Polyamines are a group of small organic molecules with physiological activity in plants, mainly putrescine (putrecine, put), cadcine (cadecine, cad), spermidine (spermidine, spd) and spermine (spermine, spm) ). They are closely related to plant growth and development, and participate in plant cell division, DNA aggregation and hormone regulation [1]. Under environmental stress, large amounts of polyamines accumulate in plants, which plays an important role in stabilizing the conformation of macromolecular substances such as cell membranes, nucleic acids, and proteins. Polyamines are also abundant in the human body and are closely related to human health [2]. Diet is one of the important sources of polyamines in the human body [3]. Accurate and rapid detection of polyamines in plant tissues is gaining more and more attention. Because polyamines are aliphatic straight-chain amines, they were mainly measured by amino acid analyzers and paper electrophoresis [4], which not only has low sample recovery rate but also low detection sensitivity. At present, the derivatization method is mostly used. The primary and secondary amino groups of the polyamine are connected to fluorescent or ultraviolet absorbing groups and separated and detected by thin layer or liquid chromatography. Benzenesulfonyl chloride and fluorescein [4 ~ 8]. Since Redmond and Tseng [5] adopted the benzoylation method in 1972, chromatographers have continuously improved this method and have become a more mature method for detecting polyamines in plants. However, in many details, such as the reaction time, the interference of side reactions, sample storage and further optimization of separation conditions, there is still a lot of work to be done. Experimental part 2. 1 instruments and reagents STI5000 liquid chromatograph. Putrescine and spermidine are Fluka products, spermine is Sigma products, methanol is chromatographically pure, water is ultra-pure water, and other reagents are domestically produced analytically pure. 2. 2 Chromatographic conditions Chromatographic column: NovapakC18 column (Waters, 150 × 3.9mm, 4um). Mobile phase: methanol: water (60:40, v / v), detection wavelength 230nm, flow rate 0.7ml / min, column temperature 30 ℃, Breeze chromatography software. 2.3 Material treatment 1.0 ~ 2.0g, add 4ml of pre-cooled 5% perchloric acid (v / v) ice bath to grind, centrifuge (15,000g, 30 minutes, 4 ℃) after immersion in ice bath for 1 hour. Take 500ul of supernatant, add 10ml plastic centrifuge tube with lid, add 7ul benzoyl chloride, then add 1ml 2M NaOH, vortex for 20s, then react in a 37 ° C water bath for 20min. Add 2ml of saturated NaCl solution, mix and extract with 2ml of ether. Take 1ml of ether phase and dry in vacuo. After vortexing with 100ul of methanol, pass 0.45um filter membrane and take 10ul for injection. 2. 4 Standard curve Make the standard product into 1mmol / L concentration stock solution, take 40ul each and carry out benzoylation according to the method of 2.3. The benzoylated polyamine standard solutions 0.03, 0.06, 0.12, 0.25, 0.50, 0.75, 1.0nm / 10ul were injected respectively, and the area was plotted against the injection volume. references 1 Zhao Fugeng, Liu Youliang. Metabolism and regulation of special morphological polyamines in higher plants. Plant Physiology Communications, 2000, 36 (1): 1 ~ 5 This article is derived from the analysis of polyamine content in plant tissues by high-performance liquid chromatography Wpc Fences,Wpc Picket Fence,Wpc Fence Panel,Composite Fencing Panels ZHEJIANG DEQING GONAT FENCE CO.,LTD , https://www.gonatfence.com
2 Tiburcio AF, Altabella T, Flores D et al. Manipulation of polyamines in food plants: chemical
and transgenic approaches, In: Bardóce S, White A (eds). Polyamines in Health and Nutrition. Klvwer Academic Publishers. 1999, 161 ~ 176
3 Ji Xiaojia, Liu Youliang. The role and regulation of plant-derived food polyamines. Botany Bulletin (in press)
4 Nikolaus S. Thin-layer chromatography and thin-layer electrophoresis of polyamines and their
derivatives, In: Tabor H, Tabor CW (eds). Polyamines. New York: Academic Press, 1983, 3 ~ 9
5 Redmond JW and Tseng A. High-press liquid chromatographic determination of putrescine, cadaverine, spermidine and spermine. J of chromatography. 1979, 170, 479- 481.
6 Flores HE and Galston AW. Analysis of polyamines in higher plants by high performance liquid chromatography. Plant physiol. 1982, 69 :, 701-706
7 Wu Shaobo, Guo Feng. High performance liquid chromatography separation of polyamines in rice embryo and endosperm. Chromatography, 1989, 7 (6): 389 ~ 391
8 Du Junying, Sun Bingyong, Chen Zhaozhen, etc .. OPA-mercaptoethanol pre-column derivatization reversed-phase high-performance liquid chromatography for the determination of polyamines in biological tissues. Journal of the PLA Technical College, 1997, (1): 53