Lipase; SDS-PAGE; lipolysis; 4-MU butyrate; hydrolytic rancidity; fluorescent assay
Lipase is the major protein in rice that catalyzes endogenous hydrolysis of lipids. Study on lipase activity is crucial in rice bran utilization, as it causes rancidity, severely affects the product quality and tarnishes its economic value. This experiment aimed to isolate lipase from different rice lines' tissues, detect its activity and determine its molecular weight. Proteins were extracted from grains of Bd192, Kasalath and 2514 rice lines by grinding, buffering and centrifuging. Extracted proteins were separated with SDS-PAGE and lipase activity was detected using 4-MU Butyrate.
Protein bands were visualized with Coomassie staining and molecular weight of lipase was determined by comparison with protein ladder. Bright UV bands were observed in all leaves but not grains samples. The molecular weights of lipase was approximately 36.7kDa. In conclusion, lipase was present in leaves samples of all rice lines but not in grains.
[...] Bright UV bands were observed in all leaves but not grains samples. The molecular weights of lipase was approximately 36.7 kDa. In conclusion, lipase was present in leaves samples of all rice lines but not in grains. INTRODUCTION: Brown rice and milled white rice are staple food for most part of the world's population, especially the Asians (Sugiyama et al. 2003). During rice polishing, the testa and pericarp of grains, which is mainly comprised of bran, are removed. With such enormous demands in standard diet, rice bran are produced abundantly. [...]
[...] Figure Image of Coomassie stained of SDS-PAGE for leaves and grains of different rice lines. Figure Protein ladder bands with molecular weight values. Table Distance migrated by bands, molecular weights and log [distance] of bands. Figure Graph of log [distance migrated] (units) against molecular weight of protein bands (kDa). Calculations: Given that lipase bands for all leaves samples migrated approximately 5.4 cm from initial well, Log [distance]=- 0.0076 (MWlipase)+ 0.9299 log [ 5.4 0.0076 (MWlipase)+ 0.9299 MWlipase = 30.0 kDa Given that E. [...]
[...] Silva MA, Sanches C & Amante ER 2006, ‘Prevention of hydrolytic rancidity in rice bran', Journal of Food Engineering, vol no pp. 487-491. Sugiyama Tang AC, Wakaki Y & Koyama W 2003, ‘Glycemic index of single and mixed meal foods among common Japanese foods with white rice as a reference food', European Journal of Clinical Nutrition, vol pp. 743- 752. Vijayakumar KR & Gowda LR 2012, ‘Temporal expression profiling of lipase during germination and rice caryopsis development', Plant Physiology and Biochemistry, vol pp. 245-253. [...]
[...] Rice brans contains many active enzymes, particularly lipase. Lipase (E. C ) catalyzes breakdown of triglycerides present as aqueous emulsions into free fatty acids and glycerol (Lee et al. 2003). Two isoforms of lipase have been reported- lipase type I and type II, with type II being the major class of lipase in brans (Vijayakumar & Gowda 2013). Lipase are localized mainly in testa cross layer while oil is in aleurone and germ layers. Rice processing brings the normally isolated substrate and enzyme together and initiates lipid deterioration, leading to hydrolytic rancidity of brans that shortens shelf life (Monsoor et al. [...]
[...] 1986). The weak fluorescent signals of bands in Figure 2 might be caused by presence of contaminants such as SDS that inhibited lipase activity. This could be avoided by first purifying lipase from gel before staining with substrate. The use of 4-MU butyrate was also subjected to false positive, as this compound could also be cleaved by non-lipolytic esterase and fluorescent. This could be rectify by substituting with MUH which was more resistant to non-specific attack (Gilham & Lehner 2005). [...]
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