The Effect of heavy Application of Nitrogen on Flavonoid Metabolism in Apple Skin

苹果果实（cv.以1972年起不施氮的-N区（-N）和每年4月施氮2kg/株的+N区（+N）为材料，研究了苹果果皮黄酮代谢的变化。采收时，-N处理果实着色率高于+N处理.测定了苹果皮中的黄酮类化合物：花色苷（AN）、无色花色苷（LA）和槲皮素苷（QG）。着色率达90%以上的果皮中总黄酮含量为-N果实383 nmol/cm ^2，+N果实340 nmol/cm ^2。各化合物占总黄酮的比例如下：-N果实中AN 33%、LA 13%和QG 54%;+N果实中AN 23%、LA 12%和QG 65%。结果表明，大量施氮抑制了硝酸还原酶的合成，促进了QG的合成。为了弄清这一现象的原因，将果实用玻璃滤纸吸收处理液粘在一起，然后用聚乙烯薄膜覆盖，在25 ℃下培养12小时。将果实在15 ℃连续光照（15 klux）下孵育72小时。使用硫酸铵、天冬酰胺（Asn）和多胺作为处理溶液。这些化合物的1 mM溶液抑制苹果皮中AN的发育。苯丙氨酸（Phe），二氢槲皮素（DQ）和t-肉桂酸（tC）作为AN和QG生物合成的前体进行了测试。Phe促进了-N果实中AN的发育，但对+N果实中AN的发育没有促进作用. DQ和tC促进+N果实中AN的发育。但这些化合物对-N果实中AN的发育没有影响，添加DQ和tC可减轻Asn对AN发育的抑制作用。

Apple fruits (cv. Jonathan) of -N plot (-N) receiving no nitrogen application from 1972 and of +N plot (+N) receiving 2kg N/tree in April of every year were investigated for flavonoid metabolism in apple skin. At harvest time, coloration rate of -N fruit was better than that of +N fruit. As flavonoid compounds in apple skin, anthocyanin (AN), leucoanthocyanin (LA) and quercetin glycosides (QG) were determined. The concentrations of total flavonoid compounds in fruit skin colored more than 90% were 383 nmol/cm^2 skin in -N fruit and 340 nmol/cm^2 skin in +N fruit. The ratios of each compound to total flavonoid was as follow ; AN 33%, LA 13% and QG 54% in -N fruit and AN 23% , LA 12% and QG 65% in +N fruit. These results suggest that heavy application of nitrogen inhibits the biosynthesis of AN and stimulates the biosynthesis of QG.To clarify the cause of this phenomenon, fruit was cohered with glass filter paper absorbing treatment solution, then lapped with polyethylene film and incubated for 12hr at 25ﾟC.After removing glass filter paper, the fruits were incubated under continuous light (15 klux) for 72hr at 15ﾟC.Ammonium sulfate, asparagine (Asn) and polyamines were used as treatment solutions. One mM solution of these compounds inhibited AN development in apple skin. Phenylalanine (Phe), dihydroquercetin (DQ) and t-cinnamic acid (tC) as precursors of AN and QG biosynthesis were tested. Phe stimulated AN development in -N fruit, but it did not stimulate AN development in +N fruit. DQ and tC stimulated AN development in +N fruit. But these compounds had no effect to AN development in -N fruit.The inhibitory effect of Asn on AN development was alleviated by addition of DQ and tC.