Spermidine synthetase activity determination reagent kit and method

Abstract

The invention discloses a spermidine synthetase activity determination reagent kit and method. The reagent kit comprises a reagent 1 (sodium hydrogen phosphate heptahydrate, sodium dihydrogen phosphate dehydrate, water), a reagent 2 (sodium hydrogen phosphate heptahydrate, sodium dihydrogen phosphate dihydrate, beta-mercaptoethanol, EDTA, phosphopyridoxal, water), a reagent 3 (a reagent 2, putrescine, decarbocylated S-ademetionine), a reagent 4 (perchloric acid, water), a reagent 5 (sodium hydroxide, water), a reagent 6 (saturated sodium bicarbonate), a reagent 7 (dansyl chloride, acetone), areagent 8 (ammonia), a reagent 9 (perchloric acid, water) and a reagent 10 (spermidine standard). The principle of the method is to catalyze putrescine to produce spermide through spermidine synthetase, and then to make spermidine and dansyl chloride react under an alkaline condition to produce a derivative with fluorescence, through separation by a high-efficiency liquid chromatography, a fluorescence detector is used to determine the spermidine production amount, and thus, the activity of the spermidine synthetase is further measured.

Description

technical field [0001] The invention belongs to the field of life sciences, and in particular relates to a high-performance liquid chromatography-based spermidine synthase (SPDS) activity assay kit and a method thereof. Background technique [0002] Polyamines are a class of small molecule biologically active substances that widely exist in organisms and are closely related to plant growth and development, aging and stress resistance. At present, the polyamine synthesis pathway in plants has been basically revealed, and its physiological role has been gradually elucidated at the molecular level. Spermidine synthase (SPDS) is an intermediate enzyme in the synthesis and metabolism of polyamines. Putrescine can generate spermidine under the catalysis of spermidine synthase, and the amount of spermidine generated per unit time is positively correlated with the activity of spermidine synthase, so the activity of spermidine synthase can be expressed as spermidine The amount of p...

AI Technical Summary

Problems solved by technology

The isotope labeling method, firstly, the label is expensive and difficult to prepare, and secondly, the operation process is complicated; the homogeneous time-resolved fluorescence method, firstly, due to the lack of universality of the antibody and the long preparation time, the entire time period is very long and expensive, and secondly are susceptible to cross-reaction interference; at the same time, these two methods have high requirements for equipment

In recent years, with the rapid development of chromatographic technology, the application of detection of small molecules in organisms has become more and more extensive, but there are few reports on the detection technology based on high performance liquid chromatography for the determination of spermidine synthase content in the market

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