Studies on the preparation of structured lipids from rice bran oil

Rajni, Chopra (2008) Studies on the preparation of structured lipids from rice bran oil. PhD thesis, University of Mysore.

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Abstract

Rice bran oil (RBO) is the byproduct of rice milling and it is consumed widely in Asia. It is produced from the bran and polish of rice, which are by-products of the rice milling industry. India is the second largest producer of rice in the world and has high potential to produce RBO. It is used for both edible and industrial applications. RBO is unconventional oil with its fatty acid composition very close to that of groundnut oil and has high content of unsaponifiable matter. There has been much interest in RBO in recent years because of its nutraceuticals like tocopherols, tocotrienols, and γ-oryzanol, which have been found to have several health beneficial effects. Structured lipids are triacylglycerols (TAGs) that have been modified to incorporate new fatty acids or have been restructured to change the position of fatty acids to produce novel/new TAGs. Structured lipids can be produced to alter the physical characteristics, or improve the nutritional quality of fats and oils. RBO contains approximately 38 % oleic acid, 34 % linoleic acid and 18.6 % palmitic acid and it lacks n-3 PUFA. The physical properties of these fatty acids do not confer any special properties for its use in food applications. It is interesting to note that stearic acid content of RBO is very low. Though stearic acid is a saturated fatty acid, studies have been shown non-atherogenic nature of stearic acid. Saturated fatty acids are known to impart various physico-chemical and thermal stability properties to oils and fats. Cocoa butter which is rich in stearic acid is currently the fat of choice in the confectionery industry. Because of its demand for confectionery and desirable physical and nutritional quality, cocoa butter substitutes and cocoa butter equivalents have recently become more common. It is of interest to see of whether structured lipid with RBO containing n-3 fatty acids such as α - linolenic acid, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which are having several health benefits, can be synthesized. Therefore, the objective of this study was to modify the RBO to produce structured lipids containing stearic acid and n-3 fatty acids so that utilization of RBO can be increased and value added products can be prepared from RBO. Objectives and plan of work The major objectives of the present investigation are to prepare structured lipids from RBO rich in stearic acid, α- linolenic acid and EPA+DHA by enzyme catalyzed reaction using immobilized lipase from Rhizomucor miehei. The structured lipids enriched with stearic acid and n-3 PUFA were evaluated for their physico-chemical and hypocholesterolemic and anti-aggregation effects respectively and accordingly the following work plan has been envisaged. 1. Standardization of different reaction conditions like incubation time, temperature, enzyme concentration and substrates molar ratio on the incorporation of stearic acid into RBO. Purification of reaction product and studies on physico-chemical studies. 2. Optimization different reaction conditions like incubation time, temperature, enzyme concentration and substrates molar ratio using response surface methodology on the incorporation of n-3 PUFA into RBO. Synthesis of large scale structured lipids rich in n-3 PUFA and their purification. 3. Evaluation of structured lipids rich in n-3 PUFA for their hypocholesterolemic effects and hepatic antioxidant enzymes and platelet aggregation in rats fed n-3 PUFA rich structured lipids. Chapter I: Introduction This chapter deals with the general introduction of fats and oils, lipid classification, different types of fatty acids and their physiological effects, biological significance and health benefits of n-6 and n-3 fatty acids, metabolic pathway of n-6 and n-3 fatty acid synthesis. This chapter also includes the effects of dietary lipids on health, health benefits of n-3 PUFA on cardiovascular system, enzymes regulating calcium ion channels, in the vascular retina, haemostatic system, endothelial function and nervous system. A description on the edible oils used in India, dietary recommendation of different fatty acids and need for the balance of fatty acids in the diet is given. Uses and importance of structured lipids, importance of immobilized enzymes in modification of fats and oils are described in this chapter. Chapter II: Materials and methods In this chapter brief protocols of all procedures carried out throughout the investigation have been given with appropriate references. Described here are standardization of different reaction conditions for the synthesis of structured lipids enriched in stearic acid, purification of newly synthesized TAG by thin layer chromatography, quantitation of triglycerides, analysis of fatty acids by gas chromatography and fatty acids at sn-2 position of triglyceride and differential scanning calorimetric studies of structured lipids rich in stearic acid. Analytical methods for minor constituents like γ- oryzanol, tocopherols and tocotrienols of native and modified RBO were also given. This chapter describes the preparation of free fatty acids by the hydrolysis of linseed and cod liver oil and preparation n-3 fatty acid concentrate from linseed oil by urea inclusion method. Optimization of reaction conditions like incubation time, temperature, enzyme concentration and substrates molar ratio by using response surface methodology for the acidolysis reaction were also described. Scale up of the enzyme catalyzed acidolysis and large scale preparation of structured lipids rich in n- 3 PUFA and preparation of blended oil from RBO with linseed oil and cod liver oil and nutritional evaluation of these structured lipids were enumerated. Chapter III: Synthesis of structured lipids with RBO enriched in stearic acid This chapter begins with the brief introduction of RBO, structured lipids, importance of structured lipids for the improvement of physical properties of fats and oils and importance and effect of stearic acid on cholesterol level. Enzymatic acidolysis reactions were carried out in 50 mL stoppered conical flasks under inert atmosphere. Different reaction conditions like incubation time, temperature, substrates molar ratio and enzyme concentration on the incorporation of stearic acid into RBO were studied. The effect of reaction time on the incorporation of stearic acid into RBO was studied from 0 to 48 h keeping other parameters constant. The effect of reaction temperature on the incorporation of SA into RBO was studied from 25 to 60 0C keeping other parameters constant. The effect of molar ratio of substrates on incorporation of SA into RBO was studied from 1:1 to 1:10. The effect of enzyme concentration on incorporation of SA into RBO was studied from 1 to 10% of the weights of both the substrates. Stearic acid was successfully incorporated (48.5 %) into RBO under optimal conditions. After incorporation of stearic acid into RBO, fatty acid profile and sn-2 positional analysis was carried out in modified and original RBO using pancreatic lipase catalyzed reaction. Analysis of oryzanol, tocopherols and tocotrienols were carried out in native and modified RBO. The type of the triacylglycerols in modified and native rice bran oil were analyzed by HPLC. After incorporation of stearic acid into RBO, physical properties of native and modified RBO were studied by differential scanning calorimeter and physical properties of modified RBO were compared with vanaspati and cocoa butter. The chapter was concluded by discussion of the results obtained. Chapter IV: Enrichment of RBO with n-3 PUFA by enzymatic acidolysis: optimization of parameters by RSM This chapter begins with the brief introduction about the importance of n-3 PUFA in human health and need of balance between n-6 and n-3 PUFA. The importance of response surface methodology (RSM) for optimizing different reaction conditions for incorporation of n-3 PUFA into the RBO was elaborated. Fatty acid concentrate from linseed oil (LSO) rich in α- linolenic acid (ALA) and free fatty acids from cod liver oil (CLO) were prepared and further used for acidolysis reaction to enrich RBO with ALA and EPA+DHA respectively. Two structured lipids (SL) were obtained with RBO, one was rich in ALA and other rich in EPA+DHA. RSM was used to optimize the reaction conditions for lipase-catalyzed incorporation of n-3 PUFA into RBO. A CCRD with four variables was used to study the response pattern. Purification of the reaction product was done with TLC and column chromatography. Pancreatic lipase hydrolysis was used to determine the fatty acids at sn-2 position in triacylglycerol. The optimum incorporation of ALA (n-3 PUFA) into RBO was about 49 % when incubation time and temperature were 11.5 h and 43.750C respectively and substrates molar ratio and enzyme concentration (%) was maintained at 7.75. The optimum incorporation of EPA+DHA into RBO was about 10 % at 37 h and 32.5 0C incubation time and temperature respectively, where as enzyme concentration (%) and substrates molar ratio was 7.75. The fatty acid composition of substrates and reaction products were analyzed by gas chromatography. Large scale synthesis of these SL rich in PUFA was done. The chapter ends with the discussion. Chapter V: Effects of SL and blends from RBO enriched with n-3 PUFA on liver and serum lipids in rats This chapter focused on the effect of SL with RBO rich in n-3 PUFA on the lipid profile in rats. This chapter begins with a brief mention on the role of dietary fatty acids in health and disease prevention in humans and cardiovascular protective effects of n-3 PUFA. This chapter deals with the hypocholesterolemic effect of RBO rich in n-3 PUFA. Different experimental diets were prepared by including SL with ALA and EPA+DHA and blended oils (RBO and LSO and RBO and CLO) having similar fatty acid composition was fed to rats. Growth and food intake were monitored and food efficiency (FER) was calculated. The serum lipid parameters like total cholesterol, triglycerides, phospholipids, low density and high density cholesterol were estimated. Liver lipid parameters like total cholesterol, triglycerides and phospholipids were estimated. Fatty acid analysis was done for serum, liver, heart, eye, brain and adipose tissues. The chapter was concluded with the discussion. Chapter VI: Antioxidant activity and platelet aggregation in rats fed SL and blends from RBO enriched with n-3 PUFA This chapter begins with a brief introduction about the importance of n-3 PUFA in prevention of inflammatory diseases, including cancer and coronary heart diseases. Effect of RBO rich in n-3 PUFA on platelet aggregation of rats was studied. Antioxidant enzymes like catalase, superoxide dismutase, glutathione reductase and glutathione transferase activity in liver homogenate and serum of rats fed structured lipids enriched with n-3 fatty acid was studied. Lipid peroxides in liver and serum were done. Total Na+, K+ ATPase activity in RBC membrane of rats fed SL rich in PUFA was also studied. Structured lipids enriched with n-3 fatty acid were studied for their inhibitory effects on platelet aggregation of rats. The chapter concludes with discussion. The summary and conclusions of the present findings were given and the thesis concludes with the bibliography arranged alphabetically in sequential order

Item Type:	Thesis (PhD)
Uncontrolled Keywords:	Rice bran oil; Rice milling-; By-products; Rhizomucor miehei; Fats and oils
Subjects:	500 Natural Sciences and Mathematics > 07 Life Sciences > 03 Biochemistry & Molecular Biology > 11 Lipid Biochemistry 600 Technology > 08 Food technology > 19 Lipids-oils/fats
Divisions:	Dept. of Biochemistry
Depositing User:	Food Sci. & Technol. Information Services
Date Deposited:	06 Apr 2010 04:43
Last Modified:	06 Apr 2010 04:43
URI:	http://ir.cftri.res.in/id/eprint/9411

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