山葡萄种质资源原花青素动态含量变化及提取降解

山葡萄种质资源原花青素动态含量变化及提取、降解研究

STUDIES OF EXTRACTION AND DEGRADATION IN PROCYANIDINE，AND THE DYNAMIC CONTENTS IN GERMPLASM RESOURCES ON VITIS AMURENSIS RUPR

作者：王新伟　　导师：沈育杰

中国农业科学院　　经济植物资源评价与利用2011届硕士

摘　要

本文对山葡萄籽中原花青素提取、降解进行了研究，优化了工艺路线。以40份山葡萄种质资源为试材，对山葡萄各个重要时期的叶片、茎皮、果实中的原花青素动态含量变化进行了测定，分析了原花青素在山葡萄各组织器官中的变化规律和山葡萄种质资源间原花青素含量和分布的特点与差异性，主要结果如下：

1.　确定了山葡萄黄烷-3-醇的HPLC洗脱程序：流动相配比：A：1%乙酸；B：80%乙腈，1%乙酸，0min→5min→15min→25min→30min→35min，动相A对应的梯度为95%→70%→50%→40%→95%→95%。

2.　原花青素的最佳提取工艺为：以乙醇为提取剂，温度60℃，浓度70%，时间为120min，料液比为1:5，提取率为3.12%。

3.　溶剂分离法分离制备原花青素高聚体，综合聚合度与过氧化指标，确定优化降解工艺为：乙酸为降解介质，料液比6:5，降解时间20min，降解温度70℃，聚合度从6.9下降到4.8，聚合度下降2.1。

4.　山葡萄种质资源叶片中原花青素动态含量变化趋势为：萌芽期→花后一周，缓慢上升；花后一周→浆果膨大期，含量急剧上升，并达到最大值；浆果膨大期→转色期，含量迅速下降；转色期→成熟期，含量上升。

5.　茎皮原花青素动态含量上升变化趋势为：萌芽期→花后一周，缓慢上升；花后一周→浆果膨大期，含量急剧上升，并达到最大值；浆果膨大期→浆果转色期→浆果成熟期，含量迅速下降。

6.　种子中原花青素动态含量变化趋势为：花后一周→浆果膨大期→浆果转色期，含量迅速上升并在转色期达到最大值；浆果转色期→浆果成熟期，含量迅速下降。成熟期含量最高的种质资源为：4N1,4N2，燕山。

7.　果皮中原花青素动态含量变化趋势为：花后一周→浆果膨大期，含量很低，缓慢上升；浆果膨大期→浆果转色期，含量迅速升高；浆果转色期→浆果成熟期，含量急剧上升，并达到顶峰。浆果成熟期含量最高的种质资源为：086034,8558814,8558831。

8.　山葡萄成熟果实果皮中原花青素含量高于种子，且差异性极为显著，与欧亚种酿酒葡萄相比具有明显的种间差异性。

关键词　山葡萄　原花青素　黄烷-3-醇　提取　降解　动态含量变化

Abstract

By studying conditions of extraction of proanthocyanidins from the vitis grape seeds after fermentation，the process of separations was optimized. The changes on the concentration of proanthocyanidins in leaves,barka and fruita was studied based on forty vitis amurensis germplasm resource with different developmental stages，the variation of proanthocyanidins with various organs and various grapes species. The preliminary conclusions as followed：

1.　The extraction of flvan-3-ols process was determined：mobile phase ratio was A:1% acetic acid ;B:80% acetonitrile,1% acetic acid,0min→5min→15min→25min→30min→35min，the mobile phase gradient of the corresponding to 95%→70%→50%→40%→95%→95%.

2.　The optimum condition of the proanthocyanidins extraction were determined：temperature 60℃，concentration of alcohol 70%，extraction time 120min，rate of solution to solid 5:1.Ratio of extraction was 3.12%.

3.　Respectively used hydrochloric acid and acetic acid as acid hydrolysis medium,this experiment studied the acid hydrolysis technology for procyanidin high polymer,determined the best acid hydrolysis technology is that acetic acid concentration－55%,hydrolysis time－20min ,hydrolysis temperature－70℃. In this experimental condition,the procyanidins high polymer units decreased from 6.9 to 4.8,dropped 2.1 units by test.

4.　The trends of the procyanidin dynamic content in leaves are that: procyanidin concentration increased slowly from budding to flowering;the content of procyanidin sharply rise in berries growing stage and up to maximum;the period of berry growth and turn color，the content declined rapidly when berry maturity.

5.　The dynamic content of procyanidin in barks are that：from budding to flowering is increase slowly；in the period of berries growing the procyanidin sharply rise and up to maximum；the content declined rapidly when berry maturity.

6.　The procyanidin trends in grape seeds are：from flowering to berry growing to turn color， the content are still increase and up to maximum in turn color period； until maturity the concentration of procyanidin declined rapidly. The higher content procyanidin species in berry maturity are：4N1,4N2，Yanshan.

7.　The trends in peel are：from flowering to berry growing，the concentration is low and rise；the content rise rapidly in turn color period；from turn color to maturity the procyanidin sharply increased and reach the peak. The higher content procyanidin species in berry maturity are：086034,8558814,8558831.

The content of procyanidin in pulp during the whole fruit development is very low.

8.　The content of procyanidin in vitis grape peel is significant higher than seeds in berry maturity,and compared with the Eurasian grape species have obvious differences.

Key words　Vitis amurensis　Procyanidin　Flan-3-ols　Abstraction　Degradation Dynamic