β-d-N-Acetylglucosaminidase isozymes were separated and assayed in the plasma of control healthy individuals and pregnant women by an automated method consisting in chromatofocusing on polybuffer exchanger PBE-94 column, flow-through fluorimetric determination of activity and computer assisted quantification. Under the established optimal conditions the method fractionated β-d-N-acetylglucosaminidase into four isozymes, A, I2, I1 and B, with the analytical coefficients of variation of 1.8, 2.2, 6.4 and 4.1%, respectively. Duration of a single analysis was 25 min including washing, and 10-15 successive runs could be performed on the same column with good reproducibility. A linear activity response was observed from 1-5 μl of plasma (depending on the individual isozyme) to 50 μl, and the detection limit was 0.016 mUnits. Isozyme A was heat labile. Upon sialidase treatment, isozymes A, I2 and I1 released sialic acid and were eluted from the column at less acidic pHs. In healthy individuals isozymes A, I2, I1 and B covered about 62.8, 6.9, 15.0 and 15.1% of the total β-d-N-acetyl-glucosaminidase activity, respectively. During pregnancy the plasma concentration of all isozymes increased. Isozyme I2 showed the highest enhancement (30-fold), followed by I1 (8-fold), B (5.6-fold) and A (3-fold). Interruption of pregnancy by either physiological delivery or ambulatory abortion was followed by a sharp fall of the concentration of all isozymes reaching, in a few days, the control levels.
- Automated procedure
- Human plasma
- β-d-N-Acetylglucosaminidase isozyme
ASJC Scopus subject areas
- Clinical Biochemistry