| Detection of immunoglobulin IgG content by high performance liquid chromatography, the material to be examined for the pretreatment of sample optimization of experimental conditions and optimization of chromatographic conditions, can improve the detection efficiency and the actual effect. A method for determining the IgG content of immune globulin in the infant dairy products accurately has been established in This topic. The research aims at the pre-treatment methods and high performance liquid chromatography separation detection conditions. The extraction solvent using in the sample pre-treatment、 centrifugation rate、ultrasonic time、sample dilution factor、the acid adjusting the pH and the detection wavelength、column、flow rate、column temperature and the pH of the mobile phase are discussed respectively, and adopting the retention time to determine the nature,using the peak area external standard method to quantify. The results confirm the optimal pre-treatment conditions:sample is dissolved with solution A that eight times than the sample, centrifuge under4℃with8000r/min, adjust the pH with10%phosphoric acid, ultrasonic20-30min; The best chromatography conditions:choose the Hi-trap protein G the HP Chromatographic column,0.05mol/L pH6.7phosphate buffer solution is used for the combine liquid,0.05mol/mL pH2.7glycine salt buffer is used as the eluent, the detection wavelength is280nm, flow rate is0.4mL/min, column temperature is25℃. The detection limit of the method is0.7μg/ml, the average recovery is92.16%, the standard deviation is3.34%, IgG content shows a good linear relationship with the target peak area in the0.1-1.0mg/mL: y=2519x-24.71(R2=0.9992). And with this method for analyzing the content of IgG in the actual samples, Results for:211.4、176.2、137.8、331.1mg/kg. Also proved that the method is better than the chromatographic peak of national standard. |
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