Efficient extraction method suitable for measurement of endogenous hormone of musa and heliconia plants

Abstract

The invention belongs to the technical field of plant science and discloses an efficient extraction method suitable for measurement of endogenous hormone of musa and heliconia plants. The method comprises the following steps: weighing 0.2g of banana leaves, quickly freezing liquid nitrogen, grinding the quickly frozen liquid nitrogen into powder, adding 6mL of pre-cooled 80% methanol solution, andperforming extraction over night in a refrigerator with the temperature of 4 DEG C; adding 2mL of 80% methanol for washing residues, repeating the operation twice, and mixing supernatant liquid; enabling the supernatant liquid to pass through a 0.22-micron micro-porous filtering film, adding 2mL of 80% methanol for washing residues of the micro-porous filtering film, transferring the filtered supernatant liquid into a rotary evaporation bottle, performing evaporation under reduced pressure at 36 DEG C by dim light until the water phase is about 2mL, taking the water phase, and washing the rotary evaporation bottle twice with 2mL of pure water, totally about 6mL; performing refrigeration for 10min at -80 DEG C, and then, performing freeze-drying to obtain dry powder; performing dissolutionwith 4mL of a phosphate buffer of which the pH is 3.0, performing extraction thrice with an equal volume of ethyl acetate to leave an ester phase, performing evaporation to dryness under reduced pressure at 40 DEG C by dim light, performing dissolution with a mobile phase of a pump A, and performing treatment with a Pak-ODS-C18 small column; and before HPLC analysis is performed, enabling the sample to pass through the 0.22-micron micro-porous filtering film to serve as a test solution. The method disclosed by the invention has the advantages that the existing extraction process is adopted, the operation is simple, the consumed time is short, the time and labor are saved, and the recovery rate is increased.

Description

technical field [0001] The invention belongs to the technical field of plants of the genus Musa and Helicana, and in particular relates to a high-efficiency extraction method suitable for measuring endogenous hormones of plants of the genus Musca and Helicana. Background technique [0002] At present, the existing technologies commonly used in the industry are as follows: High performance liquid chromatography (HPLC) has the characteristics of high efficiency and high sensitivity, and has become an ideal analysis method for plant endogenous hormones. The most commonly used extraction method at present is to extract with 80% icy methanol, extract overnight, collect the filtrate, spin evaporate to dryness, dissolve the raffinate with a small amount of water, freeze-thaw overnight, after melting, centrifuge to get the supernatant, adjust the pH, Add ethyl acetate for extraction, take the organic phase, and then rotate it to evaporate, and finally dissolve it with the mobile ph...

AI Technical Summary

Problems solved by technology

Musa and Helicana plants have the characteristics of relatively few inclusions and interfering substances. The existing extraction process is not only complicated and time-consuming, but also only one target can be detected at a time, with overlapping peaks and excessive crowding. , bimodal, tailing and other serious problems, but also time-consuming and labor-intensive, and the recovery rate is generally lower than 80%

[0003] In summary, the problems in the prior art are: The technological process of this method is backward in terms of the current improved pretreatment method and the progress of analytical purification instruments

It is mainly manifested in the following defects in the process of collecting the supernatant after simple centrifugation of the extract: 1. Suspended liquid generally exists at the junction of the supernatant and the sediment. If the suspension is collected and incorporated into the supernatant, it may Improve the sample recovery rate, but precipitation interference will occur in subsequent operations; 2. If the suspension liquid at the junction of the supernatant and the precipitation is discarded, the sample recovery rate will be reduced

The prior art uses buffer solution to adjust pH before extraction, which has the following defects: 1. Some samples need to add a large amount of buffer solution, frequent operations are prone to pollution, and the sample volume before extraction is greatly increased; 2. Some samples need to add a large amount of phosphoric acid during the pH adjustment process 3. The sample volume is large, and a large amount of extraction reagent needs to be added in the extraction process. Repeated extraction and long-term high-temperature rotary evaporation will obviously destroy the hormone structure (IAA is the most obvious)

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