| Soybean trypsin inhibitor is one of the major anti-nutritional factor in soybeans. The anti-nutritive roles of STI mainly reflect in inhibiting the growth of animal and causing pancreatic hyperplasia, pancreatic enlargement. But the mechanism of the pancreatic enlargement is not entirely clear. In this experiment, we made soybean trypsin inhibitor as the research object, and researched on whether oxygen free radical can result in pancreatic injury.In this paper, the raw material was defatted soybean meal. The crude extract of the soybean trypsin inhibitor was obtained by the extraction of phosphoric acid buffer(pH7.6), thermal denaturation(65℃) and ammonium sulfate precipitation. After purified by DE-52ion exchange, affinity chromatography and sephadex G-75gel filtration, STI with purification factor of73.85was obtained. The purified STI gave two protein bands in SDS-PAGE electrophoresis. Its molecular weights were estimated to be21.92kD and20.04kD.Pre-purified STI was added to mice feeds, and study on the effect of STI on pancreatic oxygen free radical levels in mice. The principal means was as follows:weight18±2g of clean grade male Kunming mice were pre-fed with normal diet for3days, and then were randomly divided into three groups. The first group as a control group was fed a normal diet; the second group was fed the feed containing the STI, the dosage of STI was200mg/100g feed; the third group was fed with STI+VC, the amount of STI, VC were200mg/100g and1500mg/kg. After pre-feeding period, began to experiment. The trial period was divided into five time points:1w,2w,3w,4w,5w; at the end of the experiment, the mice were fasted12hours, then taken blood samples from the eyes and were sacrificed by cervical dislocation taken pancreatic visceral samples. Experimental results showed that:1The effect of soybean trypsin inhibitor on mice weight gain and Viscera indexAt each growth period, daily weight gain between groups:the first group> the third group>the second group. In the first four weeks, the daily weight gain of the second group and the third group compared with the first group showed the significant difference (P<0.05), at the fifth weeks, there were no significant differences between the three groups (P>0.05). In addition, STI can result in mice pancreatic enlargement, in the first, third week and around, the pancreas index of the second group compared with the first group showed significant difference (P<0.05), and at the third week, STI had the most significant effect. That showed the soybean trypsin inhibitor can inhibit the growth of mice, result in pancreatic enlargement and VC can interfere in the effect of STI on organisms.2The effect of soybean trypsin inhibitor on oxidative and antioxidative indexes of mice serum and pancreasIn five weeks trial period, the content of MDA in mice serum and pancreas showed the ascendant trend, and at the third weeks, there were significant differences among the three groups (P<0.05). In addition, the activity of CAT, GSH-Px, T-SOD in the second group was significantly lower than that of the first group (P<0.05), hydroxyl radical inhibition ability, anti superoxide anion capacity were significantly reduced, causing the T-AOC of the second group was significantly lower than that of the first group (P<0.05). These results showed that STI can make free radicals in mice increase, leading to the oxidative stress.3The effect of soybean trypsin inhibitor on mice pancreatic exocrine functionSTI can influence the pancreatic gland secretion of digestive enzymes and change pancreatic exocrine function of the mice. In the period of growth, the trypsin activity of each group showed the significant differences (P<0.05), the second and the third group were significantly lower than that in the first group(P <0.05). While the activity of the lipase and amylase in the second and the third group were significantly higher than that in the first group (P<0.05), and it reached a significant level between each group (P<0.05). The results showed that STI can inhibit the activity of the trypsin and increase the activity of the lipase and amylase.4Correlation analysis between each index in mice serum and pancreasIn serum, the correlation coefficients between the content of MDA and total antioxidant capacity, the activity of T-SOD were-0.431,-0.873. They reached extremely significant level (P<0.01), and the correlation coefficients between MDA and CAT activity was-0.352, reached a significant level (P<0.05). In pancreas, the correlation coefficients between the content of MDA and anti-superoxide anion ability, the activity of T-SOD were-0.705,-0.832, they reached extremely significant level (P<0.01).This showed STI inhibited the activity of antioxidant enzymes by increasing the content of free radical in the mice and resulted in the oxidative stress. In the serum of mice, the correlation coefficients between the activity of trypsin and the activity of lipase, the activity of amylase were-0.615,-0.431.They reached the extremely significant level (P<0.01). In pancreas, the correlation coefficients between the activity of trypsin and the activity of lipase, the activity of amylase were-0.595,-0.570. They reached the extremely significant level (P <0.01). This meant the combination of STI and trypsin resulted in pancreatic enlargement. The pancreas compensatory secreted the trypsin, meanwhile, the secretion of lipase and amylase was increased. In mice serum, the correlation coefficients between the activity of trypsin and T-AOC, anti-superoxide anion, the ability of inhibiting hydroxyl radical, CAT, T-SOD, MDA were0.860,0.670,0.635,0.845,0.627.-0.691, all of them reached the extremely significant level (P<0.01). In the pancreas, the correlation coefficients between the activity of trypsin and anti-superoxide anion, the ability of inhibiting hydroxyl radical, CAT, GSH-Px, T-SOD, MDA were0.441,0.596,0.622,0.673,0.456,-0.640, they all reached the extremely significant level(P<0.01). The correlation between the activity of trypsin and T-AOC was0.376,reached the significant level(P<0.05). The results showed that STI leaded to the oxidative stress by the increasing of the free radicals, and what’s more, It induced the oxidative damage of the pancreas, and then effected the secretion of the pancreas. |
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