Agar plate method using Lactobacillus plantarum for biotin determination in serum and urine

An improved agar plate method of biotin bioassay using Lactobacillus plantarum ATCC 8014 and bromocresol purple was established to determine biotin levels in human serum and urine. Samples were treated with 4.5 N H2SO4 to liberate free biotin, autoclaved for 1 h and neutralized by 4.5 N NaOH, then 10 microliters was added to wells in each plate. The biotin levels were measured in 190 serum and 59 urine samples, and the means were 2.7 +/- 0.53 ng/ml and 12.4 +/- 5.56 ng/mg of creatinine, respectively. The intra-assay coefficient varience (CV) were 3.2 (n = 20) and 1.3% (n = 23), respectively. The recovery of biotin added (10 ng/ml) to serum was 110.7%, and to urine was 99.6%. These findings suggest that this assay is sufficiently accurate and reproducible for routine use in the clinical laboratory. The excretion of orally administered biotin was also demonstrated by the method.