Determination Of Folic Acid In Milk Powder By SPE-RP-HPLC

Ultrasonic extraction,high chlorine acid solution to remove protein,solid phase extraction technology purification and HPLC methods were applied for determination of folic acid(FA) content in milk powder.Aimed at establishing a rapid and accurate determination of folic acid content in milk powder.Through the research on experimental conditions,establishing the method that detect the content of folic acid in milk powder,specific operating procedure is as follows:(1) Ssample pretreatment:5g milk samples were taken exactly(accurate to 0.0001 g) in 50mL conical beaker;add 30mL deionized water(40～50℃) dissolve;ultrasonic oscillator extraction for 15min in 50℃condition;add 60%of high chlorine acid 1.5 mL for deposition protein;after deposition completely,the whole solution were metastasize to 50mL volumetric flask;add deionized water for constant volume to scale,shake equably;centrifugal 15min in 4000r/min condition;after filter,reserving in 0～4℃refrigerator.(2) Solid-phase extraction:①Uuse 5mL 100%methanol to activate extraction column; add 5mL water washing methanol for balance column;②Aadd 5mL sample solution to the column;use solid-phase extraction apparatus to control flow velocity in the 40d/min for making sample through column;③After all samples through solid-phase extraction column, washing out interferent with 6mL 1%deionized wash;④Ccollect the eluent with 5mL 100% methanol and dry with nitrogen;save backup.(3) Dderivatives methods:dissolve sample with 1mL water after dry nitrogen and scroll oscillation;add 1 mL 0.2g/100mL potassium permanganate derivatives,1mL 0.1mol/L sodium hydroxide and 2mL deionized water and scroll oscillation.Then add water heating 40min at 60℃.Rinse with water,scroll oscillation and use organic film filtration into the autofilling bottle,on standby.(4) Cchromatographic detection conditions:BDS:HYPERSIL C18 column(250×4.6 mm i.d.5μm);Column temperature 30℃;detector is detected wavelength,fluorescence detector:Em 285nm for Ex:for 437nm;Mobile phase for 0.1 mol/L sodium dihydrogen phosphate solution,Velocity 1mL/min,Enter the sample weight 20μL.Experimental results show that the method has following advantages:simple,precise, accurate and easy to extend.Methods can meet dairy enterprises detection folic acid content in milk powder.