阅读说明：本技术 一种橄榄叶中苦苷和羟基酪醇含量的快速检测方法 (Method for rapidly detecting content of picroside and hydroxytyrosol in olive leaves ) 是由 冯士令 丁春邦 陈涛 周莉君 张春艳 李天� 袁明 于 2020-06-02 设计创作，主要内容包括：本发明公开了一种橄榄叶中苦苷和羟基酪醇含量的快速检测方法,通过将橄榄叶粉末与甲醇的水溶液混合提取,离心取上清液过滤备用,确定流动相和梯度洗脱程序,将待测样品液体进样,设定检测条件,与标准品进行比较而获得的提取物中羟基酪醇和苦苷的含量。本发明方法检测不同储存和干燥条件下橄榄叶中苦苷和羟基酪醇重复性好,稳定性强,且ZORBAX SB-C18反相色谱柱低成本具有实验室通用性,可以广泛推广使用。(The invention discloses a method for rapidly detecting the contents of picroside and hydroxytyrosol in olive leaves. The method disclosed by the invention has the advantages that the repeatability and the stability of detecting the picroside and the hydroxytyrosol in the olive leaves under different storage and drying conditions are good, the ZORBAX SB-C18 reversed phase chromatographic column is low in cost, has the laboratory universality, and can be widely popularized and used.)

1. A method for rapidly detecting the contents of picroside and hydroxytyrosol in olive leaves is characterized in that the contents of the picroside and the picroside in an extract are obtained by mixing and extracting olive leaf powder and an aqueous solution of methanol, centrifuging, taking supernate, filtering for later use, determining a mobile phase and a gradient elution program, injecting sample liquid of a sample to be detected, setting detection conditions and comparing the sample liquid with a standard substance.

2. The method as claimed in claim 1, wherein the concentration of methanol in the aqueous solution of methanol is 78% by volume.

3. The method as claimed in claim 1, wherein the mass-to-volume ratio of the olive leaf powder to the methanol aqueous solution is 1:50g/m L.

4. The method as claimed in claim 1, wherein the mobile phase is water (A) -acetonitrile (B), and the gradient elution procedure comprises: 0-10min, 90% A-10% B; 14-15min, 80% A-20% B; 30min, 90% A-10% B.

5. The method of claim 1, wherein the conditions of rapid detection of picroside and hydroxytyrosol content in olive leaf are that the flow rate is 0.8m L/min, the chromatographic column is ZORBAX SB-C18, the column temperature is 30 ℃, the sample amount is 10 μ L, and the detection wavelength is 280 nm.

Technical Field

The invention belongs to the technical field of natural product detection, and particularly relates to a method for rapidly detecting the content of picroside and hydroxytyrosol in olive leaves.

Background

Olive (Olea europaea L.) as an economic crop is mainly planted in Greece, Italy and other countries in the mediterranean coast at first and then introduced into China, and is mainly distributed in Yunnan, Sichuan, Gansu, Guizhou and other places.

The olive leaves mainly contain polyphenols, flavonoids, secoiridoids, biflavones and the like as active substances, wherein Oleuropein (O L E) and Hydroxytyrosol (HT) are two main polyphenol compounds.

Oleuropein and hydroxytyrosol have wide pharmacological activity and are phenolic compounds in fresh olive leaves, but the oleuropein and the hydroxytyrosol are low in content and are difficult to be efficiently detected by an HP L C method.

Disclosure of Invention

The invention aims to provide a method for rapidly detecting the contents of oleuropein and hydroxytyrosol in olive leaves, which is optimized on the basis of an HP L C detection technology, changes the mobile phase components (water and acetonitrile) of a new optimized HP L C detection method into (water and acetonitrile), shortens the total detection time to only 35min, and has no foreign peaks around the hydroxytyrosol.

The technical purpose of the invention is realized by the following technical scheme:

a method for rapidly detecting the contents of picroside and hydroxytyrosol in olive leaves comprises the steps of mixing and extracting olive leaf powder and an aqueous solution of methanol, centrifuging, taking supernatant, filtering for later use, determining a mobile phase and a gradient elution program, injecting sample liquid to be detected, setting detection conditions, and comparing with a standard substance to obtain the contents of the hydroxytyrosol and the picroside in an extract.

The volume concentration of the methanol in the methanol water solution is 78%.

The mass volume ratio of the olive leaf powder to the methanol water solution is 1:50g/m L.

The mobile phase is water (A) -acetonitrile (B), and the gradient elution procedure is as follows: 0-10min, 90% A-10% B; 14-15min, 80% A-20% B; 30min, 90% A-10% B.

The detection conditions comprise a flow rate of 0.8m L/min, a chromatographic column of ZORBAXSB-C18, a column temperature of 30 ℃, a sample injection amount of 10 mu L and a detection wavelength of 280 nm.

The invention has the beneficial effects that:

1) the peak time of hydroxytyrosol is 29.3min, which is at least 10min shorter than other methods.

2) The method has the advantages that the content of the hydroxytyrosol in the olive leaves is low, the hydroxytyrosol is easy to degrade under the influence of storage and drying conditions, and the hydroxytyrosol and oleuropein signals under different storage conditions (20 ℃, 4 ℃ and room temperature) and the oleuropein signals under different drying conditions (freeze-drying, normal-temperature air drying and 70 ℃) can be detected at the same time, so that the method is high in sensitivity and strong in stability.

3) The ZORBAXSB-C18 reversed phase chromatographic column used by the method has low cost and wide application.

Drawings

FIG. 1 is an HP L C chromatogram of Hydroxytyrosol (HT) and oleuropein (O L E) standards at 280 nm;

FIG. 2 is an HP L C chromatogram of olive leaf samples Hydroxytyrosol (HT) and oleuropein (O L E) at 254, 260 and 280 nm;

FIG. 3 is a chromatogram of Hydroxytyrosol (HT) and oleuropein (O L E) at-20 ℃;

FIG. 4 is a chromatogram of Hydroxytyrosol (HT) and oleuropein (O L E) at 4 ℃;

FIG. 5 is a chromatogram of Hydroxytyrosol (HT) and oleuropein (O L E) at room temperature;

FIG. 6 is a chromatogram of Hydroxytyrosol (HT) and oleuropein (O L E) under lyophilization conditions;

FIG. 7 is a chromatogram of Hydroxytyrosol (HT) and oleuropein (O L E) under normal temperature drying conditions;

FIG. 8 is a chromatogram of Hydroxytyrosol (HT) and oleuropein (O L E) under dry conditions at 70 ℃.

Detailed Description

The technical solutions of the present invention will be described clearly and completely with reference to specific embodiments of the present invention, and it should be understood that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.