发酵果酒中高浓度花色苷影响DNS比色法检测还原糖的准确性,尤其发酵末期的果酒色深而含糖量低,增加了检测难度。该研究采用聚乙烯吡咯烷酮(polyvinylpyrrolidone,PVP)脱色、空白对照和样品稀释3种方法,减少花色苷对DNS比色法检测的干扰。结果发现PVP-K30具有吸附花色苷的作用,但无法完全排除颜色干扰。对于高花色苷样品,采用空白对照法检测结果更接近真实值,具体做法是选取200 μL待测样,添加150 μL DNS试剂,不经沸水浴作为空白对照。从排除色素干扰角度,稀释倍数越大花色苷干扰越小。而在果酒检测中,稀释倍数的确认须兼顾待测样品的初始还原糖浓度,研究发现存在最佳稀释倍数范围。100~200 g/L还原糖样品,稀释200倍最佳,50~100 g/L还原糖样品,稀释100倍以内为佳,还原糖10~50 g/L样品,50倍以内稀释准确性较好,说明从还原糖角度,稀释后样品含糖量在0.2~1 g/L时,检测效果较好。因此,深色果酒还原糖的检测建议结合空白对照和样品稀释两种操作提高准确性。该研究进一步以酿酒酵母CECA发酵蓝莓酒为例,采用改进DNS法监测4个发酵时期还原糖含量,检测5个还原糖代谢相关基因表达量。发酵前期GLK1和HXK1表达水平较高,而中期、中后期HXK2、PFK1和PFK2均呈高表达水平,后期5个基因表达水平均较低。还原糖含量与GLK1、HXK1基因表达量正相关。该研究为深色果酒发酵中还原糖快速精准监测提供方法参考,并探索蓝莓酒发酵中还原糖相关基因表达与糖代谢关联,对提升产品质量和改进监测方法具有理论研究价值。
High concentrations of anthocyanins in fermented fruit wine affect the accuracy of reducing sugar detection by DNS colorimetry.Particularly in the late stage of fermentation, the fruit wine is dark in color and has a low sugar content, which increases the difficulty of detection.In this study, three methods, decolorization with polyvinylpyrrolidone (PVP), blank control, and sample dilution, were used to reduce the interference of anthocyanins on DNS colorimetry.It was found that PVP-K30 has the function of adsorbing anthocyanins, but it could not completely eliminate color interference.For samples with high anthocyanin content, the detection result obtained by the blank control method was closer to the real value.The specific approach was to select 200 μL of the sample to be tested, add 150 μL of DNS reagent, and use it without boiling treatment as a blank control.From the perspective of eliminating pigment interference, the greater the dilution factor, the smaller the anthocyanin interference.In the detection of fruit wine, the confirmation of the dilution factor also needed to consider the initial reducing sugar concentration of the sample.The study found that there is an optimal dilution factor range.For samples with 100-200 g/L reducing sugar, a 200-fold dilution was the best.For samples with 50-100 g/L reducing sugar, a dilution within 100 times was better.For samples with 10-50 g/L reducing sugar, a dilution within 50 times had better accuracy.It indicated that from the perspective of reducing sugar when the sugar content of the diluted sample was 0.2-1 g/L, the detection effect was better.Therefore, for the detection of reducing sugar in dark fruit wine, it was recommended to combine blank control and sample dilution operations to improve accuracy.This study further took blueberry wine fermented by Saccharomyces cerevisiae CECA as an example and used the improved DNS method to monitor the reducing sugar content in four fermentation periods.The expression levels of five genes related to reducing sugar metabolism were analyzed.In the early stage, the expression levels of GLK1 and HXK1 were relatively high.In the middle and late stages, HXK2, PFK1, and PFK2 were all highly expressed.In the late stage, the expression levels of all five genes were relatively low.The reducing sugar content was positively correlated with the expression levels of GLK1 and HXK1 genes.This study provides a methodological reference for the rapid and accurate monitoring of reducing sugar in the fermentation of dark fruit wine and explores the association between the expression of reducing sugar-related genes and sugar metabolism in blueberry wine fermentation, which has theoretical research value for improving product quality and monitoring approaches.
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