jojoba plant, Simmondsia chinensis may place this family among other families which are rich in flavonol methyl ethers and flavonoid content which make the pericarp a valuable source for antioxidant and hepato-protective compounds (El-Halwany, 2002). This plant extract has been reported to be useful as a dietary supplement for use in a weight control regiment in humans, a component of functional food, a food additive, a medical food, or as a therapeutic agent (Teague et al., 2005). Ginseng is an herbaceous perennial belonging to the Aralia family. It is used early for medicinal purposes and used widely in herbal, health food and cosmetic applications (Rangahau, 2001). Various formulations prepared from the Panax ginseng root have been marketed as dietary supplements, especially in China where it is frequently used as a food additive and as raw materials of healthy food rather than therapeutic agents (Gillis, 1997; Shen et al., 2003). Bioactive compounds from medicinal plants including ginseng are known to protect against oxidative stress from reactive oxygen species and prevent lipid per-oxidation (Sievenpiper et al., 2003; Fuzzati, 2004). The rhizome of the popular ginger species, Zinger officinale, is widely used as a spice and food seasoning due to its sweet aroma and pungent taste. It is well known to have antioxidant activity (Jitoe et al., 1992; Zia-ur-Rehman et al., 2003,) and effective antimicrobial agents. A ginger rhizome extract exhibited the highest antioxidant activity (Mansour and Khalil. 2000) due to the effect of its total phenols (Stoilova et al., 2007). The interest in the antioxidant activity of plant extracts has become greater and very important (Joyeux et al., 1995; Azaizeh et al., 2005; Alma 140

grasas y aceites,

et al., 2003) due to the fact that free radicals e.g. reactive oxygen species (ROS) can be responsible for various diseases, e.g. heart diseases, stroke, arteriosclerosis and cancer, as well as being involved in the aging process (Willcox et al., 2004). The effects of plant extracts or essential oils classified as greatly recognized as safe (GRAS) following their addition, have been studied extensively and reported in a variety of meat types, including pork (Nissen et al., 2004); beef (Solomakos et al., 2008); lamb (Camo et al., 2008). Lamb meat contains higher levels of v´ -3 polyunsaturated fatty acids (PUFAs) compared to beef or pork, which is beneficial to human health (Wood et al., 1999); however, PUFAs increase the susceptibility of meat to oxidative processes such as lipid oxidation ultimately leading to off odors and warmed over flavor (Jeremiah, 2001). The objectives of the present study were: i) To establish the optimum concentrations of some natural plant extracts: jojoba (Jo), jatropha (Ja), ginseng (Gg) and ginger (Gr) as sources of natural antioxidants and/or antimicrobial agents to be added to lamb patties in order to diminish oxidative and microbiological deterioration. ii) To evaluate the effects of the natural extracts at the optimum concentrations on the evolution of quality parameters (thiobarbituric acid reactive substances (TBARS), biogenic amines (BA), microbiological count) in the prepared lamb patties stored at 4°C for 9 days. 2.  MATERIALS AND METHODS 2.1.  Materials Four natural plants (Jojoba Simmondsia chinensis, jatropha curcas, Panax ginseng and ginger, Zinger officinale) were used as sources of antioxidants and antimicrobial agents. Jojoba (pericarp) was purchased from the Egyptian Natural Oils Company, Cairo, Egypt. Jatropha curcas (leaves and roots) were obtained from the Ministry of Agriculture and Reclamation Land., Egypt. Jojoba and Jatropha were air-dried, powdered and kept in tightly closed amber glass containers. Ginger rhizomes (Zingiber officinal) were purchased from the local market. a ginseng extract was obtained from the Korean Society of ginseng, Seaul, Korea as a gift. Thiobarbituric acid (TBA), 1,1,3,3– tetraethoxypropan (TEP) and other chemicals used were of analytical grade and obtained from Sigma Chemical Co (St. Louis, MO). 2.2.  Preparation of the plant extracts Jojoba extract Air-dried powdered pericarp (one kg) of jojoba was exhaustively defatted using hexane and then extracted with ethanol 70% by percolation. The ethanolic extract was combined and evaporated under reduced pressure to yield 50gm of dry residue.

62 (2),

abril-junio,

139-148, 2011,

issn:

0017-3495,

doi:

10.3989/gya.066510

antioxidant and antimicrobial effects of some natural plant extracts added to lamb patties DURING STORAGE

The residue was suspended in water (250ml) and partitioned successively with chloroform (5 3 50ml) followed by ethyl acetate (5 3 50ml) and n-butanol (7 3 50ml). The solvents were evaporated under reduced pressure to give chloroform fraction (11gm), ethyl acetate fraction (4gm) and butanol fraction (10gm) (El-Halwany, 2002). Jatropha extract Ten grams of the air-dried powder of the leaves and roots of jatropha were extracted successively under shaking with chloroform (CHCl3) three to five times at room temperature, with 90% methanol (CH3OH) in a water-bath at 50ºC three to five times and finely with water in a water-bath at 70ºC. The obtained extract was filtered and evaporated using a vacuum evaporator to give the crude dried extract (Mothana and Likdequist, 2005). Ginger extract Ginger rhizomes were ground and passed through a 60 mesh screen. One hundred grams of ground ginger were defatted by shaking three times with four volumes of petroleum ether in a rotary shaker for 1 h. The residue obtained after filtration was dried overnight under a hood until all traces of petroleum ether were removed. The dried residue was extracted three times with four volumes of 90% ethanol by shaking for 1 h. and filtered. The combined filtrate was concentrated in a rotavapor and placed under a hood to remove the residual ethanol. The obtained aqueous extract was frozen overnight and freeze-dried at − 60°C (Dura-Dry, USA). The freeze-dried extract was stored in airtight containers at 5°C until use (Mansour and khalil, 2000). Ginseng extract Korean red ginseng extract was obtained from the Korean Society of ginseng, Seaul, Korea by Prof Dr. Mosaad A. Abdel-Wahhab, Food Toxicology and Contamination Dept., National Research Center, Cairo, Egypt; who supplied it for us as a gift 2.3. Determination of optimum concentration of the natural extracts Jojoba (Jo), jatropha (Ja), ginseng (Gg) and ginger (Gr) extracts were screened at levels ranging from 0 to 1% to determine their optimum working concentrations. Fresh lamb meat was obtained from a local slaughter house and kept overnight at 0ºC. The lamb meat was minced using a meat mincer. The four natural plant extracts were mixed with the minced lamb meat and formed into patties using a patty former. The potential antioxidants of the test extracts were determined through thiobarbituric acid reactive substances (TBARS), which were assessed in the lamb patties.



grasas y aceites,

62 (2),

abril-junio,

139-148, 2011,

issn:

0017-3495,

doi:

The optimum concentrations for the individual test extracts were identified during the screening trials and assessed simultaneously. 2.4.  Preparation of Lamb Patties Minced lamb meat was subdivided into five equal parts. Lamb patties were prepared to provide five treatment samples. A control treatment was formulated without plant extracts. The other treatments were prepared by adding the optimum concentrations determined of the tested extracts to lamb meat as follows: 0.1% jojoba (Jo) extract (sample patties with Jo), 0.1% jatropha (Ja) extract (sample patties with Ja), 0.25% ginseng (Gg) extract (sample patties with Gg) and 0.25% ginger (Gr) extract (sample patties with Gg); then mixed well and formed into patties (100g) using a meat former. Lamb patties were placed on plastic foam meat trays, wrapped with polyethylene film and kept in a refrigerator at 4°C for 9 days. The effect of the optimum concentration of the test extracts on thiobarbituric acid reactive substances (TBARS), pH, biogenic amines(BAs), mould and yeast counts & aerobic plate count (APC) were determined in lamb patties for 0, 3, 6, and 9 days of storage time at 4ºC. Hence, the patties quality and safety could be assessed. 2.5.  Chemical analyses pH determination A lamb patty sample (10 g) was homogenized in 100 ml distilled water for 1 min in a blender and the pH was measured using a digital pH-meter (HAANA, HI902 meter, Germany).Two readings were taken from each of the three lamb patty samples. Thiobarbituric acid reactive substances (TBARS) value The TBARS values were determined spectrophotometricaly according to Byun et al., (2001). Patty samples were analyzed for the optimum concentration of each extract. Homogenized patty samples (2g) were taken and TBARS were extracted twice with 10 ml of 0.4M perchloric acid. Extracts were collected and made up to 25ml with 0.4M perchloric acid and then centrifuged for 5 min at 1790g. After centrifugation, 1ml of extract was poured into a glass stoppered test-tube. A TBARS reagent (5ml) was added and the extract was heated in a boiling water bath for 35 min. After cooling under tap-water, the absorbance of the sample was read against the appropriate blank at 538nm. A standard curve was prepared using 1,1,3,3 - tetraethoxypropane (TEP). Biogenic amines Histamine, tyramine and putrescine were extracted as follows: five grams of the sample were blended

10.3989/gya.066510

141

h. m. ibraim, a. a. abou-arab and f. m. abu salem

with 25ml 5% trichloroacetic acid. Filtration was achieved using whatman filter paper No.1. Five ml. of the extract were transferred into a suitable culture tube with 4g NaCl and 1ml of 50%Na OH and then shaken for 2 min. Centrifugation were carried out for 5 min at 5000 xg and the upper layer was transferred to a 50ml separating funnel. 15ml of n-heptane were added to the upper layer extract and extracted 3 times with 1ml portions of 0.2 N HCl. The extracts were collected in a glass stoppered tube and evaporated to dryness using a water bath at 95ºC with the aid of a gentle current of air. This was followed by the formation of Dansylamines as described by Maijala and Eerola, (1993). Biogenic amine concentrations were determined according to Deabes, (2000) using the HPLC. The HPLC system was equipped with a (Waters 600) delivery system. The HPLC column was a reverse phase C18 Nucleosil column 250  4mm, 10µm packing, (Macherey-Naggl). The detection was performed using a U.V detector (waters 486) at 254nm wavelength, using a linear program of 25 min. periods and 1ml/min constant solvent flow rate. Data were integrated and recorded using Millennium Chromatography; Manger software 2010, (Waters, Milford MA 01757) 2.6.  Microbial determinations Aerobic plate count (APC) The aerobic plate count was determined on nutrient agar medium as recommended by the American Public Health Association for food stuff examination (APHA, 1992). Plates seeded with serial dilutions of the samples were incubated at 37˚C for 24-48 h. Mould and yeast counts Mould and yeast counts were estimated on Potato Agar according to APHA, (1992).The medium was acidified to pH 3.5 by adding a sterile 10% lactic acid solution; incubation was carried out at 25-28˚C for 72 h. 2.7.  Statistical analysis The conventional statistical methods were used to calculate means and standard deviations. All the

measurements were performed in triplicate and the data are presented as mean  SD. The effects of the addition of natural antioxidant extracts and storage time were analyzed and the obtained data were subjected to analysis of variance (ANOVA) according to PC-STAT, Version I A Copyright 1985, the University of Georgia. 3.  RESUTS AND DISCUSSION 3.1. Optimum concentration of natural plant extracts Based on preliminary trials, the optimum concentrations of the four tested natural plant (Jo, Ja, Gg, Gr) extracts were incorporated into lamb patties. Potential antioxidant properties of each of the tested natural plant extracts was determined through TBARS analysis. The concentration range employed for each test extract screened was from 0 to 1.0%. Doubling of the natural extracts addition rates was used in patty processing i.e. 0, 0.01, 0.05, 0.10, 0.25, 0.50, and 1.0% in order to have a greater effect on the assessment of the extracts’ performance. Owing to the huge amount of data generated during screening; only the optimum concentrations of each of the four tested natural plant extracts are presented in Table 1. Thus, the optimum test extract addition rates based on the identified levels of antioxidant activity were determined as: jojoba (0.1%), Jatropha (0.1%), ginseng (0.25%) and ginger (0.25%). 3.2.  pH changes The effect of the optimum concentrations of natural plant extracts under investigation on the pH values of lamb patties stored at 4°C for 9 days is shown in Table 2. At zero time the pH of the control and all tested samples had the same value (5.92). Control samples, generally, had higher pH values than the other samples throughout the storage time. The pH values of the control and tested lamb patties containing natural antioxidant extracts were significantly (pBE>PE, the rank order of TP/TF content of EAE and BE were different according to antioxidant ability. The overall results showed that the EAE and BE were richer in phenolics and flavonoids than petroleum ether extract (PE), and may represent a good source of antioxidants. KEY-WORDS: Antioxidant activity – Free radical scavenger – HPLC – Jasminum nervosum Lour – Total flavaonoid and total phenolic.

1.  INTRODUCTION Reactive oxygen species (ROS) are major free radicals generated in many redox processes, which may induce some damage to the human body. Increased production of various forms of activated oxygen species, such as oxygen radicals and nonfree radical species is considered to be the main contributor to oxidative stress, which has been linked to several diseases like atherosclerosis, Parkinson’s disease, Alzheimer’s disease, stroke, arthritis, chronic inflammatory diseases, cancers, and other degenerative diseases (Halliwell and Grootveld, 1987; McDermott, 2000). Antioxidants are vital substances which possess the ability to protect the body from damage caused by free radical induced oxidative stress (Ozsoy et al., 2008). Synthetic antioxidants, such as butylated hydroxytoluene (BHT) and butylated hydroxyanisole (BHA), are commonly used in processed foods; these compounds have been reported to cause DNA damage induction and are carcinogenic (Sasaki et al., 2002; Ku and Mun, 2007). Therefore, the search for natural antioxidants that may be used in foods to replace synthetic antioxidants is necessary. Within the antioxidant compounds, flavonoids and phenolics with a large distribution in nature have been studied (Li 149

l. huo, r. lu, p. li, y. liao, r. chen, ch. deng, ch. lu, x. wei and y. li

et al., 2009). In a study by Bushra, fruit peel extracts exhibited high TP, TF, flavonol and scavenging power (2008). It is reported that the wild edible plants in the Black Sea Region of Turkey have strong antioxidant power due to their high concentrations of total phenolic compounds, flavonoids and anthocyanins (Tevfik, 2010). Phenolics or polyphenols, including flavonoids, have received considerable attention because of their physiological functions such as antioxidant, antimutagenic and antitumor activities (Othman et al., 2007). Mohammad et al. also explicitly demonstrated that pharmacological effects may be attributed, at least in part, to the presence of phenols and flavonoids in the extracts (2010). The compounds such as quercetin, rutin, narigin, catechins, caffeic acid, gallic acid and chlorogenic acid, are very important plant constituents because of their antioxidant activities (Paganga et al., 1999), which are also used widly as standard substances today. Jasminum nervosum Lour. belonging to the Oleaceae family, is a Scandent shrub of 1-5 m height, found in the Guangdong, Hainan and Guangxi provinces of China. The leaves and the stems are widely used for various applications in traditional medicine, exhibiting a remedy for diarrhea, malaria, sores and ulearations (Editorial committee of the National Chinese Medicine Administrative Bureau. 2005). Some earlier works have been reported on the essential oils, phenylpropanoids, flavonoid and iridoid glycosides of various Jasminum species (Jin W. et al., 2006) which possess good bioactivities such as antioxidant activity, antiviral, antibiosis, antitumour, liver and gallbladder-protection, eliminating pain and relieving spasm, strengthening immunity as well as anti-diabetes and anti-hyperlipidenmia activity, playing significant roles in clinical medicine. The literature shows that no information exists regarding the use of Jasminum nervosum Lour. stems as a source of natural antioxidants. Therefore, the objectives of this study were to evaluate the stems of Jasminum nervosum Lour. as a source of natural antioxidants using different fractions to determine their antioxidant capacities. Several in vitro assays, including radical-scavenging assays with 1,1-Diphenyl-2-picryl hydrazyl (DPPH), 2,2’-Azobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and reducing power were carried out to evaluate the antioxidant activity of the extracts from Jasminum nervosum Lour. stems. Because of the important roles of the phenolics and flavonoids as antioxidants, the amounts of total phenolics (TP) and total flavonoids (TF) in the extracts were also determined by spectrophotometric and HPLC methods, expressed as Gallic acid and Lutin equivalents, respectively.

2,2’-Azobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and Butylated hydroxyltoluene (BHT) were purchased from Sigma Aldrich Co., St. Louis, MO, USA; Gallic acid standard and Lutin standard was purchased from J&K Scientific Ltd., Beijing, China; Acetonitrile (HPLC grade), Methanol (HPLC grade) and Acetic acid (reagent grade) were purchased from Merck&Co., Inc, Germany; Other chemicals were purchased from China National Medicine Group Shanghai Corporation, Shanghai, China. All chemicals and solvents used were of analytical grade. 2.2.  Preparation of extracts Jasminum nervosum Lour. was obtained from the Guangxi Provnice, in August 2009 (summer) before the flowering stage, and authenticated by professor Song Ji Wei, Department of Zhuang Pharmacy, GuangXi traditional Chinese Medical University. The samples were initially air-dried and then reduced to small particles. The particles selected for analysis were passed through a 40-mesh screen and suspended in 95% ethanol for 48 h at room temperature. The extracts were concentrated, suspended in deionized water and sequentially partitioned with petroleum ether, ethyl acetate, and n-butanol to obtain three different fractions (water part was discarded). Fractions were collected, dried under a rotary evaporator and kept in the dark at 4°C until testing. The extract yields with petroleum ether (PE), ethyl acetate (EAE), and n-butanol (BE) from the stems of Jasminum nervosum Lour. were 0.71%, 0.73%, and 1.87% (w/w), respectively. 2.3.  Spectrophotometric procedures Determination of TP content Total soluble phenolics were determined using the Folin-Ciocalteau reagent according to the method of Slinkard and Singleton (1977) and gallic acid as an internal standard. Briefly, the fraction solution (0.5 ml, 1 mg ml21) was diluted with distilled water (46 ml) in a volumetric flask. Folin-Ciocalteau reagent (1 ml) was added and mixed thoroughly. After 3 min, a sodium carbonate solution (3 ml, 2%) was added and the mixture was allowed to stand for 2 h with intermittent shaking. The absorbance was measured at 760 nm. The concentration of TP compounds in the extracts was determined as micrograms of Gallic acid equivalent per gram of dry matter. All tests were done in triplicate. Determination of TF content

2.  MATERIALS AND METHODS 2.1.  Chemicals 1,1-Diphenyl-2-picryl hydrazyl (DPPH) (purity 98%) was purchased from Wako Chemicals, Japan; 150

grasas y aceites,

TF content was determined following the procedure by Dewanto et al. (2002), and lutin as an internal standard. A fraction solution (3 ml, 0.8 mg ml21) was placed in a 10-ml volumetric flask and 5 ml of distilled water were added followed by a NaNO2

62 (2),

abril-junio,

149-154, 2011,

issn:

0017-3495,

doi:

10.3989/gya.066810

antioxidant activity, total phenolic, and total flavonoid of extracts from the stems…

solution (0.3 ml, 5%). After 5 min, an AlCl3 solution (0.6 ml, 10%) was added. After another 5 min, a NaOH solution (2 ml, 1 M) was added and the volume was made up with 95% ethanol. The solution was mixed thoroughly and absorbance was measured at 510 nm. TF amounts were expressed as micrograms of Lutin equivalent per gram of dry matter of extract. All tests were performed in triplicate. 2.4. High-performance liquid chromatography (HPLC) analysis Preparation of standard and sample solutions The phenolic and flavonoid compounds in the stem extracts of Jasminum nervosum Lour. were determined by HPLC, performed with a Waters 600 diode array detector system equipped with a dualistic pump. The analyses were carried out on a Diamonsil C18 column (4.6 mm  250 mm, 2.5 μm). Standard stock solutions of Gallic acid and Lutin were prepared in 95% EtOH, at a concentration of 0.5 mg ml21. All sample solutions of PE , EAE and BE (1 mg/ml21) were filtered through a 0.45 μm membrane filter (Millipore) and injected directly. HPLC analysis of phenolic compounds The mobile phase was composed of sovent A (0.3% Acetic acid) and sovent B (Acetonitrile). The gradient programme was as follows: (0–5 min, 20% B; 5–10 min, 90% B; 10–15 min, 10% B; 15–25 min, 20% B); flow rate 1 ml/min; volume injected 20 μl; temperature 25°C; UV detection wavelength 280 nm. All experimental data are expressed as means  SD values (n 5 3).

control)]  100%. The control contains all reagents except the extract. The DPPH radical scavenging activity of BHT (0.5 mg ml21) was also assayed for comparison. All tests were performed in triplicate and means were centered. 2.6.  ABTS+• radical cation scavenging The antioxidant activities of various solvent extracts in the reaction with the stable ABTS1• radical cation were determined according to the method of Re et al. (1999) with slight modifications. The reaction between ABTS and potassium persulfate directly generates the blue/ green ABTS1• chromophore, which can be reduced by an antioxidant, thereby resulting in a loss of absorbance at 734 nm. ABTS was dissolved in water to a 7 mM concentration. ABTS radical cation (ABTS1•) was produced by reacting an ABTS stock solution with 2.45 mM potassium persulfate (final concentration) and allowing the mixture to stand in the dark at room temperature for 12–16 h. The ABTS1• solution (stable for 2 days) was diluted with a phosphate buffer (2 mM, PH 7.4) to achieve an absorbance of 0.70.05 at 734 nm. Extract solutions (10 μl) were mixed with ABTS1• solution (5 ml), and absorbance was read at ambient temperature after 1 min. PBS solution was used as a control. All tests were done in triplicate. The radical-scavenging activity of the samples was expressed as SC% 5 [(Acontrol–Atest)/ Acontrol]  100%, in which Acontrol is the absorbance of the control (ABTS1• solution without test sample) and Atest is the absorbance of the test sample (ABTS1• solution plus extract). BHT (0.2, 0.5, 0.8 and 1.2 mg ml-1) was also assayed for comparison. 2.7.  Reducing power

HPLC analysis of flavonoid compounds The mobile phase was composed of sovents of MeOH-Water-Acetic acid (40:57.5:2.5) at a flow rate of 1.0 ml/min. The column temperature was set at 25°C. The injection volume was 20 μl. The UV detection wavelength was monitored at 254 nm. All experimental data are expressed as meansSD values (n 5 3). 2.5.  Scavenging activity on DPPH radical The scavenging effect of different Fractions on the DPPH radical was measured using a modified version of the method described by Shimada et al. (1992). In brief, extract solutions (20 μl) in 95% ethanol at different concentrations (0.2, 0.5, 0.8 and 1.2 mg ml21) were added to 8 ml 0.0004% (w/v) solution of DPPH in 95% ethanol. The reaction mixtures were incubated at 28°C. The scavenging activities on DPPH radicals were determined by measuring the absorbance at 515 nm every 10 min until the reaction reached the steady state. The antioxidant activity was expressed as a percentage of scavenging of DPPH: SC% 5 [1–­(absorbance of sample)/(absorbance of



grasas y aceites,

62 (2),

abril-junio,

149-154, 2011,

issn:

0017-3495,

doi:

The reducing power of Jasminum nervosum Lour. stems was determined according to the method of Oyaizu (1986). Fraction solutions (100 μl) in 95% ethanol (0.2, 0.5, 0.8 and 1.2 mg ml21) were mixed with phosphate buffer (2.5 ml, 0.2 M, pH 7.4) and potassium ferricyanide [K3Fe(CN)6] (2.5 ml, 1%). The mixture was incubated at 50°C for 20 min. A portion 2.5 ml of trichloroacetic acid (10%) was added to the mixture, which was then centrifuged at 3000 rpm for 10 min. The upper layer of solution (2.5 ml) was mixed with distilled water (2.5 ml) and FeCl3 (0.5 ml, 0.1%), and the absorbance was measured at 700 nm. Increased absorbance of the reaction mixture indicated increased reducing power. The reducing power of BHT (0.2, 0.5, 0.8 and 1.2 mg ml21) were also assayed for comparison. 3.  RESULTS AND DISCESSION 3.1. Determined of the TP and TF content by Spectrophotometric procedures The total phenolics content (TP) and the total flavonoid content (TF) in the extracts were

10.3989/gya.066810

151

l. huo, r. lu, p. li, y. liao, r. chen, ch. deng, ch. lu, x. wei and y. li

determined from a regression equation for the calibration curve (y 5 0.0796x 1 0.0091, R2 5 0.9925) and (y 5 0.8836x 2 0.0255, R2 5 0.9998), respectively. The amount of TP and TF of PE, EAE, and BE are demonstrated in Table 1. EAE revealed the highest total phenolic content at 120.11  0.42 mg g-1 dry weight, expressed as Gallic acid equivalents, followed by BE and PE. In Addition, BE had the highest flavonoid content (219.221.77 mg g-1 dry weight, expressed as Lutin equivalents) compared to that of other extracts. No significant differences were observed in TP/TF contents between BE and EE. From the introduction, we know that TP and TF content may contribute directly to antioxidant action. Therefore, ethyl acetate and n-butanol may be the most effective extracting solvents for antioxidants from the stems of Jasminum nervosum Lour. These results show that the antioxidant activity of extracts of the stems of this plant may be related to their phenolic or flavonoid substrates. Table 1 TP contents and TF contents in different extracts from the stems of Jasminum nervosum Lour Sample

TP

a

identified in the PE, EAE, and BE according to their retention times and the spectral characteristics of their peaks against those of standards. As shown in Table 2, EAE contained the highest content of Lutin and Gallic acid. No gallic acid peak was found in PE. By comparing the different fractions, the content of Gallic acid and Lutin decreased in the same order of EAE.BE.PE, and the rank order of EAE and BE was different according to their antioxidant pontency and free radical-scavenging ability. This result indicates that, besides Gallic acid and Lutin, the other compounds in Jasminum nervosum Lour. stem extracts may also be responsible for the antioxidant activity. The ethyl acetate solution is more suitable for the extraction of both Gallic acid and Lutin.

Table 2 Contents of Lutin and Gallic acid of different extracts from the stems of Jasminum nervosum Lour. by HPLC Sample

Gallic acid a

Lutin b

PE

20.601.02

ND

TF

EAE

92.540.59

87.660.88

BE

55.260.73

67.421.33

b

PE

52.002.96

117.143.42

EAE

123.212.82

195.692.83

BE

108.610.31

219.221.77

Results are mean  standard deviation of three parallel measurements. a Milligrams of Gallic acid per 1 g (dry weight) of extract. b Milligrams of Lutin per 1 g (dry weight) of extract.

Results are mean  standard deviation of three parallel measurements. a Milligrams of Gallic acid per 1 g (dry weight) of extract. b Milligrams of Lutin per 1 g (dry weight) of extract. ND, Not detected.

3.3.  Effect of DPPH• scavenging activity

Figure 1 HPLC chromatogram of reference standards (1) Gallic acid, (2) Lutin and (3), (4) unknown residues.

3.2. Quantitative analyses of Lutin and Gallic acid by HPLC RP-HPLC coupled with UV–Vis DAD was employed to separate, identify and quantify phenolic compounds in the fractions of Jasminum nervosum Lour. stems. The concentrations were determined by calculating the HPLC peak areas which are proportional to the amount of analytes in a peak and presented as the mean of three determinations which were highly repeatable. The standard curves for Gallic acid and Lutin were y 5 3397.9413 1 10474.6674  (R2 5 0.9905) and y 5 4.8920 1 16640.1563  (R2 5 0.9981), respectively. Figure 1 shows the chromatogram of reference standards of Gallic acid and Lutin. These compounds have been 152

grasas y aceites,

The scavenging abilities of various extracts against DPPH radical were concentration-dependent, and increased steadily with time (Figure 2). According to the results, PE showed no obvious scavenging activity (Figure 2 A). EAE showed just a little higher antioxidant activity than BE in this assay system except for the concentration of 1.2 mg ml21 in BE, which reached the maximum scavenging effect among all the extracts (Figure 2 B and C). The scavenging activities of EAE on the DPPH radical were superior to the positive control, BHT (0.5mg ml21). However, at a low concentration (0.2 mg ml21), the scavenging activity of EAE was not as effective as BHT (Figure 2 B). Only the scavenging activity of BE at concentration (0.8 and 1.2 mg ml21) was better than BHT (Figure 2 C). It should be noted that EE and BE have high amounts of TP and TF and also exhibited good DPPH radical scavenging activity. 3.4.  Effect of ABTS1• scavenging activity All extracts tested showed significant ABTS scavenging capacity (Figure 3). The control (BHT) and the three fractions exhibited concentrationdependent ABTS1• scavenging activity. EAE and BE exhibited similar high scavenging capacity, whereas the lowest activity was found in the PE, which was in accordance with the TP and TF

62 (2),

abril-junio,

149-154, 2011,

issn:

0017-3495,

doi:

10.3989/gya.066810

antioxidant activity, total phenolic, and total flavonoid of extracts from the stems…

Figure 2 DPPH free radical scavenging activity of PE (A), EAE (B) and BE(C) from Jasminum nervosum Lour. stems compared with BHT (0.5 mg ml21). Results are mean  standard deviation of three parallel measurements.

content. The ABTS1• radical scavenging of EAE and BE increased rapidly with sample concentration. After 1min the scavenging rate could reach 50% to 95%, but increased slowly at higher concentrations. Under the same experimental conditions, the scavenging activity of the three fractions was lower than the synthetic antioxidant, BHT, but at high concentrations in BE, EAE and BHT, no obvious differences were detected among them.

donating capacities in a concentration-dependent manner and BE was the most potent reducing agent. The reducing power of the three fractions was: BE.EE.PE. The reductive capability of BE was slightly higher than EE. It is noted that the reducing power of the three fractions was related to the TP and TF contents. Only BE, at low concentrations of 0.2 mg ml21 and 0.5 mg ml21, had a stronger reductive capacity than BHT.

3.5.  Reducing power

4.  CONCLUSIONS

Figure 4 depicts the reducing properties of each fraction at different concentrations. The results suggest that as electron donors, each extract could convert free radicals into more stable products, leading to the termination of radical chain reactions. Various extracts showed some degree of electron

The highest amount of TP was 123.212.82 mg per gram of dry extract in EAE, while the greatest TF content was 219.221.77 mg per gram of dry extract in BE. The HPLC data indicated that Jasminum nervosum Lour. stems extracts contained phenolic and flavoniod compounds, such as Gallic acid and



grasas y aceites,

62 (2),

abril-junio,

149-154, 2011,

issn:

0017-3495,

doi:

10.3989/gya.066810

153

l. huo, r. lu, p. li, y. liao, r. chen, ch. deng, ch. lu, x. wei and y. li

Figure 3 ABTS1 free radical scavenging activity of PE, EAE and BE from Jasminum nervosum Lour. stems with BHT as a positive control. Results are mean ± standard deviation of three parallel measurements.

Lutin, except that there’s no Lutin peak found in PE. PE was proved to be the worst part in this plant for antioxidant axtivity as determined by both methods due to its low content of phenolics or flavonoids. The overall results showed that the extracts of Jasminum nervosum Lour. stems exhibited varying degrees of TP/TF contents, and antioxidant activity in vitro and that it can be used as an easily accessible source of natural antioxidants. Future work is advised to provide additional information about the chemical composition of the extracts in order to better understand their mechanism of action as antioxidants. ACKNOWLEDGMENTS This investigation was supported by a grant (P2009035) for subject for Guangxi traditional Chinese Medical University, China. REFERENCES Bushra S, Farooq A, Muhammad RA, Shahzad ASC. 2008. Antioxidant potential of extracts from different agro wastes: Stabilization of corn oil. Grasas y Aceites, 59, 205–217. Dewanto V, Wu X, Adom K K, Liu R H. 2002. Thermal processing enhances the nutritional value of tomatoes by increasing total antioxidant activity. J. Agric. Food Chem. 50, 3010–3014. Editorial committee of the National Chinese Medicine Administrative bureau. 2005. Chinese materia medica. Shanghai Scientific and Technical Education Publishing House, Shanghai. Halliwell B, Grootveld M. 1987. The measurement of free radical reactions in humans. FEBS Lett. 213, 9–14. Jin W, Jan GF. 2006. Biological activities of iridoids, Herald of Medicine., 25, 530–533. Li H, Hao Z, Wang X, Huang L, Li J. 2009. Antioxidant activities of extracts and fractions from Lysimachia foenum-graecum Hance. Bioresour Technol. 100, 970–974.

154

grasas y aceites,

Figure 4 The reducing power of PE, EAE and BE from Jasminum nervosum Lour. stems with BHT as a positive control. Results are mean  standard deviation of three parallel measurements.

Ku CS, Mun S P. 2007. Antioxidant activities of ethanol extracts from seeds in fresh Bokbunja (Rubus coreanus Miq.) and wine processing waste. Bioresour Technol. 99, 2852–2856. McDermott JH. 2000. Antioxidant nutrients: current dietary recommendations and research update. J. Am. Pharm. Assoc. 40, 785–799 Mohammad AE, Seyed FN, Seyed MN, Bahman E. 2010. Antihypoxic and antioxidant activity of Hibiscus esculentus sedes. Grasas y Aceites, 61, 30–36. Othman A, Ismail A, Ghani AN, Adenan I. 2007. Antioxidant capacity and phenolic content of cocoa beans. Food Chemistry 100, 1523–1530. Oyaizu M. 1986. Studies on product of browning reaction prepared from glucose amine. Jpn. J. Nutr. 44, 307– 315. Ozsoy N, Can A, Yanardag R, Akev N. 2008. Antioxidant activity of Smilax excelsa L. leaf extracts. Food Chemistry 110, 571–583. Paganga G, Miller N, Rice-Evans CA. 1999. The polyphenolic content of fruit and vegetables and their antioxidant activities. What does a serving constitute?. Free Radical. Res. 30, 153-162. Re R, Pellegrini N, Proteggente A, Pannala A, Yang M, Rice Evans C. 1999. Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic. Biol. Med. 26, 1231–1237. Sasak YF, Kawaguchi S, Kamaya A, Ohshita M, Kabasawa K, Iwama K, Taniguchi K, Tsuda S. 2002. The comet assay with 8 mouse organs: Results with 39 currently used food additives. Mutation Research/ Genetic Toxicology and Environmental Mutagenesis. 519, 103–119. Shimada K, Fujikawa K, Yahara K, Nakamura T. 1992. Antioxidative properties of xanthan on the antioxidation of soybean oil in cyclodextrin emulsion. J. Agric. Food Chem. 40, 945–948. Slinkard K, Singleton VL. 1977. Total phenol analyses: automation and comparison with manual methods. Am. J. Enol. Vitic. 28, 49–55. Tevfik Özen. 2010. Antioxidant activity of wild edible plants in the Black Sea Region of Turkey. Grasas y Aceites, 61, 86–94. Recibido: 24/6/10 Aceptado: 24/8/10

62 (2),

abril-junio,

149-154, 2011,

issn:

0017-3495,

doi:

10.3989/gya.066810

grasas y aceites,

62 (2), 155-161, 2011, issn: 0017-3495 doi: 10.3989/gya.067210

abril-junio,

Oxidative stability of lard and sunflower oil supplemented with coffee extracts under storage conditions · By Graz·yna Budryn,* Ewa Nebesny and Dorota Zyz·elewicz Department of Chemical Food Technology, Faculty of Biotechnology and Food Sciences, Technical University of Lodz, Stefanowskiego 4/10, 90-924 Lodz, Poland (*Corresponding author: [email protected])

RESUMEN Estabilidad oxidativa de manteca y aceite de girasol suplementados con extractos de café bajo condiciones de almacenamiento. La estabilidad oxidativa de manteca y aceite de girasol suplementados con extractos acuosos de granos de café verde o tostado Arábica y Robusta fue estimada. Un descenso en la velocidad de las reacciones de oxidación de la grasa durante el almacenamiento de las muestras durante 12 semana a temperatura ambiente, que resulto de la adición de los extractos de café, fue evaluada usando métodos químicos estándares tales como la determinación de peróxidos y el índice de paranisidina y ensayos de dienos y trienos conjugados, así como métodos físicos tales como la determinación del perfil térmico por DSC. También las propiedades sensoriales de todas las grasas fueron estimadas. Estas medidas mostraron que extractos acuosos de café al 0.1% en la grasa decrecieron (p  0.05) los valores obtenidos por los métodos químicos con respecto al control y aproximadamente redujo a la mitad la velocidad de oxidación. También el análisis del perfil térmico revelo que la suplementación con extractos de café redujo la extensión de cambios negativos en las propiedades de las grasas. La efectividad de los extractos de café ensayados decrecieron en el orden: Robusta verde . Arábica verde . Robusta tostado . Arábica tostado. PALABRAS-CLAVE: Aceite de girasol – Estabilidad oxidativa – DSC – Extractos de café – Manteca. SUMMARY Oxidative stability of lard and sunflower oil supplemented with coffee extracts under storage conditions. The oxidative stability of sunflower oil and lard supplemented with water extracts of green and roasted, Arabica and Robusta coffee beans was estimated. A decrease in the rate of fat oxidation reactions during the storage of samples for 12 weeks at ambient temperature which resulted from the addition of coffee extracts was evaluated using standard chemical methods such as the determination of peroxide and p-anisidine value and the assays of conjugated dienes and trienes as well as physical methods such as the determination of thermal profile by DSC. The sensory properties of all fat samples were also determined. These measurements showed that 0.1% water coffee extracts in fats decreased (p0.05) the quantities assayed by the chemical methods as compared to the control samples and approximately halved the rate of fat oxidation. In addition, the thermal profile analysis revealed



that supplementing with coffee extracts reduced the extent of negative changes in the thermal properties of fats. The effectiveness of the tested coffee extracts decreased in the order: green Robusta . green Arabica . roasted Robusta . roasted Arabica. KEY-WORDS: Coffee extracts – DSC – Lard – Oxidative stability – Sunflower oil.

1.  INTRODUCTION The majority of oils and edible fats are chemically unstable, in particular when exposed to oxygen and light which bring about oxidation even under standard storage conditions. The oxidation of fats not only gives rise to an unpleasant smell and taste but also generates potentially harmful or toxic compounds (Luzia et al., 1997). The consumption of oxidized lipids leads to the formation of reactive forms of oxygen and free radicals which in turn oxidize biological membranes and display potential mutagenic, genotoxic and angiotoxic activities. They also contribute to the development of cardiovascular system disorders, faster ageing processes, a loss in vitamins, mainly tocopherols, and act as inhibitors of enzymes (Sikwese and Duodu, 2007). Oxidation not only decreases the stability of oils and fats during long term storage but also negatively affects the stability of lipid containing foods, in particular those which were thermally processed like fried foodstuffs (Anwar et al., 2006). To some extent these disadvantageous processes taking place during the storage of fats can be slowed down through supplementation with antioxidants. The most frequently applied, synthetic antioxidants pose a threat to human health and therefore natural antioxidants have attracted attention. The occurrence of many natural antioxidants in raw materials, mainly of plant origin, has been well documented (Bandonien[ et al., 2000). Up to now, the most promising results come from investigations of extracts from rosemary and sage that stabilized oils used in frying (Irwandi et al., 2000). Potentially rich sources of antioxidants are also such plants like rice, garlic, grapes, green tea, and coffee (Renuka Devi et al., 2007, Iqbal and Bhanger, 2007). Previously, roasted 155

G. BUDRYN, E. NEBESNY AND D. ¶Y¶ELEWICZ

coffee was added to foods as an antioxidant, as it is usually consumed after roasting (Nissen et al., 2004). However, green coffee contains much more (even 10-fold) polyphenols than roasted coffee (Budryn et al., 2009). Anwar et al. (2006) investigated the effects of sunflower oil supplementing with methanolic green coffee extracts. However, chlorogenic acids that are responsible for the antioxidant activity of green coffee beans are more soluble in water than in alcohols. Besides, water is nontoxic and therefore aqueous coffee bean extracts should also be used in studies on the oxidative stability of fats. Various reactions taking place during fat storage can be monitored by using diverse analytical methods enabling the determination of polar compounds and free fatty acid contents along with iodine, peroxide and p-anisidine values. However, these methods are time and labor consuming, and usually require the application of toxic reagents (Tan and Che Man, 1999a). Therefore they are frequently replaced by instrumental methods including spectrophotometric ones and the differential scanning calorimetry (DSC) (Gloria and Aguilera, 1998). The latter are well correlated with the results of chemical methods. The highest correlation was observed between the results of DSC and polar compound assays (Tan and Che Man, 1999b). The objective of this work was to determine the effect of the aqueous extracts of green and roasted coffee beans on the long term oxidative stability of lard and sunflower oil as representative edible fats. The analyses were carried out using standard and instrumental methods. 2.  MATERIALS AND METHODS 2.1.  Materials The tested materials were pork lard from a local market and sunflower oil purchased from “Kruszwica” ZPT (Poland). They were supplemented with dried water extracts from green and roasted Robusta coffee beans (Coffea canephora), which were grown in Indonesia and processed by the dry method as well as from green and roasted Arabica coffee beans (Coffea arabica), from Columbia, processed by the wet method. Green Robusta and Arabica beans were purchased from Aspol Ltd. (Poland). 2.2.  Coffee bean roasting Both types of coffee beans were roasted in a laboratory, convective coffee roaster with an automatic cooler, Precission Heartware (USA), until a 17% decrease in their weight was achieved (a difference in weight before and after roasting per 100 g coffee beans (Daglia et al., 2000). The temperature inside the roasting chamber was measured using a NiCr-NiAl thermocouple (Poland). In the last stage of roasting it reached 240 and 235oC for Robusta and Arabica (roasting duration of 12 and 9.5 min), respectively. The humidity of 156

grasas y aceites,

coffee beans was determined by drying at 103oC until constant weight (Lakenbrink, 2000). It was 0.8 and 1.1% for Robusta and Arabica, respectively. The color of green and roasted coffee beans was measured according to the CIE L*a*b* system (Lopez-Galilea et al., 2006) using a Specord M-40 spectrophotometer, Carl-Zeiss Jena (Germany). Changes in color parameters caused by the roasting of Robusta coffee beans were as follows: L* decreased from 70.33 to 42.65, a* increased from 2.83 to 5.34, b* decreased from 21.57 to 20.66. The changes for Arabica coffee beans were similar: L*: decreased from 75.51 to 41.84, a*: increased from 1.68 to 4.68, b*: decreased from 20.28 to 19.47. Green coffee beans were ground in a laboratory mill WZ-1, ZBPP (Poland), which was adapted to the grinding of very hard beans. The roasted coffee beans were ground in a Il Macinino F.A.C.E.M Spa – Tre Spade mill (Italy). The dimensions of the ground particles (suspended in paraffin oil) were measured using a micrometric screw NSK Digitrix Mark II Electronic Micrometer, Japan Micrometer MFG.CO.LTD (Japan). Their dimensions ranged from 480 to 680 μm. 2.3.  Preparing of coffee extracts Extracts from ground, green and roasted, Arabica and Robusta coffee beans were prepared at a coffee: water ratio of 1:5.75, which ensured approximately 5% solid concentration, that was optimal for extract lyophilization. The extracts were obtained through boiling in water at 110oC for 10 min in a pressure cooker PS-5682 First (Austria). Rapid cooling and filtration under reduced pressure were carried out in a vacuum pump KNF Neuberger N 035.3 AT.18 (Germany). After that, the extracts were freeze-dried in a DELTA 1-24LSC Christ lyophilizer (Germany) to prevent deterioration of the preparations and to limit the addition of water to the fat samples (Shishikura et al., 2006). The solid substance content in the extracts was assayed through drying at 103oC. It was 1.3, 2.5, 1.0 and 2.8% for extracts from green and roasted Robusta and green and roasted Arabica, respectively. The concentrations of chlorogenic acids (determined by HPLC) were 37.6, 3.74, 19.56 and 6.19% of dried extracts and the pH values of solutions prepared by dissolving 2 g of dry extract in water to 100 mL volume measured with a Schott CG Schott Geräte GmbH pH-meter (Germany) were 6.0, 5.3, 5.9 and 5.1, respectively. 2.4. Preparation of fat samples supplemented with coffee bean extracts The concentration of the lyophilized coffee extracts added to lard and sunflower oil was 0.1%. This concentration was selected based on the results of earlier studies aimed at compromising between the antioxidant effect and sensory properties. After the addition of coffee extracts to sunflower oil

62 (2),

abril-junio,

155-161, 2011,

issn:

0017-3495,

doi:

10.3989/gya.067210

OXIDATIVE STABILITY OF LARD AND SUNFLOWER OIL SUPPLEMENTED WITH COFFEE EXTRACTS UNDER STORAGE CONDITIONS

3.  RESULTS AND DISCUSSION

62 (2),

abril-junio,

155-161, 2011,

-1

10

issn:

0017-3495,

doi:

8

6

4

2

0 0

80

3

6

3

6

weeks

9

12

9

12

b

70 60 50 40 30

10

Detection of the initial products of fatty acid oxidation is of particular importance during investigations of long term storage at ambient temperature when comparing to high temperature conditions, where these substances are quickly destroyed. Fig. 1 shows changes in peroxide value



a

20

3.1. Oxidative stability evaluated by chemical methods

grasas y aceites,

12

meq O2 x kg

The extent of oxidative changes during storage was estimated based on changes in iodine, peroxide and p-anisidine values, indexes of conjugated dienes and trienes contents (according to standard methods) as well as on changes in the thermal profile as evaluated by DSC. The thermal analysis enabling estimation of the intensity of oxidative changes in fats during their storage was conducted using a DSC 111 (Setaram, France). Fat melting was carried out in the temperature range of 20 and 60oC for lard and -55 and 5oC for sunflower oil. Lard and sunflower oil samples were cooled starting from an ambient temperature until complete solidification and were kept for 5 min at temperatures of 20 and -55oC, respectively. The heating rate was 3oC  min21. Results of the thermal analysis were processed using the scanning calorimeter software. Melting temperature was determined based on peaks on melting curves and the enthalpy of phase transition was computed based on the surface area under peaks I and II and on a sample mass basis. Calibration of temperature and energy was carried out using the indium standard and the same scanning rate as for tested samples. Evaluation of the sensory properties of fats was carried out by 5 well trained panelists having sufficiently high sensory sensitivity and knowledge related to the evaluated properties. Results were described in a 5-point intensity scale in which 1 corresponded to extremely unaccepted, changed and indicating oxidative deterioration features while 5 corresponded to typical, characteristic and well accepted properties. The taste, aroma and color of fats were evaluated according to this scale and the overall score was a mean of these 3 partial scores. All evaluations were carried out in triplicate. Their results were subjected to variance analysis (ANOVA) and the Duncan test (DMRT). Statistical significance of the results was determined at p  0.05.

meq O2 x kg

2.5. Analysis of fats

(PV) during the storage of lard and sunflower oil supplemented with extracts from green and roasted, Robusta and Arabica coffee beans. After 12 weeks of storage the PV of lard without coffee antioxidants (control) was more than 15 times higher than the initial PV (0.6 meq O2  kg21) and reached 10.6 meq O2  kg21 (Fig. 1a). The significant differences (p  0.05) in PV between the control and samples supplemented with coffee extracts were observed. Extracts from both types of green coffee considerably slowed down (p0.05) the growth of PV starting from the third week of storage and after 12 weeks PV values were only approximately twice that of the fresh material (1.1 and 1.3 meq O2  kg21 for extracts from green Robusta and Arabica, respectively). The positive effect of extracts from roasted Robusta and Arabica coffee beans on PV was observed (p