阅读说明：本技术 一种测定饲料中β-阿朴-8＇-胡萝卜素酸乙酯着色剂含量的方法 (Method for measuring β -apo-8' -ethyl carotenoate colorant content in feed ) 是由 章厉劼 王庆红 孙真峥 赵贵 栾庆祥 钱莘莘 杨强 孙播东 罗永成 焦仁刚 金晓 于 2019-11-07 设计创作，主要内容包括：本发明公开了一种测定饲料中β-阿朴-8＇-胡萝卜素酸乙酯着色剂含量的方法,包括：(1)预处理,称取待测饲料样品,加入水、饱和氯化钠、0.05％乙酸溶液、混合提取液涡旋混合后离心,吸取上层萃取液；用甲基叔丁基醚+乙腈溶液溶解残渣,过滤得供试样品溶液；(2)液相色谱仪分析,测定β-阿朴-8＇-胡萝卜素酸乙酯着色剂的峰面积；(3)利用β-阿朴-8＇-胡萝卜素酸乙酯线性回归方程计算待测饲料中β-阿朴-8＇-胡萝卜素酸乙酯着色剂的质量浓度,再计算待测饲料中β-阿朴-8＇-胡萝卜素酸乙酯着色剂的含量。本发明方法具有简单、快速、准确、灵敏的特点,适用于饲料中β-阿朴-8＇-胡萝卜素酸乙酯着色剂含量的检测分析。(The invention discloses a method for determining the content of β -apo-8 ' -ethyl caronate colorant in feed, which comprises the steps of (1) preprocessing, weighing a feed sample to be determined, adding water, saturated sodium chloride, 0.05% acetic acid solution and mixed extracting solution, performing vortex mixing, centrifuging, absorbing upper extraction liquid, dissolving residues by using methyl tert-butyl ether and acetonitrile solution, filtering to obtain a sample solution, (2) analyzing by using a liquid chromatograph, determining the peak area of β -apo-8 ' -ethyl caronate colorant, and (3) calculating the mass concentration of β -apo-8 ' -ethyl caronate colorant in the feed to be determined by using a β -apo-8 ' -ethyl caronate linear regression equation, and then calculating the content of β -apo-8 ' -ethyl caronate colorant in the feed to be determined.)

1. A method for determining the content of β -apo-8' -ethyl carotenoate colorant in a feed, comprising:

(1) pretreatment of

Weighing a feed sample to be detected, adding water, saturated sodium chloride, 0.05% acetic acid solution and mixed extracting solution, mixing in a vortex mode, centrifuging, and sucking upper-layer extracting solution;

repeatedly extracting with a mixed extracting solution, wherein the mixed extracting solution is a mixed solution of methanol, ethyl acetate and n-hexane;

mixing the extractive solutions, adding anhydrous ethanol, and mixing; drying under nitrogen;

with methyl tert-butyl ether: dissolving residues in an acetonitrile solution, and filtering to obtain a sample solution;

(2) liquid chromatograph analysis

Analyzing the test sample solution on a liquid chromatograph, and determining the peak area of β -apo-8' -ethyl carotenoate coloring agent;

(3) calculating the content of β -apo-8' -ethyl carotenoate colorant in the feed

Determining the peak area of β -apo-8 '-ethyl caronate by utilizing a plurality of groups of β -apo-8' -ethyl caronate solutions with known mass concentrations, performing linear regression by taking the mass concentration of β -apo-8 '-ethyl caronate as a horizontal coordinate and the corresponding peak area as a vertical coordinate, and establishing a β -apo-8' -ethyl caronate linear regression equation;

according to the peak area of β -apo-8 '-ethyl carotenoate of the feed sample to be detected, calculating the mass concentration of the β -apo-8' -ethyl carotenoate coloring agent in the feed to be detected by using a β -apo-8 '-ethyl carotenoate linear regression equation, and then calculating the content of the β -apo-8' -ethyl carotenoate coloring agent in the feed to be detected.

2. The method for determining the content of β -apo-8 '-carotenoic acid ethyl ester colorant in a feed according to claim 1, wherein the β -apo-8' -carotenoic acid ethyl ester linear regression equation is:

y＝37985x-8487.3

wherein y is the peak area of β -apo-8 '-ethyl carotenoate, and x is the mass concentration of β -apo-8' -ethyl carotenoate colorant in the feed to be detected.

3. The method for determining the content of β -apo-8 '-carotenoic acid ethyl ester colorant in feed according to claim 1, wherein the formula for calculating the content of β -apo-8' -carotenoic acid ethyl ester colorant in feed is:

in the formula, W is the content of β -apo-8 '-ethyl carotenoate colorant in the feed to be detected, x is the mass concentration of β -apo-8' -ethyl carotenoate colorant in a sample to be detected, V is the volume of a methyl tert-butyl ether and acetonitrile solution of a dissolution residue, and m is the mass of the feed sample.

4. The method for determining the content of β -apo-8' -ethyl carotenoate colorant in a feed according to claim 1, wherein the volume ratio of methyl tert-butyl ether to acetonitrile in the methyl tert-butyl ether + acetonitrile solution is 1: 9.

5. The method for determining the content of β -apo-8' -ethyl carotenoate colorant in a feed according to claim 1, wherein the volume ratio of methanol to ethyl acetate to n-hexane in the mixed extracting solution is 1: 1: 1.

6. The method for determining the content of β -apo-8 '-carotenoic acid ethyl ester colorant in feed according to claim 1, wherein the detection range of the content of β -apo-8' -carotenoic acid ethyl ester colorant in the feed is 0.2-50 mg/kg.

Technical Field

The invention relates to a method for measuring the content of β -apo-8 '-ethyl carotenoate colorant in feed, belonging to the technical field of β -apo-8' -ethyl carotenoate colorant content detection.

Background

According to specific stipulations in No. 2625 of Ministry of agriculture of the people's republic of China, the highest limit of β -apo-8 ' - β -carotene-4, 4-dione (canthaxanthin) in the compound feed is 80 mg/kg. at present, no national standard and industrial standard method exists for detecting β -apo-8 ' -ethyl caronate colorant in the feed in China, so that the detection of the colorant in the feed is blank, and the disadvantages of feed product quality safety risk and supervision are increased.

β -apo-8 '-ethyl caronate is also called apocaronate or apoester, which is an oxygen-containing apocaronate compound in carotenoid, the product is mainly applied to coloring edible oil, margarine, jam, jelly and beverage products in the food industry, at present, in the detection of β -apo-8' -ethyl caronate colorant in feed, due to the interference of lipid and protein in a sample on a result and the unstable factors of the β -apo-8 '-ethyl caronate colorant, the problems of poor accuracy, low sensitivity, slow analysis speed and the like exist in the detection of the β -apo-8' -ethyl caronate colorant in feed.

Disclosure of Invention

The invention aims to solve the technical problems that the method for determining the content of β -apo-8 '-ethyl carotenoate colorant in feed has high sensitivity, good accuracy and high analysis speed, and the interference of lipid and protein in a sample on a detection result and the instability of the β -apo-8' -ethyl carotenoate colorant in an object to be detected in the prior art are solved through reasonable sample extraction and purification conditions.

The technical scheme of the invention is that the method for measuring the content of β -apo-8' -ethyl carotenoate colorant in the feed comprises the following steps:

(1) pretreatment of

Weighing a feed sample to be detected, adding water, saturated sodium chloride, 0.05% acetic acid solution and mixed extracting solution, mixing in a vortex mode, centrifuging, and sucking upper-layer extracting solution;

repeatedly extracting with mixed extracting solution, wherein the mixed extracting solution is methanol: ethyl acetate: a mixed solution of n-hexane;

mixing the extractive solutions, adding anhydrous ethanol, and mixing; drying under nitrogen;

dissolving the residue with a methyl tert-butyl ether and acetonitrile solution, and filtering to obtain a sample solution;

(2) liquid chromatograph analysis

Analyzing the test sample solution on a liquid chromatograph, and determining the peak area of β -apo-8' -ethyl carotenoate coloring agent;

(3) calculating the content of β -apo-8' -ethyl carotenoate colorant in the feed

Determining the peak area of β -apo-8 '-ethyl caronate by utilizing a plurality of groups of β -apo-8' -ethyl caronate solutions with known mass concentrations, performing linear regression by taking the mass concentration of β -apo-8 '-ethyl caronate as a horizontal coordinate and the corresponding peak area as a vertical coordinate, and establishing a β -apo-8' -ethyl caronate linear regression equation;

according to the peak area of β -apo-8 '-ethyl carotenoate of the feed sample to be detected, calculating the mass concentration of the β -apo-8' -ethyl carotenoate coloring agent in the feed to be detected by using a β -apo-8 '-ethyl carotenoate linear regression equation, and then calculating the content of the β -apo-8' -ethyl carotenoate coloring agent in the feed to be detected.

Preferably, the β -apo-8' -ethyl carotenoate linear regression equation is:

y＝37985x-8487.3

wherein y is the peak area of β -apo-8 '-ethyl carotenoate, and x is the mass concentration of β -apo-8' -ethyl carotenoate colorant in the feed to be detected.

Preferably, the content of β -apo-8' -ethyl carotenoate colorant in the feed is calculated by the formula:

in the formula, W is the content of β -apo-8 '-ethyl carotenoate colorant in the feed to be detected, x is the mass concentration of β -apo-8' -ethyl carotenoate colorant in a sample to be detected, V is the volume of a methyl tert-butyl ether and acetonitrile solution of a dissolution residue, and m is the mass of the feed sample.

Preferably, the methyl tert-butyl ether: methyl tert-butyl ether in acetonitrile solution: the volume ratio of acetonitrile is 1: 9.

preferably, the ratio of methanol: ethyl acetate: the volume ratio of n-hexane is 1: 1: 1.

preferably, the detection range of the content of the β -apo-8' -ethyl carotenoate colorant in the feed is 0.2-50 mg/kg.

The method has the beneficial effects that β -apo-8 ' -ethyl carotenoate as a colorant under the colorant item in feed additive safety use specifications is taken as a research object, a feed sample to be detected is mixed with water, saturated sodium chloride and 0.05% acetic acid solution to dissolve starch coated on the surface of β -apo-8 ' -ethyl carotenoate, mixed extract liquid of methanol, ethyl acetate and n-hexane is used for liquid-liquid extraction, and after the upper layer solution is concentrated and redissolved, the ultra-high performance liquid chromatography is used for determination, and the result shows that the method has the characteristics of simplicity, rapidness, accuracy and sensitivity, and is suitable for the content detection and analysis of the β -apo-8 ' -ethyl carotenoate colorant in the feed.

The method is beneficial to establishing β -apo-8 ' -ethyl carotenoate colorant content detection standards in the feed to provide a large amount of basic data, gradually perfects the basis for monitoring the feed quality safety risk, evaluates the quality safety of β -apo-8 ' -ethyl carotenoate, masters the loss of β -apo-8 ' -ethyl carotenoate in the feed production, guides feed enterprises and farms to scientifically and reasonably use the colorant, ensures that no major poultry and livestock product quality safety events occur, and reduces the production cost under the condition of meeting the color requirements of consumers.

Drawings

FIG. 1 shows a liquid chromatogram of β -apo-8' -ethyl carotenoate standard sample.

Detailed Description

The present invention will be described in detail with reference to the following embodiments in order to make the aforementioned objects, features and advantages of the invention more comprehensible. In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein, but rather should be construed as broadly as the present invention is capable of modification in various respects, all without departing from the spirit and scope of the present invention.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.