烟台近海3种贝类中呈味核苷酸和氨基酸的测定及比较分析

摘要

对水产品中呈味核苷酸和氨基酸的高效液相色谱检测方法进行了优化,并对烟台近海长牡蛎(Ostrea gigas)、栉孔扇贝(Chlamys farreri)和中国蛤蜊(Mactra chinensis)3种贝类中呈味核苷酸和氨基酸的含量进行了比较分析,从而为开发贝类复合调味料提供科学依据。以纯水提取呈味物质,采用DABS-Cl柱前衍生反相高效液相色谱法测定游离氨基酸,流动相为17 mmoL/L柠檬酸溶液(pH 6.4,含4%DMF)和乙腈溶液(含4%DMF),梯度洗脱,检测波长436 nm。反相离子对色谱法测定呈味核苷酸, pH 4.5的磷酸二氢钾溶液和乙腈溶液为流动相,磷酸二氢钾溶液内含5 mmoL/L四甲基氢氧化氨(TMAOH),梯度洗脱,检测波长254 nm。方法加标回收率为90.2%~108%,相对标准偏差(RSD)为0.7%~7.1%。准确度、精密度均满足分析要求。测定结果显示：长牡蛎中牛磺酸、丙氨酸和谷氨酸含量较高,栉孔扇贝中甘氨酸、牛磺酸和丙氨酸含量较高,中国蛤蜊含有较多的甘氨酸、牛磺酸和丙氨酸；3种贝类中,丙氨酸、谷氨酸的TAV均大于1,栉孔扇贝和中国蛤蜊中,甘氨酸的TAV大于1,对呈味具有显著贡献。呈味核苷酸中,长牡蛎中IMP含量最高,占总呈味核苷酸的80%,其次是GMP。中国蛤蜊中AMP含量最高,其次是IMP。而栉孔扇贝中仅检测到IMP。3种贝类中呈味核苷酸含量虽然都较低, TAV未超过1,但核苷酸之间、核苷酸和氨基酸之间的协同增效作用,会对贝类的呈味产生重要影响。本研究通过对烟台近海3种贝类中各类鲜味物质和多种辅助呈味物质的含量、比例关系及其相互作用规律进行探讨,旨在为充分利用贝类营养氨基本作为复合调味料及其功能性开发和评价提供科学依据。%A high-performance liquid chromatography (HPLC) method was optimized for the determination of the free amino acids and flavor-enhancer nucleotides in aquatic products. The flavor-enhancer nucleotides and amino acids of Pacific oyster (Crassostrea gigas), Farrer’s scallop (Chlamys farreri) and sunray surf clam (Mactra chinensis) were determined to provide basic data for exploiting shellfish seasoning compounds. After sample preparation using purified water, free amino acids were determined by dabsyl chloride (Dabs-Cl) pre-column derivatization and reversed-phase HPLC. The best separation was obtained using a 17 mmol/L citric acid solution (pH 6.4, containing 4%dimethylfor-mamide (DMF)) and acetonitrile (containing 4%DMF), gradient elution and a detection wavelength of 436 nm. Flavor nucleotides were determined by reversed-phase ion-pair HPLC, with a mobile phase of 0.02 mol/L KH2PO4 (pH 4.5, 5 mmol/L tetramethylammonium hydroxide (TMAOH)) and acetonitrile, and a detection wavelength of 254 nm. This method displayed better precision and higher accuracy in meeting the analytical requirements. Recoveries varied from 90.2%to 108%and RSD varied from 0.7%to 7.1%. The results showed that high levels of taurine, alanine and glu-tamic acid were found in Pacific oyster; glycine, taurine and alanine were the principal free amino acids in Farrer’s scallop and high percentages of glycine, taurine and alanine were detected in sunray surf clam. The taste activity values (TAVs) of alanine and glutamic acid in the three shellfish were >1; only the TAVs of glycine in Farrer’s scallop and sunray surf clam were>1, indicating that they make a significant contribution to taste. Regarding flavor-enhancing nu-cleotides, Pacific oyster contained more inosine 5′-monophosphate (IMP), accounting for 80%, while Farrer’s scallop contained the highest adenosine monophosphate (AMP) level followed by IMP;sunray surf clam contained IMP only. Although the concentration of flavor-enhancing nucleotides in three shellfish were low and TAVs were<1, the synergis-tic effect between nucleotides and amino acids has an important impact on shellfish flavor. Pacific oyster, Farrer’s scal-lop and sunray surf clam from offshore Yantai are rich in taste, and the flavor-enhancing nucleotides and amino acids, such as taurine, are valuable for human health. The study also provides basic data for the extraction of active substances for the preparation of seafood seasoning compounds.