Quantitative Detection Of Cow Milk In Goat Milk Based On Non-protein Biomarker

The purchase price of goat milk was higher than cow milk.Driven by economic interests,the phenomenon of adding cow milk to goat milk has appeared.Enzyme-linked immunoassay based on the difference in protein molecular structure between goat milk and cow milk is currently the most widely used method to identify the above-mentioned adulteration behavior.However,this technology cannot detect heat-treated cow milk in goat milk,which limits the application and promotion of this method.In order to overcome this limitation,this research developed a method for detecting the above-mentioned adulteration behavior based on the key non-protein biomarkers of cow milk.The main results obtained are as follows:We used cold acetone to precipitate and remove proteins in goat milk and cow milk,and used SYLON HTP reagent for silylation derivatization.Then,a full-scan analysis of the derivatives was performed by gas chromatography-mass spectrometry,and there were two chromatographic peaks in the cow milk derivatives,which did not appear in the goat milk derivatives.After searching through the NIST library and comparing the standard substances,it is confirmed that the two chromatographic peaks were the same substance,which is N-acetylglucosamine.Then,based on this non-protein biomarker,methods based on gas chromatography-mass spectrometry and high performance liquid chromatography were developed to quantitatively detect the cow milk in goat milk.The method based on gas chromatography-mass spectrometry to quantitatively detect the cow milk in goat milk was established.The protein in the sample was removed by precipitation with cold acetone,and the sample was derivatized with the silylating reagent SYLON HTP.The content of N-acetylglucosamine in the sample was analyzed by gas chromatography-mass spectrometry in the SIM mode(the content of the marker in cow milk is 147.8 mg/L),and the amount of milk in the sample was calculated.The detection limit of this method is 1.5% of milk addition,and the quantification limit is 5.0% of milk addition.The linearity is good in the range of quantification limit to 100% milk addition,and the recovery rate is 92.3% ～ 97.0%,the spiked concentrations of 10,50 and 100 mg/L correspond to intraday and intraday precisions(RSD)of 8.7% and 9.5%,7.5% and 9.0%,5.3%and 6.2%,respectively.Due to the slightly lower sensitivity and precision of the above method,the method for quantitative detection of cow milk mixed with goat milk based on high-performance liquid chromatography with higher sensitivity and precision was developed.The protein in the sample was removed by precipitation with cold acetone,and the sample was derivatized with1-phenyl-3-methyl-5-pyrazolone.The content of N-acetylglucosamine in the sample was analyzed by high performance liquid chromatography with a UV detector,and the amount of milk in the sample was calculated.The detection limit of this method is 0.3% of milk addition,and the quantification limit is 1.0% of milk addition.The linearity is good in the range of quantification limit to 100% milk addition,and the recovery rate is 100.4% ～105.1%,the spiked concentrations of 10,50 and 100 mg/L correspond to intraday and intraday precisions(RSD)of 1.8% and 2%,1.7% and 3.7%,2.6% and 2.7%,respectively.