Immobilization of horseradish peroxidase on PMMA nanofibers incorporated with nanodiamond

Figures & data

Table 1. The effect of nanodiamond concentration, which adsorbed on PMMA nanofibers and pH medium on the immobilization efficiency of HRP.

Nanodiamond %	Immobilization efficiency%
	pH 6.0	pH 7.0	pH 8.0
0	8	16	12
1	23	63	42
5	25	45	33
10	42	96	62
15	17	41	28
20	9	39	16

Each value represents the mean of two experiments.

Figure 1. ATR-FT-IR spectra of ND, nfPMMA, nfPMMA-ND and nfPMMA-ND-HRP samples.

Figure 2. SEM Images of (a) nfPMMA low magnification (b) nfPMMA high magnification (c) nfPMMA-ND low magnification (d) nfPMMA-ND high magnification.

Figure 3. SEM Images of (e) nfPMMA-ND-HRP low magnification (f) nfPMMA-ND-HRP high magnification.

Figure 4. TEM Images of (a) nfPMMA-ND-HRP with 10 µm scale (b) nfPMMA-ND-HRP 0.5 with µm scale (c) nfPMMA-ND-HRP with 50 nm scale.

Figure 5. Reuse of immobilized HRP. Each point represents the mean of three experiments ± SE.

Figure 6. Optimum pH of soluble and immobilized HRP. The enzyme activity was measured at different pH’s ranging from 4.0 to 9.0. Each point represents the average of two experiments.

Figure 7. Optimum temperature of soluble and immobilized HRP. The enzyme activity was measured at different temperatures ranging from 30 to 80  C. Each point represents the average of two experiments.

Figure 8. Effect of temperature on the thermal stability ofsoluble HRP and immobilized HRP. Each point represents the average of two experiments.

Figure 9. Lineweaver-Burk plots relating soluble HRP and immobilized HRP reaction velocity to guaiacol (a) and H₂O₂ (b) concentrations. Each point represents the average of two experiments.

Table 2. The substrate specificity of soluble HRP and immobilized HRP.

Substrate	Relative activity %
	Soluble HRP	Immobilized HRP
Guaiacol	100	100
o-Phenylenediamine	69	312
o-Dianisidine	34	605
p-Aminoantipyrine	25	36
Pyrogallol	17	40
ABTS	2	9

Each value represents the mean of two experiments.

Table 3. The effect of 5 mM metal ions on the activity of soluble HRP and immobilized HRP.

Metal ion	Relative activity%
	Soluble HRP	Immobilized HRP
Control	100	100
Co^2+	73	97
Ca^2+	109	155
Pb^2+	75	87
Ni^2+	66	80
Zn^2+	65	83
Hg^2+	16	83
Cu^2+	89	289
Fe^3+	168	293
Cd^2+	86	92

Each value represents the mean of two experiments.

Table 4. The effect of chemical compounds on soluble HRP and immobilized HRP.

Chemical compounds	Relative activity%
	concentration	Soluble HRP	Immobilized HRP
Control		100	100
Urea	2M	60	82
	4M	24	50
Isopropanol	5%	39	77
	10%	31	87
n-Butanol	5%	75	92
	10%	63	76
Heptane	5%	68	96
	10%	55	84

Each value represents the mean of two experiments.

Figure 10. The effect of trypsin concentration on the activity of soluble HRP and immobilized HRP. Each point represents the average of two experiments.