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Measurement of milk beta-hydroxybutyrate with a simple UV spectrophotometer method:an alternative assay method

Measurement of milk beta-hydroxybutyrate with a simple UV sp...

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中国畜牧兽医学会动物毒物学分会2009年学术研讨会

作者： ZhANG Z G~(1,2),LIU G W~1,LI X B~1,WANG Z~(1*),LI Y F~2,XU S W~2 (1.College of Animal Science and Veterinary Medicine,Jilin University, Changchun 130062 2.College of Veterinary Medicine,Northeast Agriculture University,harbin,150030)

Ketosis is a common metabolic disorder frequently observed in dairy cows during the early lactation *** is a metabolic condition characterized by increased levels of ketone bodies in blood,urine and *** major ketone body is beta-hydroxybutyrate (BhBA).Determination of BhBA in milk samples is an important tool in the diagnosis of subclinical/clinical ketosis in dairy *** this report, we describe a novel method based on the traditional spectrophotometric approach for measuring BhBA levels in milk of dairy *** two commercial dairy farms,milk samples were taken from dairy cows within 2 months after *** spectrophotometric approach involve d two ***,in the prese nce of NAD,was oxidized by BhBA dehydrogenase to AcAc and NADh. Then NADh and nitroblue tetrazoli um were reduced by diaphorase to fo rmazan and NAD,*** amount of formazan was measured *** were prepa red as follows:milk was deproteiniz ed with perchloric acid,centrifuged, the supernatant was neutralized with KOh *** standard curve was made by a series of dilutions the BhBA standard solution.A reaction buffer was prepared,and comprised Tris buffer,Triton x-100,NBT,BhB A dehydrogenase,diaphorase and oxamic ***-buffer(ph 8.5), the reaction solution and prepared milk sample were added to centrifug e tubes,in order,and shaken vigorou *** resulting mixture was kept at 30 min at room temperature in the dark and assayed using a UV spectro *** following paramete rs were established:linearity,recove ry,repeatability,decision limit and *** fluorometric detectio n procedure as *** sample buffer comprised Tris-buffer ph 8.5, oxamic acid,NAD,rezasurin,Triton X-100,BhBA dehydrogenase and *** end-point process was read after 20 *** results showed that the standard-curve equa tion of the UV spectrophotometer method was y=0.2582x+0.0269 (R=0.9967).The reaction buffer did not react with AcAc,L-s

关键词： dairy cows beta-h ydroxybutyrate assay milk

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