5.0 Summary and conclusion

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Stewart Stevens
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1 5.0 Summary and conclusion Trans fatty acids are the geometric isomers of fatty acids that are formed industrially by partial hydrogenation of unsaturated fatty acids from vegetable and marine oil. Trans isomer generally have more density, lower solubility and higher melting point. Often food manufacturers use artificial trans fat in food products because it is inexpensive and it increases the food s shelf life, stability, and texture. The trans configuration has an effect on the functional and physiochemical properties of these fatty acids which in turn effects their metabolism in humans. Through studies, it is found that consumption of Trans fatty acid in diet is related to various health problems like coronary heart diseases, breast cancer, prostrate cancer, obesity etc. also it may effect cell membrane, auto immune system, damage brain cells. The regulations and declaration of the trans fatty acid content in nutrients have caused a request for a database on trans fatty content in foods consumed in different countries. Since there is no database available for trans fatty acids in food from India, the research report will help to generate data about trans fatty acid content in food. Furthermore a detailed monitoring of the cooking fat/oil is necessary in order to investigate TFA s formation in edible fat/oil during various domestic or commercial processing conditions. In view of this, present study was conducted with the following objectives that are to analyze trans fatty acid by various analytical techniques, to quantify trans fatty acids in selected processed foods available in market, to study the effect of thermal and nonthermal processing on trans fatty acids in selected oils/fats and to find the effect of processing on quality parameters of oil. For study on Gas chromatography and Fourier transform infra red spectroscopy, 10 branded samples were selected on which trans fat content was mentioned. The results obtained from these two methods were then compared to select a suitable method. It was found that both the methods are equally suitable for trans fat estimation and no significance difference between the trans fatty acid content analysed by the two methods (t-value obtained was , for p<0.05). However, both these methods have their certain advantages and disadvantages. In the second objective six different types of products traditionally considered as important suppliers of TFA viz. fat/oil, baked products, confectionary, snacks, indigenous 1

2 sweets and indigenous snacks were analysed. For each type of product, premium brands and some store brands were analysed, in order to assess a potential effect on the TFA content. The highest fat content is found in chocolates ranging from 17-56%, followed by chips/bhuji samples 12-25% and then fries 8-29%. Fat content in pastry, cakes and pizza ranged from 7-12%, 11-23% and 10-15% respectively. Among indigenous snacks, fat content in samosa, pakora and mathri ranged from 12-25%, 10-21% and 13-17% respectively. In indigenous sweets, total fat content ranged from %, % and % respectively for jalebi, gulab jamun and laddoo samples. A large variation has been found in the total fat and trans fatty acid profile of these food samples within these groups and also within the various brands among these groups. The trans fat content in non-branded vanaspati samples ranged from 9-23%, which is beyond the limit regulated by FSSAI. In margarine samples trans fat % ranged from %, while in butter sample TFA analysed was 1.8%. In selected bakery products a large variability in trans fat content was found, however a comparatively less TFA was found in this category but one cake sample shows a TFA content of 5.8%. In the confectionaries, Chocolate sample analysed for TFA shows a very high range of trans fat in them with 32% and 26% trans fat in some of the samples whereas in cookies also TFA was found to be high up to 5.8%. in snacks, aloo bhujia has less variability in TFA content ranging from 2.1% to 2.6%. Among Indigenous foods samosa and pakora samples total fat and trans fat % ranged from % and % respectively for samosa and % and % respectively for pakora samples. Mathri samples show trans fat content ranging from1.0 to 4.9%. The high level of trans fat in these sample were basically due to high trans containing vanaspati that are used for frying. In the same way the samples of gulab jamun, jalebi and laddoo very popular in Indian market have their total fat and trans fat % ranged from % and % respectively for gulab jamun, % and % respectively for jalebi and % and % for laddoo respectively. In bhujia samples total fat and trans fat % ranged from % and % respectively. Similarly in potato fries and pizza samples total fat and trans fat % in sample ranged from % and % respectively for fries and % and % respectively for pizza samples. Among all the selected categories a wide variability was observed within the selected samples for 2

3 total fat content and trans fatty acids. The results could not separate the categories in terms of high fat containing or low fat containing groups. Different amount of TFA was found in same food item sourced from different outlets such as branded sweet shop, local sweet shop and roadside vendors. It was observed that none of the food sample, exceeded the maximum limit of peroxide value for oils/fats (maximum permissible limit of peroxide value in oil/fats is specified to be 10 meq/kg of oil, as per FSSAI regulation). Results obtained for the free fatty acid content in various branded and non-branded food samples shows that, among the branded samples 11 out 30 samples were found to contain free fatty acid more than 0.5%, whereas in non-branded samples 21 out of 30 samples contains more than 0.5% FFA,, which is the maximum permissible limit for free fatty acids specified by FSSAI. To study the effect of industrial processing (refining), samples of soybean oil and palm oil at different stages of refining were procured from Mayur oil refinery, Kanpur. No significant change was observed in trans fatty acids during degumming, neutralization and bleaching stages. However, during de-odorization a very significant increase in trans fatty acid content was observed. Palm oil sample show an increase in TFA by 75.9%, whereas in soybean oil sample TFA was found to be increased by 62.3% during de-odorization. Among the two oils selected, palm oil (having lower initial linolenic acid content and higher linoleic acid content) undergoes a higher degree of isomerization corresponding to soybean oil (with higher initial linolenic acid content). Free fatty acid, peroxide value and iodine value of the samples were also analysed in the study and it was found that there was a very significant decrease in free fatty acid (%) and peroxide value in both the oils after degumming. This may be due to the removal of FFA and other oxidation products in oil during degumming. Iodine value of oils also showed a slightly significant change during refining process. In order to study the effect of various thermal and non-thermal processing treatment, five major oils/fat consumed in India viz; mustard oil, soybean oil, palm oil, hydrogenated fat (vanaspati) and desi ghee (Nutrition Foundation of India, 2010) were selected. These oils/fats have different degree of unsaturation (double bonds) which will affect the trans isomerization process. Various thermal and non-thermal treatments were given to oil samples and the processed samples were then analysed for trans fatty acid, 3

4 peroxide value, anisidine value, TBA value, free fatty acid, refracrive index, iodine value and total polar compound. In order to assess the impact of frying on selected oils, raw, sliced Potato fingers were fried in the five selected commercial oil samples at 200 ºC -220ºC. During each frying cycle 100 g potato fingers were fried in 1000 ml oil samples till golden brown (for 10 min appx).. For 2 nd and 3 rd frying cycle fresh potato fingers were fried in the same oil and oil was for analysis was taken after every 2 nd frying. On comparing the relative percentage of TFAs during the frying sessions, a clear increase in TFAs was observed. The main trans fatty acid isomers found in palm oil and hydrogenated fat are elaidic acid (C18:1,t), in concentration, 0.45±0.12 g/100g and 6.13±0.36 g/100g respectively. In soybean oil, mustard oil and clarified butter, the predominant trans fatty acid present was linolenic acid in the concentration 0.32±0.04 g/100g, 0.39±0.08 g/100g and 2.68±0.13 g/100g, however mustard oil contain a good amount of elaidic acid also (0.33±0.10 g/100g). Among the five oils selected, maximum increase in trans fat was found in hydrogenated fat, followed by clarified butter. Soybean oil, palm oil and mustard oil have also shown a significant increase in trans fat content. In order to study the impact of heat treatment on trans fatty acid, 600 ml of oil was taken in a 1 litre beaker and kept on thermostat hot plate with continuous shaking. Heat treatment of each selected oil was carried out at 100 ºC, 200 ºC, 300 ºC and ºC for 1 hour. The temperature of oil was maintained with ±5ºC variation. Oil samples were procured after 1 hour heating at each temperature. The trans fatty acid composition of each oil before and after treatment was observed and it was found that no significant increase in TFA content occur at 100 ºC and 200 ºC temperature, however at higher temperature 300 ºC and 400 ºC trans fatty acid content increases significantly. The total trans fatty acid content in the oil samples varied from 2.57 to 19.20%. The most significant increase was found in hydrogenated fat. If each oil was observed separately, then in soybean oil and hydrogenated fat maximum increase was found in trans linoleic acid content, whereas mustard oil and clarified butter show significant increase in linolenic acid content. In palm oil, trans linolenic acid and palmitelaidic acid content increases more significantly. For the purpose of microwave treatment, 250ml each of the edible oil samples 4

5 was taken for treatment and treated in the microwave oven at 2450 MHz and 150 Watt power level for and 40 minutes. Analysis of trans fatty acids in treated samples showed a significant increase in trans fat content. Out of the five oils, maximum increase was found in hydrogenated fat and soybean oil, followed by clarified butter and palm oil. Mustard oil showed relatively less increase in trans fat level. It was also found that the increase in isomerization of fatty acid is proportional to the time of microwave treatment. Irradiation of samples was done at the Defence Food Research Laboratory, Mysore. The applied radiation doses were 2, 4, and 6 kgy by Gamma rays in γ-chamber 5000 at the dose rate of KGy/hr. In order to study the effect of Irradiation on trans fat formation, potato fingers were fried in the three oil samples (soybean, mustard and palm oil) and were processed at 2, 4 and 6 KGy at ambient temperature. Slight but statistically significant changes (P<0.05) of the main polyunsaturated trans fatty acids such as C18:2-9c,12c, C18:1-9n were observed in potato fries for each of the 3 oils selected with the irradiation dose of 4 kgy compared with the control sample, at a dose of 2 KGy, no significant change was observed in trans fatty acid content. Hence it can be concluded that irradiation induced the formation of TFAs in food when the absorbed dose exceeded 4 kgy. Among the three oils selected for study, maximum difference was observed in soybean oil followed by palm oil and then mustard oil. To study the effect of High Pressure processing on trans fat formation, the soybean, mustard and palm oil samples were processed at 200, 400 and 600 MPa pressure at ambient temperature (27-29 C ). Water and propellant glycol was used as coolant. Treatments on HPP were conducted at Defence Food Research Laboratory, Mysore. After processing the samples were analysed for trans fatty acids on gas chromatography. It was found that there is no significant difference in trans fat content by application of HPP in all the three oil. The F-Value obtained by application of one way ANOVA was 0.197, and for soybean, mustard and palm oils respectively which are below significance for p<0.5. If the increase in TFA content was compared for all the processing treatments then, it was found that TFA formation during frying was highest in all the oils. Among heat treatment and microwave, formation of trans fat is more pronounced in microwave treatment, with a significant rise in TFA after 10 mins of treatment, whereas in heating, 5

6 significant increase in trans fat was found after 200ºC temperature. Irradiation treatment shows slightly significant increase at the end of 6 KGy radiation, however high pressure processing did not affect the trans fat formation at all. If the increase in free fatty acid (% of oleic acid) was observed for all the processing treatment, it was found that frying treatment significantly increases the FFA value of oils by 0.86±0.21, 1.07±0.37, 1.21±0.31, 0.86±0.12 and 2.62±0.30 for soybean oil, mustard oil, palm oil, hydrogenated fat and clarified butter respectively. For microwave and heat treatment ffa value was found to increase slightly with more pronounced increase in microwave treatment (with co-efficient of variation ranging from 0.32 to 0.46) followed by heat treatment (0.25 to 0.38). among the five oil/fat selected maximum free fatty acid was formed in clarified butter. This may be attributed due to the presence of short chain fatty acids in it. Whereas hydrogenated fat shows least effect on FFA content during various processing. If the increase in total polar compound (% g/100g) was observed for all the processing treatment, it was found that frying treatment significantly increases the TPC value of oils by 1.53±0.11, 0.95±0.18, 0.89±0.11, 1.06±0.07and 1.47±0.21for soybean oil, mustard oil, palm oil, hydrogenated fat and clarified butter respectively, with a maximum increase in soybean oil (C.V ) followed by mustard oil (C.V ). In general, oils with higher levels of unsaturated fatty acids produced more polar compounds compared to the more saturated oils. Whereas hydrogenated fat shows least effect on FFA content during various processing. For microwave and heat treatment TPC value was found to increase. Irradiation and HPP have insignificant effect on total pola compound of oils. A very high significant change was observed in peroxide value of oils after frying, heating, microwave and irradiation treatment. Change was more pronounced in frying followed by microwave treatment. However, none of the sample exceeds the maximum permissible limit for peroxide value (10mEq/Kg of oil) during the entire processing. Maximum icrease in peroxide value was found in soybean oil followed by mustard oil. In mustard oil, palm oil and clarified butter, a slight decrease in peroxide value was observed during the last frying treatment. This decrease in peroxide value can be due to the conversion of primary oxidation products into secondary oxidation product. 6

7 The analysis of Iodine value of oils reveals that there was a drop of about 14,10, 7, 5, 7 degree for soybean oil, mustard oil, palm oil, hydrogenated fat and clarified butter respectively during frying. Whereas during heat treatment decrease in iodine value observed was 11, 8, 6, 3, and 5 for soybean oil, mustard oil, palm oil, hydrogenated fat and clarified butter respectively. Similar trend was observed for microwave treatment also. It was followed that the more saturated the oil is the lesser is the degradation of oil into secondary metabolites. As shown, the highest significant (p < 0.05) change in the IV was shown by the mustard oil and soybean oil, thus indicating that the highest decrease in double bonds occurred due to oxidative rancidity in the proposed frying media. This observation could be due to the presence of a high amount of PUFAs in frying medium 7

8 Conclusion GC and FTIR both methods were found equally suitable for trans fat analysis as no significant difference was found in the trans fat content, analysed by these two methods. Maximum trans fats were found in chocolate samples followed by cookies and bhujia samples. Mathri and laddoo samples were also found to be high in trans fat. Local food samples found to have more trans fat and free fatty acid content compared to branded samples. The trans fat was found to be increased significantly during the de-odorization of oil. Among the various thermal and non-thermal processing treatments, frying produced more trans fat and total polar compound followed by microwave and heat treatment, however high pressure processing does not increased the trans fat as well as TPC in oils. Frying showed a significant increase in Total polar compound (TPC) followed by microwave and heat treatment. Irradiation and HPP did not showed any significant increase in TPC. 8

9 APPENDIX I FTIR graph for triolein standard FTIR graph for trielaidin standard FTIR graphs for triolein and trielaidin standards 9

10 APPENDIX II GC chromatogram for Elaidic acid methyl ester GC chromatogram for Palmitelaidic acid methyl ester 10

11 GC chromatogram for linolenaidic acid methyl ester GC chromatogram for linoleic acid methyl ester 11

12 GC chromatogram for linoleic acid methyl ester 12

Overview of the food science behind fatty acid technology

Overview of the food science behind fatty acid technology Pamela J. White, Ph.D. Food Chemist/Scientist Food Science and Human Nutrition Dept. & Center for Crops Utilization Research Iowa State University