Efficient Enzyme Immobilization by Combining Adsorption and Cellulose Membrane Coating †

doi:10.7503/cjcu20190185

Abstract

Abstract:

The papain was used as a model enzyme to develop an immobilized enzyme method based on combination of adsorption and cellulose membrane coating. The type of adsorption carries, cellulose category and the solvent type for dissolving cellulose, protectant category and concentration, drying mode and time were systematically optimized. The best adsorption-cellulose coating combined immobilization process was adopting diatomite or HPD-417(macroporous resin) as adsorption carrier, methylcellulose(molecular weight 40000—50000) acetone solution as coating solution, 6%(mass fraction) polyethylene glycol or maltose as the protectant for membrane coating, and drying at 4 ℃ for 9 h. The prepared immobilized papain showed high enzyme activity recovery rate, which attained to 96.50% for diatomite adsorbing-cellulose coating immobilized papain and 93.92% for HPD-417 adsorbing-cellulose coating immobilized papain. Finally, the properties of the immobilized enzyme were investigated. The results showed that the adsorbing-cellulose coating immobilized papain had better thermal stability. After being stored at 80 ℃ for 12 h, the residual activity of the immobilized enzyme remained about 90%. In the range of pH=4.5—9.5, the stability of the immobilized enzyme was better. After being continuously used for 9 batches, the residual activity of the immobilized enzyme remained about 95%.

Key words: Enzyme immobilization, Adsorption, Cellulose coating, Combined immobilization, Methyl cellulose, Protectant, Papain

CLC Number:

O629
Q556

TrendMD:

WU Rong, DONG Qihui, SUN Yiyi, SU Erzheng. Efficient Enzyme Immobilization by Combining Adsorption and Cellulose Membrane Coating ^†[J]. Chem. J. Chinese Universities, 2019, 40(9): 1888.

Figures/Tables 10

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Adsorption material	Protein immobilization yield(%)	Immobilized enzyme activity/(U·g^-1)	Enzyme activity recovery rate(%)
D130	94.91±0.07	11.68±0.040	41.91±1.57
HPD-450	99.01±0.06	12.18±0.016	40.49±0.27
ADS-17	95.95±0.18	25.68±0.054	66.51±0.59
HPD-417	95.11±0.07	28.57±0.032	60.33±0.46
D101	96.21±0.33	9.38±0.042	30.38±0.34
AB-8	95.65±0.03	10.34±0.12	39.63±0.90
Activated carbon(30 mesh)	34.12±0.49	6.01±0.012	43.29±1.34
Activated carbon(40 mesh)	35.49±0.33	6.09±0.022	37.64±0.46
Zeolite	38.69±0.25	9.44±0.026	42.96±0.61
Kaolin	55.55±0.25	29.07±0.016	98.13±1.12
Imvite	100.00	32.76±0.026	99.47±1.28
Attapulgite	58.92±0.40	13.05±0.032	53.08±1.76
Diatomite	51.47±0.09	30.61±0.022	98.38±2.62

Adsorption material	Protein immobilization yield(%)	Immobilized enzyme activity/(U·g^-1)	Enzyme activity recovery rate(%)
D130	94.91±0.07	11.68±0.040	41.91±1.57
HPD-450	99.01±0.06	12.18±0.016	40.49±0.27
ADS-17	95.95±0.18	25.68±0.054	66.51±0.59
HPD-417	95.11±0.07	28.57±0.032	60.33±0.46
D101	96.21±0.33	9.38±0.042	30.38±0.34
AB-8	95.65±0.03	10.34±0.12	39.63±0.90
Activated carbon(30 mesh)	34.12±0.49	6.01±0.012	43.29±1.34
Activated carbon(40 mesh)	35.49±0.33	6.09±0.022	37.64±0.46
Zeolite	38.69±0.25	9.44±0.026	42.96±0.61
Kaolin	55.55±0.25	29.07±0.016	98.13±1.12
Imvite	100.00	32.76±0.026	99.47±1.28
Attapulgite	58.92±0.40	13.05±0.032	53.08±1.76
Diatomite	51.47±0.09	30.61±0.022	98.38±2.62

Cellulose	Adsorption material	Immobilized enzyme activity by adsorption/(U·g^-1)	Immobilized enzyme activity after coating/(U·g^-1)	Residual enzyme activity after coating(%)
Cellulose acetate	Diatomite	30.94±0.27	18.68±0.88	60.37±0.73
	ADS-17	25.56±1.92	17.60±0.93	68.88±0.39
	HPD-417	27.66±0.83	19.91±1.21	71.99±1.38
Diacetate cellulose	Diatomite	30.94±0.27	20.90±0.96	67.54±1.91
	ADS-17	25.56±1.92	18.76±0.77	73.41±1.28
	HPD-417	27.66±0.83	20.78±0.35	75.11±0.93
Methyl cellulose	Diatomite	30.94±0.27	21.03±0.93	67.97±1.36
40000—50000	ADS-17	25.56±1.92	19.26±1.24	75.34±1.21
	HPD-417	27.66±0.83	21.55±0.82	77.91±0.63
Methyl cellulose	Diatomite	30.94±0.27	21.47±0.69	69.38±1.91
12000—13000	ADS-17	25.56±1.92	20.36±0.79	79.67±0.86
	HPD-417	27.66±0.83	21.18±0.83	76.56±1.22

Cellulose	Adsorption material	Immobilized enzyme activity by adsorption/(U·g^-1)	Immobilized enzyme activity after coating/(U·g^-1)	Residual enzyme activity after coating(%)
Cellulose acetate	Diatomite	30.94±0.27	18.68±0.88	60.37±0.73
	ADS-17	25.56±1.92	17.60±0.93	68.88±0.39
	HPD-417	27.66±0.83	19.91±1.21	71.99±1.38
Diacetate cellulose	Diatomite	30.94±0.27	20.90±0.96	67.54±1.91
	ADS-17	25.56±1.92	18.76±0.77	73.41±1.28
	HPD-417	27.66±0.83	20.78±0.35	75.11±0.93
Methyl cellulose	Diatomite	30.94±0.27	21.03±0.93	67.97±1.36
40000—50000	ADS-17	25.56±1.92	19.26±1.24	75.34±1.21
	HPD-417	27.66±0.83	21.55±0.82	77.91±0.63
Methyl cellulose	Diatomite	30.94±0.27	21.47±0.69	69.38±1.91
12000—13000	ADS-17	25.56±1.92	20.36±0.79	79.67±0.86
	HPD-417	27.66±0.83	21.18±0.83	76.56±1.22

Cellulose	Adsorption material	Immobilized enzyme activity by adsorption/(U·g^-1)	Immobilized enzyme activity after coating/(U·g^-1)	Residual enzyme activity after coating(%)
Cellulose acetate	Diatomite	30.94±0.27	19.64±0.91	63.47±0.72
	ADS-17	25.56±1.92	17.24±1.21	67.47±0.69
	HPD-417	27.66±0.83	19.29±0.66	69.74±1.35
Diacetate cellulose	Diatomite	30.94±0.27	21.02±0.71	67.93±1.22
	ADS-17	25.56±1.92	17.30±1.92	67.69±0.82
	HPD-417	27.66±0.83	19.89±1.25	71.89±0.69
Methylcellulose	Diatomite	30.94±0.27	21.44±0.52	69.35±0.77
40000—50000	ADS-17	25.56±1.92	18.79±1.12	73.51±0.96
	HPD-417	27.66±0.83	20.31±0.57	73.42±1.02
Methylcellulose	Diatomite	30.94±0.27	21.60±0.82	69.81±1.28
12000—13000	ADS-17	25.56±1.92	18.56±0.72	72.61±0.65
	HPD-417	27.66±0.83	21.04±0.91	76.07±0.74