Ascorbate Oxidase Minimizes Interference by High-Concentration Ascorbic Acid in Total Cholesterol Assays

Dear Editor, Ascorbic acid interferes with certain clinical chemistry assays based on peroxidase and redox indicators [1]. Ascorbic acid is reported to interfere with the measurement of glucose, total cholesterol, triglycerides, and uric acid [2-5]. We recently encountered several cases of interference from ascorbic acid in serum total cholesterol assays based on colorimetric enzymatic reactions. A 48-yr-old-female with recurrent ovarian cancer underwent palliative segmental resection surgery of the small intestine. The patient showed progressive renal impairment following surgery, and total cholesterol level was <3 mg/dL, as measured using the OSR6516 total cholesterol assay (Beckman Coulter, Brea, CA, USA) on an AU5800 analyzer (Beckman Coulter). We repeated the test with the same sample, using the Pureauto S CHO-N total cholesterol assay (Sekisui Medical Co., Tokyo, Japan) on a Hitachi7600 analyzer (Hitachi Co., Tokyo, Japan) and observed a concentration of 121 mg/dL. We did not repeat the OSR6516 assay after a certain period. The specimen was checked for lipemic, hemolytic, and icteric indices to rule out other possible interferents. The patient was administered 30 g of ascorbic acid intravenously daily for 22 days prior to total cholesterol measurements. Dipstick analysis of the patient’s urine was performed by using an URiSCAN SUPER analyzer (YD Diagnostics, Seoul, Korea), which showed a value of 2+ (equivalent to 25 mg/dL) for ascorbic acid. Three additional cases characterized by spuriously low total cholesterol values owing to ascorbic acid interference are summarized in Table 1. Although we could not determine serum ascorbic acid levels, we detected its presence by using a urine dipstick assay in three of the four cases. Several studies recommend the addition of ascorbate oxidase to minimize ascorbic acid interference in assays to assess uric acid, triglyceride, oxalate, and cholesterol [5-8]. Most cholesterol assays in clinical laboratories use an enzymatic colorimetric method with the Trinder end-point reaction, in which hydrogen peroxide reacts with a chromogen via peroxidase to form a colored product; absorbance of this product is proportional to the concentration of total cholesterol in the sample [1]. Ascorbic acid interferes with peroxidase-based oxidation of the chromogen [6]. Since dehydroascorbic acid is already oxidized and has lost its reducing power, interference via ascorbic acid can be successfully prevented. Commercially available total cholesterol assays are summarized in Table 2. Ascorbate oxidase included in the Sekisui total cholesterol assay effectively converts serum