The Development And Application Of High Performance Liquid Chromatographic Detection Of Biogenic Amines In Foods

Biogenic amines(BAs)can hurt body's nervous system and cardiovascular after excessive intake.Thus,it is important to establish a method to detect the content of biogenic amines in food.Two methods of detecting biogenic amines were established in this paper.One method is that simultaneous detection of fifteen biogenic amines in cheese was established by on-line solid phase extraction(SPE)coupled with capillary liquid chromatography.The other is that fifteen biogenic amines in animal derived products simultaneously detected by high performance liquid chromatography with off-line solid phase extraction.The main parameters of on-line SPE were optimized by single factor variable test.A Zorbax SB-C18 column(0.5 x 35 mm,5?m)was chosen as SPE column.5%acetonitrile ?water worked as mobile phase with isocratic elution.The flow rate is 10?L min-1 and injection volume is 1 ?L.The six-way valve was switched at 3 min.The capillary liquid chromatography conditions were also optimized.The Zorbax SB-C18 colummn(0.5 x 150 mm,5 ?m)was chosen as analytical column to separate biogenic amines using ultra-pure water and acetonitrile with gradient elution.The column temperature was 35? and the detection wavelength was 245 nm.Fifteen kinds of biogenic amines have good linearity(R2?0.9981)within 0.25-50.0 mg L-1,and the limits of detection(LOD)were in the range of 0.02-0.10 mg kg-1.At spiked levels of 1?20,40 mg kg-1,recoveries of fifteen biogenic amines on four kinds of cheese ranged from 79.65-118.66%except methylamine,ethylamine,3-methylbutanamine and 5-hydroxytryptamine;with RSDs from 0.33-14.93%except 3-methylbutanamine and 5-hydroxytryptamine.This indicated that the method was suitable for the determination of biogenic amines in cheese samples.The main parameters of off-line SPE were also optimized.Waters Sep-Pak C18 was chosen as SPE column.The pH of sample solution was adjusted to 9 and 10%acetone solution was used as the washing solution.5 mL of ethyl acetate worked as elution solution.HPLC separation conditions were also optimized.Inertsil ODS-3 C18(250 x 4.6 mm,5?m)column was used to separate fifteen biogenic amines using ultra-pure water and acetonitrile with gradient elusion.The incubator temperature was 35? and flow rate is 0.8 mL min-1.The injection volume was 20 ?L.All biogenic amines were able to separate successively and linear relationship of fifteen kinds of biogenic amines was good(R2>0.9990)in 0.02-5 mg L-1.The limits of detection were between 0.001 to 0.02 mg L-1 as well as recoveries of meat products and fish samples between 70.49-121.16%,with RSDs from 0.71-15.99%,indicating that the established method was able to determine the content of biogenic amines in meat products.The contents of biogenic amines in pork and beef meat stored at different temperatures(room temperature,-4?,-20?)and different time(14 h,7 d,30 d)were measured by established method.The results showed that the contents of methylamine,cadaverine,histamine,5-hydroxy-tryptamine and tyramine changed obviously in pork and beef,indicating they were relevant to the level of meat corruption.Therefore,biogenic amines can be used as indicators of meat freshness.And the contents of biogenic amines in meat stored at a low temperature can significantly be reduced,which it is important for people's health.