Masaru Yamakoshi

An enzymatic method for the measurement of glycated albumin in biological samples

BACKGROUND In order to determine glycated albumin more easily and rapidly, we developed a new enzymatic method for glycated albumin in blood samples. METHODS The method involves use of albumin-specific proteinase, ketoamine oxidase and serum albumin assay reagent. In the assay, glycated albumin is hydrolyzed to glycated amino acids by proteinase digestion, and ketoamine oxidase oxidizes the glycated amino acids to produce hydrogen peroxide, which is quantitatively measured. Glycated albumin is calculated as the percentage of glycated albumin in total albumin. RESULTS The calibration curve for glycated albumin concentration was linear (r(p)=0.999) between 0.0 and 50.0 g/l and that for albumin concentration was linear (r(p)=0.999) between 0.0 and 60.0 g/l. The analytical recoveries of exogenous glycated albumin added to serum were 100-102.5%. The within-run and between-run CVs were 0.45-0.67% and 1.09-1.26%, respectively. This method was free from interference by bilirubin, chyle, glucose, globulins and labile intermediate. Weak interference by hemoglobin and ascorbic acid was observed. Glycated albumin detected by the present method was significantly correlated with glycated albumin detected by high-performance liquid-chromatographic (HPLC) method (serum: r(s)=0.989, plasma: r(p)=0.992). CONCLUSIONS This new enzymatic method is simple, rapid, allows multiple determinations and enables quantitative analysis of glycated albumin.

Determination of urinary myo-inositol concentration by an improved enzymatic cycling method using myo-inositol dehydrogenase from Flavobacterium sp.

BACKGROUND To determine myo-inositol more accurately, we improved the enzymatic cycling method. METHODS We screened myo-inositol dehydrogenase (MIDH; EC.1.1.1.18) from Flavobacterium sp., which was highly specific to myo-inositol. We measured urinary myo-inositol/creatinine ratio 2 h after 75-g oral glucose tolerance test (2 h MI) of 71 volunteers, and investigated the relationship between diabetes and urinary myo-inositol concentration. RESULTS The calibration curve was linear (r = 1.00) up to 2000 micromol/l, and the detection limit was 10 micromol/l. Within-run and between-run CVs were 0.5-1.1% and 0.4-1.3%, respectively. The 2 h MI of impaired fasting glycemia (IFG; 65.1 +/- 46.6 mg/g Cr, P < 0.005), impaired glucose tolerance (IGT; 85.0 +/- 73.7 mg/g Cr, P < 0.001) and diabetes (163.4 +/- 73.7 mg/g Cr, P < 0.0001) increased significantly compared with that of normal glucose tolerance (NGT; 24.0 +/- 14.4 mg/g Cr). From receiver operating characteristic analyses on 2 h MI, with 50 mg/g Cr as a tentative cutoff value to detect diabetes, the sensitivity and specificity were 100% and 77%, respectively. With 40 mg/g Cr as a tentative cutoff value to detect NGT, the sensitivity and specificity were 74% and 85%, respectively. CONCLUSIONS The myo-inositol measurement method demonstrated high specificity and yielded accurate results. The results of clinical trials suggested that 2 h MI could not only determine diabetes but also distinguish IFG and IGT from NGT.

Lucica® MI Urinary Myoinositol Kit

Abstract• Diabetes mellitus is a growing healthcare problem internationally, and poses a major burden from both a individual and societal perspective.• Diabetes causes potentially life-threatening complications that are preventable if the disease is detected early and appropriate interventions are put in place. Early detection is therefore imperative for preventing diabetes-related morbidity and mortality.• Current methods of detection, including the oral glucose tolerance test (OGTT), and measures of fasting plasma glucose, glycated hemoglobin (HbA1c), or glycated albumin, can be time-consuming and uncomfortable for patients.• Myoinositol can be measured in urine and has been found to be elevated in patients with diabetes and glucose intolerance; it has thus proven useful as a marker for the early detection of these conditions.• Lucica® MI is a diagnostic kit for the measurement of urinary myoinositol; it is used to detect glucose intolerance and diabetes mellitus at an early stage in disease progression. The test is based on an enzymatic method that uses liquid reagents requiring no preparation. Clinical trial results demonstrate that the test could be used to detect not only diabetes mellitus, but also to distinguish impaired fasting glucose and impaired glucose tolerance from normal glucose tolerance.