Effect of ATPase/ATP Synthetase inhibitors on As(III) biosorption by Aspergillus niger X₃₀₀

S. Ganguly*

Department of Biological Sciences, Sankrail Abhoy Charan High School (H.S), Sankrail, Howrah, West Bengal, India

ABSTRACT:

An experimental study was conducted to examine the involvement of metabolic energy in the biosorption of As(III) by Aspergillus niger X_300. Five major inhibitors of ATPase was added to the medium to inhibit ATPase. But no one exhibed adverse impact on the biosorption, suggesting there by, biosorption of As (III) by Aspergillus niger X₃₀₀ is not dependent on the cost of metabolic energy.

KEYWORDS: Experimental, biosorption, Aspergillus niger X_300, inhibitors, ATPase

INTRODUCTION:

ATPase are a group of enzymes that catalyze the hydrolysis of ATP into ADP into Pi[1].The decomposition releases energy which drives biochemical reactions [2].Thus ,any process requires metabolic energy cannot be operated when ATPase activity is blocked by inhibitors.The present study was intended to examine whether or not the biosorption of As(III) by Aspergillus niger X₃₀₀ requires metabolic energy.For this purpose , four major ATPase inhibitors( namely:2,3 butanidione 2-monoxime, 4 hydroxynonenal,bafilomycin A-1 and amiloride hydrochloride were examined.

MATERIALS AND METHODS:

Microorganism:

An experimentally developed As(III) resistant strain Aspergillus niger X₃₀₀ used throughout the study [3].

Physical condition for fungal growth:

The following growth conditions were used for optimum growth of the fungus: pH ,4.5; temperature, 30⁰C; spore density, 7x10⁸ spores or cells/ml; shaker speed, 200 rpm, volume of medium, 100ml, age of inoculum, 7 days and incubation period, 7 days[4].

Composition of synthetic medium of the following composition of the synthetic medium was used in the present study:

Glucose, 12%; (NH₄)₂SO₄, 0,8%; K₂HPO₄,0.01% ; KH₂PO₄, 0.01% ; CaCl₂.2H₂O, 0.03%; NaCl, 0.02%; MgSO₄.7H₂O, 0.03%; MnSO₄.4H₂O, 0.02 µg/ml and thiamine-HCl ,0.01µg/ml[5].

Estimation of As(III):

The concentration of As(III) in the broth was estimated by the method as reporte by Cernansky et al .(2007) [6].

Estimation of spore/ Dry cell weight:

Dry Spore/cell weight was estimated by the method as proposed by Shah et al. (2000)[7].

Addition of ATPase inhibitors:

Full grown fungus were incubated in the media containing As(III) and ATPase inhibitors ( 0.01-0.08 mM /ml).

Statistical analysis:

All data were expressed as mean± SEM ,where n=6. Data were analyzed by one way ANOVA followed by Dunett’s post hoc multiple comparison test considering p<0.05 as significant and p<0.01 as highly significant using prism 4.0 (Graph pad inc., USA).

RESULTS:

TABLE 1 : OPTIMIZATION OF CONTACT TIME BETWEEN Aspergillus niger X300 AND As(III)
Time (min)	As(III) concentration (mg/L)
	Initial	Final
00(Control)	1500	1500.0±8.213
30	1500	*1325.6±5.991
45	1500	**1176.5±6.212
60	1500	**921.8±6.732
90	1500	**776.6±7.213
105	1500	**622.2±8.116
120	1500	**421.8±9.163
135	1500	**176.4±8.881
150	1500	**100.6±6.732
ʘ165	1500	**89.3±8.991
180	1500	**89.3±7.771

(Values were expressed as Mean ± SEM , where n=6 ; *p<0.05, **p<0.01 when compared to control. ʘ stands for maximum biosorption)

From this Table 1 it is clear that maximum biosorption was resulted by the full grown fungus at 165 minutes of contact.

TABLE 2: EFFECT OF ATPase INHIBITORS ON As(III) BIOSORPTION BY Aspergillus niger X300
Inhibitors	Concentration(s) [Mm]	As(III) Biosorption (mg/L)
		Initial	Final
2, 3 butanidione 2-monoxime, 4 hydroxynonenal, bafilomycin A-1 and amiloride hydrochloride	0.01	1500	**89.3±0.991
	0.02	1500	**89.3±1.683
	0.03	1500	**89.3±0.882
	0.04	1500	**89.3±0.718
	0.05	1500	**89.3±0.996
	0.06	1500	**89.3±1.774
	0.07	1500	**89.3±0.913
	0.08	1500	**89.3±0.832

(Values were expressed as Mean ± SEM , where n=6 ; *p<0.05, **p<0.01 when compared to control.

From this table 2 , it can easily be stated that , among these four ATPase inhibitors examined , none of them exerted any sort of significant effect on As(III) biosorption by this fungus. It can therefore be concluded that , As(III) biosorption by this fungus is a passive process,it doesnot any sort of metabolic energy.

ACKNOWLEDGEMENT:

I express my sincere gratitude to the Department of Chemical Engineering, University of Calcutta, Bose Institute, Kolkata, Indian Institute of Chemical Biology (IICB), Kolkata, Department of Food technology and Biochemical Engineering, Jadavpur University, Kolkata for their kind cooperation.

REFERENCES:

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2. Sarkadi B, Price EM, Boucher RC, German UA and Scarborough GA, Expression of the human multi drug resistant c-DNA in insect cells generates a high activity drug stimulated membrane ATPase, Journal of Biological Chemistry, 267, 1992 : 4854-4858.

3. Ganguly S, Selection of suitable fungus , development of Arsenic resistant strain and kinetic analysis for As(III) biosorption, Indian Journal of Applied Research , 3,2013:9-10.

4. Ganguly S , Optimization of Physical Conditions for As(III) Biosorption by Aspergillus niger X₃₀₀ , International Journal of Pharma medicine and Biological Sciences , 2 , 2013:46-49.

5. Ganguly S, Selection of Suitable Synthetic Medium For As(III) Biosorption By Aspergillus niger X₃₀₀, Asian Journal Research in Chemistry (in press).

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Received on 12.04.2013

Modified on 16.05.2013

Accepted on 11.06.2013

© A&V Publication all right reserved

Research J. Pharmacology and Pharmacodynamics. 5(4): July–August 2013, 205-206