A new affinity method for purification of bovine testicular hyaluronidase enzyme and an investigation of the effects of some compounds on this enzyme

Figures & data

Figure 1. Optimization of pH on BTH (after 60% (NH₄)₂SO₄ precipitation) with m-anisidine compound.

Figure 2. Protein values at 280 nm and BTH activities in elution fractions.

Figure 3. SDS-PAGE of BTH. The pooled fractions from ammonium sulfate precipitation and affinity chromatography (sepharose-4B, l-tyrosine and m-anisidine) were analyzed by SDS-PAGE (10% and 3%) and revealed by Coomassie blue staining. Experimental conditions were as described in the method.

Table 1. Summary of the purification of BTH.

Step	Volume (ml)	Activity (U ml^−1)	Total activity (U)	Protein amount (mg ml^−1)	Total protein (mg)	Specific activity (U mg^−1)	Overall yield (%)	Overall purification (fold)
Bovine testis serum	85	9.67	821.95	3.15	268	3.07	100	–
Ammonium sulfate fractionation	6	17.8	106.8	4.73	28.39	3.76	12.99	1.23
Affinity chromatography	2	69.67	139.34	0.02	0.04	3317.62	16.95	881.78

Figure 4. Activity (%) graphs of mepiquat chloride (1,1-dimethylpiperidinium chloride (PIX), 2-naphthoxyacetic acid (β-NOA); and gibberellic acid on BTH.

Table 2. IC₅₀ values of 3-methoxyaniline; (m-anisidine) mepiquat chloride (1,1-dimethylpiperidinium chloride (PIX); 2-naphthoxyacetic acid (β-NOA); and gibberellic acid.

Compound name	IC50 (mg/mL)
1,1-Dimethyl piperidinium chloride	9079
β-Naphthoxyacetic acid	3534
Gibberellic acid	0718
m-Anisidine	1.73