INTRODUCTION:

Nitrogen is a basic component of all living organisms, but animals are not able to metabolize the atmospheric molecular nitrogen and their nitrogen sources might be phyto-organic. Food is the major source of nitrogen, in the form of nitrates. A typical diet provides an average of 75 to 100 mg per day of nitrate. Vegetables such as spinach, celery, beets, lettuce, and root vegetables are responsible for most of the dietary intake. Release of nitrogen into air, water and soil leads to cascade of problems for human health and the environment. Particularly, nitrates are a health hazard because of their metabolic reduction to nitrites.

Once these are ingested, their conversion takes place immediately by oral bacteria in the saliva of people of all age groups, but in particular in the gastrointestinal tract of infants. Nitrites are then absorbed into the blood, may convert hemoglobin into methenoglobin¹.The concentration of nitrogen in drinking water is usually less than 10 mg/l, in the absence of bacterial contamination. In areas where nitrate contamination is higher, steps must be taken to lower it to avoid nitrate induced methemoglobinemia in infants. Ten mg/l of nitric nitrogen has been accepted by the U.S. Environmental Protection Agency as standard level in the Primary Drinking Water Regulations, mainly to protect young infants. They are able to convert approximately 10% of ingested nitrate to nitrite, compared to the 5% conversion in older children and adults. High nitrate-nitrogen concentration levels in drinking water can cause an illness, called “Blue-Baby”, in small children.

Nitrogen containing compounds released into environment can create serious problems, such as deterioration of water quality and is a potential hazard to human health. Furthermore, nitrate and nitrite have the potential to form nitrous compounds, which are potent carcinogens². Technologies currently adopted for the removal of nitrates comprises of; biological denitrification, reverse osmosis, ion exchange and electrochemical method.

Ion exchange and biological denitrification are the most frequently chosen technologies when nitrate removal is the only concern³. Conversely, the use of ion-exchange resins, as well as the recourse to reverse osmosis and electrodialysis, require frequent regeneration and lead to the production of secondary solutions. It is known that different factors such as the electrolyte, current density, pH and temperature of the electrolytic medium may affect the products and efficiency of electrochemical nitrate reduction⁴. These occur from the accumulation and concentration of undestroyed nitrates, and are of higher salinity, which requires them to be then disposed off. In addition, both the ion exchange resins and the reverse-osmosis membranes have high initial and operational costs.

Other more recent technologies for the reduction of nitrate include chemical, catalytic denitrification⁵ electrolytic denitrification and electrodeionization⁶ but all are still under development. Bioremediation is one of the most economic and eco-friendly technique which have been used in recent years. Bioremediation of congo red dye removal was studied by Kowsalya et al⁷. Debi Prasad Patnaik et al have studied about phytoremediation of heavy metals⁸.

MATERIALS AND METHODS:

Collection of Seaweed:

Marine algal species such as Centerocerasclavulatum, Enteromorpha flexuosa, Grateloupialithophila, Enteromorpha intestinalis, Ulva lactuca., and Chaetomorphaantennina were collected from Covelong, Chennai. Samples of Sargassum sp., Amphiroa sp., Ulva sp., and Hypnea sp. were collected from Kanyakumari whereas Chaetomorphaantennina from Puducherry. They were identified by Dr. M. Baluswamy, Professor and Head, Department of Plant biology and Plant Biotechnology, Madras Christian College, Tambaram, Chennai. The collected samples were thoroughly washed and were dried under sun light. Then the dried seaweeds were powdered and stored.

Preparation of Algal Extracts:

Estimated amount of powdered samples were soaked in 50 ml of non polar solvents such as benzene and chloroform for two days to extract the phytochemical constituents completely. They were also soaked in 50ml of polar solvents such as methanol, ethanol and water for 24 hrs and were extracted. Table.1 shows the various extracts of seaweed and their abbreviations used in this study for convenience. Table. 2 shows the raw seaweed powder which was used as an adsorbent and their abbreviations.

Table.1 Seaweed extracts using in this study

S. No	Name of Seaweed	Collection Place	Water	Chloroform	Benzene	Methanol	Ethanol
1	Enteromorpha intestinalis	Covelong	EIWC	EICC	EIBC	EIMC	EIEC
2	Ulva sp.	Kanyakumari	UWK	UCK	UBK	UMK	UEK
3	Hypnea sp.	Kanyakumari	HWK	HCK	HBK	HMK	HEK
4	Grateloupialithophila	Covelong	GLWC	GLCC	GLBC	GLMC	GLEC
5	Sargassum sp.	Kanyakumari	SWK	SCK	SBK	SMK	SEK
6	Chaetomorphaantennina	Puducherry	CAWP	CACP	CABP	CAMP	CAEP
7	Centerocerusclavulatum	Covelong	CCWC	CCCC	CCBC	CCMC	CCEC
8	Ulva lactuca	Covelong	ULWC	ULCC	ULBC	ULMC	ULEC
9	Enteromorpha flexuosa	Covelong	EFWC	EFCC	EFBC	EFMC	EFEC
10	Chaetomorphaattenina	Covelong	CAWC	CACC	CABC	CAMC	CAEC
11	Amphiroa sp.	Kanyakumari	AWK	ACK	ABK	AMK	AEK

Table. 2 Raw algal powder

S. No	Name of sample	Place	Abbreviation
1	Enteromorpha intestinalis	Covelong	EIC
2	Ulva sp.	Kanyakumari	UK
3	Hypnea sp.	Kanyakumari	HK
4	Grateloupialithophila	Covelong	GLC
5	Sargassum sp.	Kanyakumari	SK
6	Chaetomorphaantennina	Pondicherry	CAP
7	Centerocerusclavulatum	Covelong	CCC
8	Ulva lactuca	Covelong	ULC
9	Enteromorpha flexuosa	Covelong	EFC
10	Chaetomorphaattenina	Covelong	CAC
11	Amphiroa sp.	Kanyakumari	AK

Fig.1 Effluent collection site

Collection of Effluent:

Leather industry effluent was collected from the canal which was connected to the Peria Eri located at Nagalkeni village (Fig.1), Pallavaram, Chennai, Tamil Nadu, India and was preserved in a refrigerator to avoid further microbial contamination.

Phytochemical Analysis:

Phytochemical constituents of algal extracts were analysed qualitatively by standard procedure.

Estimation of nitrate concentration:

Nitrate concentration was analysed by standard procedure⁹.

RESULTS AND DISCUSSION:

Nitrate contamination of water resources has become an increasing problem globally due to the extensive use of nitrogen fertilizers and improper treatment of waste water from industrial sites¹⁰. Within the human body, nitrates may be reduced to nitrites that combine with haemoglobin to form methaemoglobin, which can be fatal to neonates¹¹. Because of its health risks, several methods that serve to reduce nitrate in drinking water have been presented^12-14. Conventional nitrate removal technologies include ion-exchange and reverse osmosis (RO). These processes¹⁵ cannot transfer nitrate into harmless compounds but only concentrate nitrate from water to high salinity waste stream that has very limited disposal options requiring further treatment. Recently, researchers have focused^16-22 on the electrochemical reduction of nitrate with different cathodes due to its high treatment efficiency, the small area occupied by the plant and relatively low investment costs. But the nitrate reduction rate slightly decreased with increasing NaCl concentration. Biological denitrification is a mature technology in wastewater treatment commonly used for the removal of nitrate from wastewater. However, it is an emerging technology for water treatment and without the addition of a carbon source, biological denitrification is difficult^23,24 and the problem of contamination by dead bacteria has to be solved to make such processes safe enough to utilize in drinking water treatment²⁵. Seaweeds were used as adsorbent and was used for treating textile industries effluent²⁶.In this work, all these disadvantages were overcome by using algal extracts for reducing nitrate content in the effluent. Untreated sample showed nitrate concentration of 540 mg/l. But the allowable limit of nitrate in the effluent by The National Environment standard is 10 mg/l.

The basic parameters of treated effluent were analysed after treatment and the results were categorized as A, B and C (Table.3) based on their percentage efficiency in the remediation of the effluent. The phytochemical constituents of all the extracts were also analysed to correlate with the results obtained from the effluent treatment with marine algae.

Phytochemical results revealed that the terpenoids were absent only in EIEC, UEK, SEK, GLEC, CCCC and CCWC. Phenol was found to be absent in the aqueous extracts such as CAWC, ULWC, AWK, UWK, CCWC, HWK, GLWC, SWK and also in methanolic extracts of Chaetomorpha sp., from Covelong and Amphoroa sp., from Kanyakumari. Since solubility of phenol is very less at room temperature. Aqueous extracts showed absence of carboxylic acid except in E.intestinalis collected from Covelong and in case of chloroform extracts it was found only in ULCC and CACP. Seven extracts of ethanol showed presence of carboxylic acid except EFEC, CCEC and AEK. All the aqueous extracts showed presence of amino acids and it was found to be absent in all benzene extracts except in EFBC and also absent in chloroform extract except in EFCC. Other extracts showed presence of amino acid except in ULEC, CACP, AEK, CAEC, HMK, SMK, CAMP, ULMC, EIMC, GLMC, and UMK. Phytochemical report showed that most of the extracts contained carbonyl compounds except in UEK, CAEP. HEK, AEK, CAEC, HMK, CAMP, CCMC, GLMC, UMK, CACP, CCCC, CACC, ACK, HWK, GLWC, EIWC. All the methanolic extracts showed presence of carbonyl compounds. In this study, carbonyl compounds were found to be the main constituent for the reduction of harmful compounds present in the effluent. (Data not shown here).

Table.3 Different Categories of Reduction

S. No	Category	Percentage Reduction	Remark
1	Category A	75 % - 100 %	Good
2	Category B	50 % - 75 %	Medium
3	Category C	Below 50 %	Poor

This study showed that good reduction in nitrate was obtained by ULBC, ULCC, EIBC, EFBC, EIMC, GLMC, EFEC, CCMC, ULEC, EFMC, ULMC, CCCC, EFCC, ULWC, CCEC, CCBC, HMK, UMK, EFWC (Fig.2) and medium reduction (Category B) was obtained GLBC, CAEC, CAMC, EIEC, HEK, GLEC, CABP, SEK, AMK, SMK, CAWP, CABC, HBK, CCWC, UBK, SBK, ABK, GLCC and SCK (Fig.3).

Poor reduction was found in EICC, HCK, UCK, CAWC, AWK, GLWC, CAMP, CACP, HWK, SWK, CACC, ACK, UEK, UWK, AEK, and EIWC (Fig 4). Algal powder was not able to reduce much nitrate content present in the effluent (Table.4).

Table.4 Concentration of nitrate in the treated effluent by algal powder

Extract	Nitrate content of Treated Effluent mg/l
HK	500
SK	450
GLC	540
UK	480
AK	400
CAC	375
CAP	375
EIC	510
EFK	500

In biological denitrification process, addition of carbon source was found to be difficult. In this work, carbonyl compounds may act as carbon source for reducing nitrate content, hence there is no need of adding additional carbon source to the effluent to reduce nitrate content.

CONCLUSION:

Degradation of harmful content present in effluent by biological method is an effective method and highly focused area in this era. Using of extracts of seaweed for treating waste water is an innovative technology. In future, these extracts may be used for treating other harmful contents present in Industrial as well as other waste waters which is polluting environment in higher level.

REFERENCES:

1. EPA (Environmental Protection Agency), Public health goals for nitrate and nitrite in drinking water, Pesticide and Environmental Toxicology Section, Office of Environmental Health Hazard Assessment, California. 1997.

2. Forman D. Nitrate exposure and human cancer. In: Bogardi, I., Kuzelka, R. (Eds.), Nitrate Contamination, RD NATO ASI Series. 1991; vol. G 30. Springer-Verlag.

3. Kapoora A, Viraraghavana T and Cullimoreb DR. Removal of heavy metals using the fungus Aspergillus niger, Bioresource Technology. 1999; 70: 95-103.

4. Englehardt D, De JD and Kalu EE. Electroreduction of nitrate and nitrite ion on a platinum-group-metal catalyst-modified carbon fiber electrodechrono amperometry and mechanism studies. J. Electrochem. Soc. 2000; 147: 4573–4579.

5. Murphy AP. Chemical removal of nitrate from water. Nature. 1991; 350: 223-225.

6. Salem K, Sandeaux J, Molenat J, Sandeaux R and Gavach C. Elimination of nitrate from drinking water by using electrochemical processes. Desalination. 1995; 101: 123-131.

7. Debi Prasad Pattanaik, Sanjeev Mishra, Anshuman Mishra, Sharmila S, Dhanalakshmi V, Anbuselvi S, and Jeyanthi Rebecca L. Phytoremediation of Mercury, Aluminium and Chromium using Raphanus sativus and Zea mays. International Journal of Biotechnology and Bioengineering Research. 2011; 2 (2): 277–286.

8. Kowsalya E, Sharmila S and Jeyanthi Rebbecca L. Removal of congo red dye from effluent sample using casurina leaves as a adsorbent. Journal of Chemical and Pharmaceutical Research. 2015; 7(9): 659-665.

9. SAP (Standard Analytical procedure), Hydrology Project, Govt. of India & Govt. of Netherlands.1999.

10. Ghafari S, Hasan M and Aroua MK. Bio-electrochemical removal of nitrate from water and wastewater – a review. Bioresour. Technol. 2008; 99: 3965–3974.

11. Gupta SK, Gupta AB, Gupta RC, Seth AK, Bassain JK and Gupta A. Recurrent acute respiratory tract infections in areas with high nitrate concentrations in drinking water. Environ. Health Perspect. 2000; 108 (4): 363–365.

12. Fernandez-Nava Y, Maranon E, Soons J and Castrillón L. Denitrification of wastewater containing high nitrate and calcium concentrations. Bioresour. Technol. 2008; 99: 7976–7981.

13. Virkutyte J and Jegatheesan V. Electro-Fenton, hydrogenotrophic and Fe2+ ions mediated TOC and nitrate removal from aquaculture system: different experimental strategies. Bioresour. Technol. 2009, 100: 2189–2197.

14. Wang Q, Feng C, Zhao Y and Hao C, Denitrification of nitrate contaminated groundwater with a fiber-based biofilm reactor. Bioresour. Technol. 2009, 100: 2222–2227.

15. Samatya S, Kabay N, Yüksel ü, Rda M and Yüksel M, Removal of nitrate from aqueous solution by nitrate selective ion exchange resins. React. Funct. Polym. 2006; 6 (11): 1206–1214.

16. Petri OA and Safonova TY. Electroreduction of nitrate and nitrite anions on platinum metals: a model process for elucidating the nature of the passivation by hydrogen adsorption. J. Electroanal. Chem. 1992; 331: 897–912.

17. Ureta-Zanartu S and Yanez C. Electroreduction of nitrate ion on Pt, Ir and on 70:30 Pt: Ir alloy. Electrochim. Acta. 1997; 42: 1725–1731.

18. Duarte HA, Jha K and Weidner JW. Electrochemical reduction of nitrates and nitrites in alkaline media in the presence of hexavalent chromium. J. Appl. Electrochem. 1998; 28: 811–817.

19. Huang CP, Wang HW and Chiu PC. Nitrate reduction by metallic iron. Water Res. 1998; 32: 2257–2264.

20. Vooys ACA, Santen RA and Veen JAR. Electrocatalytic reduction of NO_ 3 on palladium/copper electrodes. J. Mol. Catal. A Chem. 2000; 154: 203–215.

21. Wang Y, Qu J, Wu R and Lei P. The electrocatalytic reduction of nitrate in water on Pd/Sn-modified activated carbon fiber electrode. Water Res. 2006; 40: 1224–1232.

22. Li M, Feng CP, Zhang ZY and Sugiura N. Efficient electrochemical reduction of nitrate to nitrogen using Ti/IrO2–Pt anode and different cathodes. Electrochim. Acta. 2009; 54: 4600–4606.

23. Kesseru P, Kiss I, Bihari Z and Polya´k B. Biological denitrification in a continuous-flow pilot bioreactor containing immobilized Pseudomonas butanovora cells. Bioresour. Technol. 2003; 87: 75–80.

24. Dhamole PB, Nair RR, D’Souza SF and Lele SS. Simultaneous removal of carbon and nitrate in an airlift bioreactor. Bioresour. Technol. 2009; 100: 1082–1086.

25. Lei X and Maekawa T. Electrochemical treatment of anaerobic digestion effluent using a Ti/Pt–IrO2 electrode. Bioresour. Technol. 2007; 98: 3521–3525.

26. Amaraselvam K, Sharmila S and Jeyanthi Rebecca L. Removal of colour from textile industry effluent using seaweed. International Journal of Pharmacy & Technology. 2016; 8(3): 15740-15747.

27. Metcalf and Eddy. Waste water engineering, treatment, disposal, reuse, New York: McGraw-Hill. 1979.

Received on 20.02.2019 Modified on 13.03.2019

Research J. Pharm. and Tech. 2019; 12(7):3522-3526.

DOI: 10.5958/0974-360X.2019.00599.7