Studies On The Application Of Nitrogen Fraction Method On The Degree Of Hydrolysis

On the basis of the previous studies, nitrogen fration method was put forward for thedegree of hydrolysis. This method used Kjeldahl determination through the determination oftotal nitrogen, dissolved nitrogen, pure protein nitrogen, amino nitrogen, as well as high,medium and low molecular weight nitrogen in the dissolved nitrogen, to characterize thedegree of protein hydrolysis. This method needed single equipment and was simple, intuitive,reliable, suitable for rapid detection of factory, not be influenced by the samplecharacterization.The content of of total protein, soluble protein, pure protein, pure protein in solubleprotein,and amino nitrogen were determined before and after the hydrolysis for the degree ofhydrolysis. Besides, the formaldehyde titration,TCA,GPC and the SDS-PAGE were also usedfor characterization of protein degradation. The results showed that,the results determined bynitrogen fration method was consistent with that by TCA method,formaldehyde titrationmethod，SDS-PAGE and GPC,and was more accurate than the formaldehyde titration methodin expressing the reality of hydrolysis; From the GPC map we can infer that, the molecularweight of TCA soluble protein was about10,000Da.Introducing the Lundin fraction method for the protein fraction in soluable protein,systematic studied the changes of nitrogenous substance in the process of hydrolysis anddetermined protein distinguish range of the tannins acid and phosphomolybdic acid. Theresults showed that, nitrogen fraction method was intuitive and reliable, consistent withSDS-PAGE; Mass spectrometry analysis showed that, after dealt with tannins acids,themolecular weight of peptides in enzyme solution was below2300Da, while2000Da wasobtained after being treated with phosphomolybdic acidUsing NSI and angiotensin converting enzyme (ACE) inhibition activity (IA) forsececting the best hydrolysis conditions for high NSI, ACE inhibition activity peptides. Andnitrogen fraction method characterized protein hydrolysis degree. Preliminary compared thebitterness of peptides from high temperature soybean meal and SPI by Two PairedComparison method. The results show that under the optimum conditions for NSI（62.97%）,the content of total protein was41.21%, soluble protein was37.23%, pure proteinwas4.94%,amino nitrogen was0.64%, high molecular weight nitrogen was11.54%, mediummolecular weight nitrogen was5.92%and low molecular weight nitrogen was82.63%, at thiscondition, ACE inhibition activity was18.37%; Under the optimum conditions for ACEinhibition activity（77.55%），the content of total protein was42.22%, soluble protein was 30.94%, pure protein was7.40%, amino nitrogen was0.59%, high molecular weight nitrogenwas4.04%, medium molecular weight nitrogen was4.78%and low molecular weightnitrogen was91.17%, NSI was52.33%; When enzyme quantity was1500U/g, pH8.5,temperature60℃, substrate concentration4%, NSI was61.68%, ACE inhibition activity75.51%, so this hydrolysis condition was the best condition.Under this condition, the totalprotein was41.35%, soluble protein was36.47%, pure protein was5.23%，amino nitrogenwas0.62%, high molecular weight nitrogen was8.65%, medium molecular weight nitrogenwas5.24%, low molecular weight nitrogen was86.11%.Compared with SPI hydrolysate, the bitterness of high temperature hydrolysate was low.