Lectin is one of the major anti-nutritional factors in soybeans and inhibits digestion of dietary protein. Here, an absolute quantification method was developed to detect lectin using synthetic peptide 183TTSWDLANNK192 as reference standard and corresponding isotope labeled peptide TTSWDLANNK (Alanine-13C3,15N) as internal standard to normalize results. After the ground soybeans and soy products were defatted with n-hexane and extracted with extraction buffer, the crude protein extract was digested on filter membrane by trypsin. Further, the enzymatic hydrolysis peptides were quantified using liquid chromatography-tandem mass spectrometry. The synthetic reference peptide showed a detection limit of 0.27 ng/mL and a linear relationship in the range of 3.2-1000 ng/mL (r2 > 0.997). Correspondingly, the detect limit of lectin in soybean samples was 35.5 μg/g. The results showed that the recoveries of the lectin in spiked samples ranged from 80.9% to 108.7% with intra-day precisions (% CV) less than 9%. The method was successfully used to evaluate lectin levels in hundreds of soybean seeds from different varieties and soy products from different soybean processing techniques. Furthermore, the method may provide a potential application as a general method for the ultrasensitive detection of various protein anti-nutritional factors in food.
Keywords: Anti-nutritional factors; Lectin; Liquid chromatography-tandem mass spectrometry; Protein absolute quantification; Soybean.
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