| Capsaicin is the main component of hot pepper, It is not only evaluation of pepper fruit quality index, and widely used in food, medicine, agriculture and other fields, but due to its extremely low yield, the international market is very tight. Therefore, the study of capsaicin, especially how to improve the yield of capsaicin has become a hotspot and focus of many researchers. At present, there are few studies on the metabolic regulation of capsaicin in pepper fruits, and the research results are very contradictory. In order to improve the content of capsaicin in pepper, the effects of different cultivation conditions on the synthesis of capsaicin in pepper fruits were studied. But cultivation conditions is a comprehensive factor, which is difficult to achieve. Therefore, the paper studies the formation and distribution of capsaicin in pepper plants from the physiological point of view, the contents of the enzyme, the endogenous hormone, the synthetic intermediate and the competition. The effects of exogenous ployamine spraying on the synthesis and metabolism of capsaicin, and to provide a theoretical basis for improving the content of capsaicin in pepper, and to find effective methods to improve the content of capsaicin in pepper. The main results are as follows:l.The Capsaicin chromatographic conditions HPLC method were as follows:Waters Nov a-Pak(?) C18 column (3.9mm × 150mm,4μm); the column temperature was 35 ℃; the detect ion wavelength was 281 nm; the flow rate was 1.0 mL · min-1; A mobile phase was acetonitril e; B mobile phase was 0.1%(V/V) phosphate buffer solution. When A:B was 40:60 (V/V ), the injection amount was 25 μL, Capsaicin and dihydro capsaicin can be completely separat ed from each other without interference,and repetition rate well.2.Characteristics of capsaicin accumulation in pepper fruitsThe capsaicin were all detected in the three Pepper varieties, and only in placenta and Pulp, but in the seeds and leaves. The Capsaicin content in placenta were higher.In Niujiao pepper L-10 and Shen pepper 4th, during the whole fruit development stage, the accumulation of capsaicin was a single peak, and the accumulation peak of 44d and 52d were respectively. This period is also the fruit turns red. Therefore, Optimal harvest time was fifty-two days and forty-four days after flowering. If premature, fruit development is not complete, capsaicin content is low, quality decline; too late, although the high yield, but capsaicin content decreased, while fruit softening, decline in quality.Among the tested varieties of pepper NiujiaoL-10, pepper fruit on the plants of a position higher, fruit capsaicin content is higher.3.Relationship between metabolic enzymes and the accumulation of capsaicin in pepper fruits.The experiment with Niujiao L-10 as experimental material. The results show that pericarp and placenta PAL enzyme activity and gene expression rate showed a single peak curve and peak (after anthesis 44d) early to capsaicin (after anthesis 52d), and the accumulation of capsaicin was significantly and positively correlated. The changes of Ca4H activity in the 36d and 68D, and the expression amount of the enzyme gene was the highest in the early stage after anthesis, and the accumulation of and 44d in the early stage. CS activity, gene expression almost exactly the same curve with the same period capsaicin content. Its activity and capsaicin content was significantly positive correlation. Thus, the first key enzyme of capsaicin synthesis was CS, followed by PAL.Capsaicin Accumulation and POD activity showed a significant negative correlation, but didn’t reach significant levels. Inferred, POD involved capsaicin degradation.4.HPLC detection system of the Simple phenols was determined:At 6280 nm wavelengt h UV detection, mobile phase of acetonitrile and 1% aqueous acetic acid=15:85 ((V/V)), th e column temperature was 35 ℃, the flow rate of 1.0 mL · min-1, the injection volume 15uL, various phenols can be completely separated from each other without interference,and repetiti on rate well.5.Relationship between metabolic intermediates of capsaicin biosynthesis and accumulation of capsaicin.In the tested the Niujiao L-10, in synthetic capsaicin early, detected a high level of cinnamic acid, p-coumaric acid and vanillin, with the development of pepper fruit, which cinnamic acid content decreased gradually, p-coumaric acid and vanillin in fruit development in late was not detected, and ferulic acid only in fruit development of the 52d-84d detected. From the change of total phenol content, the content of total phenol in the early stage of capsaicin was high, and the content of the content was low. It is indicated that the early stage of the simple phenols, which can provide a lot of synthesis of the accumulation of capsaicin6. Relationship between metabolic competition of capsaicin biosynthesis and accumulation of capsaicin.The results showed that Lignin content in the fruit development process showed a high-low-high-low trend, and the accumulation of the second peaks lag and the accumulation of capsaicin. Further analysis indicated that there was a negative correlation between the accumulation of lignin in the fruit and the synthesis of capsaicin. The level of tannins is fluctuant in the thoughout ftuit development stage, and the trend of the change level of tannins is not related to the change of the accumulate level of capsaicin. The content of flavonoids in the fruit of pepper was a single peak in the development of the fruit, and the accumulation of flavonoids (60d) lagged behind the accumulation of capsaicin (52d). And there was a significant negative correlation with the synthesis of capsaicin. Protein content in pepper fruit development period, with the increase of protein content in fruit, amount of enzyme involved in the synthesis also increased, increase the activity of enzyme, so as to promote pepper flesh and placenta capsaicin synthesis increased, but with capsaicin synthesis correlation was not significant. It is speculated that the biosynthesis of capsaicin and the synthesis of lignin and flavonoids have a competitive relationship, but not the synthesis of tannin and protein.7. Relationship between endogenous hormones and the accumulation of capsaicin in pepper fruit development.In Niujiao peper L-10, IAA content plunged from a higher value, in favor of synthetic ca psaicin and In this case synthetic capsaicin peaked. GA high levels is not conducive to the syn thetic of the capsaicin. The content of ZR in different developmental stages and the accumulation of capsaicin did not show a regular relationship. ETH contents peak have a higher consistency with peak capsaicin content. While the peak of ABA content and the accumulation of capsaicin were the same as 52d in the flower Thus, ABA may play an important role in regulating the synthesis of capsaicin in pepper fruits. Through the analysis of the relationship between the hormone balance and the accumulation of capsaicin, the low IAA/ABA and IAA/ETH in the test were proved to be beneficial to the formation of capsaicin in pepper fruit.8. Relationship between capsaicin accumulation and biosynthesis metabolism in Pepper Fruits.This study is clear, in the process of synthesis of capsaicin in pepper fruit, IAA and GA in early stage were lower in PAL and CS activity, With the fruit development, the content of GA and PAL decreased, the activity of IAA and CS and the expression of gene increased. While the total amount of simple phenolic compounds is higher, the content of capsaicin in this stage is also increased by the simple phenols, It may also be related to the low POD activity, which is accompanied by the synthesis of capsaicin. However, because of the low level of PPO activity, the content of CS increased gradually with the increase of CS content, and the increase of ABA content, and the effect of IAA and GA, which is the key enzyme activity, which makes the accumulation of capsaicin in pepper fruit is the highest. After that, PAL and CS activity and related gene expression decreased, and maintained low level, due to the secondary metabolism into the exuberant period, the contents of the precursors were also decreased, while the increase of POD enzyme activity, accelerated the degradation of capsaicin, and the synthesis of lignin and flavonoids, and the synthesis of capsaicin to compete in the former phenolic substances, so as to decrease the content of capsaicin.9. Exogenous Putrescine on capsaicin biosynthesis regulation mechanism in pepper fruit.This experiment by using polyamine biosynthesis key enzyme ODC inhibitor difluoromethylornithine (DFMO) inhibitor spraying pepper flowers. It is clear that the flowers of 0.1 mmol·L-1 Put can promote the accumulation of capsaicin in pepper fruits. The mechanism of exogenous Put to promote the accumulation of capsaicin was preliminarily considered that exogenous polyamine increase the level of enogenous polyamine and futher promote the growth of pepper fruits and enhance the activity of the pivotal enzymes CS and PAL in the capsaicin synthesis pathway as well as up-regulated the expression of them, and finally promote more phenolic precursor converted into capsaicin. |
PDF Full Text Request