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Research on the Extraction and Purification of Yak Blood Immunoglobulin G and Its Stability

Author PangChen
Tutor ZhangZhen
School Gansu Agricultural University
Course Nutrition and Food Hygiene
Keywords Yak blood IgG extraction purification Stability
Type Master's thesis
Year 2013
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The yak is one kind the rare and precious species which survives under the low pollutionecological environment, the content of IgG in its blood is3times higher than normal cattle.This paper researched by remove Thrombin after separation by freezing fresh yak as rawmaterials, has studied on FeCl3salting out method and polyphosphates precipitation methodextraction yak blood separately the IgG optimal technological parameter, the purificationcondition of the ion-exchange chromatography, the gel permeation chromatography and SDS-PAGE to IgG, studies under the dissimilar condition the IgG stability, the research results areas follows:(1) The extraction of FeCl3salting out method in yak serum IgG of optimal processparameters: reaction temperature34℃, ferric chloride concentrations for2.7mmol/L, reactiontime of1.8h, pH value of4.4, and under these conditions the extracting IgG8.28mg/mL.(2) The extraction of polyphosphates precipitation method in yak serum IgG of optimalprocess parameters: reaction temperature for32℃, degrees centigrade polyphosphatesconcentration for0.1%, pH value is3.7, the response time is1.7h, yak serum IgG extractionunder the technological conditions for the6.48mg/mL.(3) After the igG purified to obtain a elution peak, located in between the tubes12to27, IgGchange from light yellow to white liquid liquid state other contaminating proteins have beenfiltered, continued elution found other components. Carries on the purity appraisal afterpurification IgG its result to indicate is≥95%IgG standard banding not different also notappears other proteins with the purity.(4) The activeness of IgG will change in the different response environment,when neutralenvironment and over65℃, the IgG activeness loss speed is quick, in75℃heats up1h, IgGcomplete deactivation. When the pH is2.0, the IgG active retention rate is40%, in the pH isunder4.5conditions, after65℃and75℃heats up2h and1h respectively, the IgG completedeactivation, in the pH is under4.0conditions, after65℃and75℃heats up2h and1hrespectively, IgG complete deactivation.(5) Adds the pepsin and trypsin under the temperate reaction condition, the IgG activenesssuffers loss, after the pepsin recruitment is5.0mg/mL responded3h, the IgG active retentionrate is85.21%, under the temperate reaction condition, the denaturation of IgG was causes bythe pepsin; After the trypsin recruitment is5.0mg/mL responded3h, the IgG live retention rateis82.54%, under the mild condition, the denaturation of IgG was causes by the trypsin.(6) In the simulation artificial digesting fluid, the IgG activeness changes, IgG in the simulation gastric juice digests6h, the survival rate is1.51%, IgG is digested. IgG insimulation intestinal juices digests6h, the survival rate reaches50.23%still to have majorityof IgG maintains the activeness, the effect of intestinal tract environment on IgG is quiterelatively slow.In summary, FeCl3salting out method and polyphosphates precipitation method can be agood extraction of IgG in Yak blood. Use of ion exchange chromatography and gel filtrationchromatography of the crude extract high purity of IgG purified. Yak blood IgG in acidic andhigh temperatures easily inactivated, enzyme solution and artificial digestion can affect thestability of IgG. Yak blood has the potential to be used as an important source of IgGextracted.

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