The measurement of D-3-hydroxybutyrate (D-BHBA) in milk samples is an important tool for diagnosis of subclinical/clinical ketosis in dairy cows. We describe a simple UV spectrophotometric method for measuring the concentration of D-BHBA in milk of dairy cows. From two herds, 119 milk samples were taken from dairy cows. The standard-curve equation was y = 0.2582x + 0.0269 (R² = 0.9967). The assay was highly specific with a minimum detection limit of 0.01 mmol/L and measuring range of up to 5 mmol/L. The recovery was between 99.35% and 100.22% and repeatability was 99.8%. The comparison between the spectrophotometric method and the fluorometric method revealed a close correlation (r = 0.9939). These results show that the spectrophotometric method can be successfully used as an alternative method to measure D-BHBA content in milk.
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