The Purification And Identification Of Umami Peptides From Soy Sauce And The Study Of Its Taste Characteristic

Soy sauce is one of the necessary seasoning of daily life for Chinese. The umami andkokumi tastes of soy sauce lead to its important role in the industry of seasoning. And thus thestudy of the umami peptide of soy sauce contributes to the understanding of the basicmaterials of soy sauce and provides theories and methods for the development of seasoning.In this study, the performance and separation characteristics of four macroporous resins(SP-825, HP-20, XAD-16 and HP-2MG) for purifying umami peptides from soy sauce wereexamined. Results showed that four resins could enrich the peptides of soy sauce.Furthermore, the particle diffusion kinetics model was suitable for describing the wholeexothermic adsorption process on the SP-825 and HP-20 resins, while thepseudo-second-order kinetics model accurately described the behaviour of XAD-16 andHP-2MGL resins. The adsorption processes of the peptides on the four resins followed theFreundlich model. The XAD-16 resin was the most effective resin for the enrichment ofumami peptides due to its high adsorption and total recovery capacities for peptide. After atreatment with gradient elution on the XAD-16 resin columns, the profiles of the peptides andamino acids from soy sauce as well as the taste were analysed. Interestingly, the content ofumami amino acids from peptides of XAD-0% fraction was higher than other fractions. Inaddition, the umami taste of XAD-0% fraction was better than other fractions. Also the tasteof XAD-0% fraction was better than its compositions of artificial free amino acids or totalamino acids. And thus the umami peptides were enriched in the deionized water fraction.The XAD-0% fraction was further purified by medium pressure chromatography(MPLC),reverse high-performance liquid chromatography(RP-HPLC) and LC-MS/MS analysis. Afterseparation by MPLC, XAD-0% was divided into four fractions(M-1, M-2, M-3 and M-4),and the taste of M-1 fraction was better than other fractions with the highest content of umamiamino acids in peptides(38.64 mg/g) and it was 51.73% of the total umami amino acids inpeptides. Besides, the taste of M-1 fraction was better than its compositions of aritificial freeamino acids or total amino acids, so it was likely that the umami peptides would exist in M-1fraction. After separation of M-1 fraction by RP-HPLC, the umami taste of HPLC-3 fractionwas higher than other three fractions. UPLC-MS/MS analysis reveled the amino acidsequences of the five new umami peptides from HPLC-3 fraction were ALPEEV, LPEEV,AQALQAQA, EQQQQ and EAGIQ that originated from glycinin A1 b B2-445, glycininA1 b B2-445, cobyric acid synthase, leucine--t RNA ligase and lglycoproteinglucosyltransferase, respectively.Five peptides(ALPEEV, LPEEV, AQALQAQA, EQQQQ and EAGIQ) were synthetizedafter purified and identified from HPLC-3 fraction. Furthermore, the taste and the effect ofumami-enhancement were detected with the synthetic peptides. The result showed thatpeptides LPEEV, AQALQAQA and EQQQQ were also tasted as umami while other peptideswere not. The threshold value of peptides ALPEEV, LPEEV, AQALQAQA, EQQQQ andEAGIQ were about 0.76 mmol/L, 0.43 mmol/L, 1.25 mmol/L, 0.76 mmol/L and 0.97 mmol/L,respectively. In the MSG + peptides model, peptides ALPEEV, EAGIQ and LPEEV had theeffect of umami-enhancement with the threshold about 1.52 mmol/L, 1.94 mmol/L and 3.41mmol/L, respectively. When the concentration was 2 g/L, peptide ALPEEV, LPEEV andEAGIQ had the best umami-enhancement effect for MSG. And therefore, the umami aminoacid(Asp or Glu) did not determine the umami taste of peptides, also umami peptides mightnot have umami amino acids, besides, the position of umami amino acids in peptides was notrelated to the umami taste.