Biodegradation of Black Liquor

B.G. Gaikwad*

C.E. Division, National Chemical Laboratory, Pune-411008

*Corresponding Author E-mail: bg.gaikwad@ncl.res.in

ABSTRACT:

Bacterial strains and yeast strains were screened for treatment of black liquor. 17 bacterial cultures of genus –arthrobacter, cellulomonas, chainia, pseudomonas and glucanobacter were used. 19 yeast cultures having genus- kluyveromyces, pachysolen, pichia, rhodotorula, saccharomyces, candida and yerrowia were used for study. At pH 8.0, 58.33% sugar utilization, 16.14% phenol degradation, 37.81% decolorization was obtained by using Candida catanula NCIM 3337, C. brumptii 3402, Pseudomonas fluorescens 2653 respectively. 45.63% sugar utilization, 29.48% phenol degradation , 20.97% decolorization was observed by using Yerrowia lipolytica NCIM 3472, Pseudomonas fluorescens 2173, Yerrowia lipolytica 3589 respectively at pH 6.5. 63.49% sugar utilization was observed by Pseudomonas fluorescens 2141 at pH 4.5. 28.85% phenol degradation was done by Pachysolen tannophilus 3445 at pH 4.5. 63.26% decolorization was observed for Candida lambica 3532 at pH 4.5. Use of biomass for decolorization was attempted in this study. Fermentation of fruits was carried out. Sapota, fig, papaya, pineapple, banana, guava and tamarind were used for fermentation. Fermentation was done by using baker’s yeast. Residue from ethanol fermentation was used for decolorization of black liquor. Decolorization was obtained by residue from banana and tamarind fermentation residue was higher. It was 63.79 and 65.62% decolorization for banana and tamarind fermentation residue respectively.

KEYWORDS: Decolorization, candida, sugar, phenol, pH, pseudomonas etc.

INTRODUCTION:

Pollution from different industries is harmful for environment. Pollution from paper and pulp industry is a major problem, which is required to be resolved successfully Effluent from paper and pulp industry is called black liquor. It is also termed as sulphite waste liquor. Color and hazardous chemicals in black liquor are dangerous to aquatic livings. It can be used for drinking and other purpose. Many years researchers are working to solve it but still successful technology is in waiting list., Chai et al. (2014)¹ , Chandra and Abhishek (2011)², Oliveira et al. (2009)³, Jin (2008)⁴, Chandra et al. (2007)⁵ etc. have studied decolorization of BL. It is mentioned in results and discussions. Some researchers have studied use of biomass for color removal.

In this study we have reported here screening of strains for treatment of black liquor. Use of biomass for color removal is attempted in this work. Study has shown that it is possible to reduce pollution.

MATERIALS AND METHODS:

Chemicals and Cultures

Chemicals from Hi-Media, Mumbai and S.D. Fine Chemicals, Mumbai were used for experiment. Cultures were brought from NCIM (National Collection of Industrial Microorganism), Pune. Baker’s yeast was purchased from Local purchase.

Growth

Bacterial cultures were maintained on Nutrient –agar medium (beef extract 10, NaCl 5, peptone 10 g/L, pH 7.0, agar 20g/L.) at 4⁰C. Yeast cultures were maintained on MGYP-agar (Malt extract 0.3, glucose 1, yeast extract 0.3 and Peptone 0.5 %, pH adjusted to 6.5. 2.0% agar used). Bacterial cultures were grown on nutrient broth. Yeast cultures were grown on MGYP. Preinoculum was used for growth. Growth was carrried out at 28⁰C, 220 rpm for 24h. Cells were centrifuged and used for study.

Analysis:

Sugar was determined by DNSA method (Miller 1959)⁶. Nitroprusside method was used for phenol determination [Rodenas-Torralba et al. (2005)]⁷. Colour was measured at 465nm [Chandra and Abhishek (2011)]² at pH 8.0 and 6.5 study. While for pH 4.5, we have used wavelength 700nm.

Biodegradation:

1mL cell slurry was taken. 4mL 1:5 diluted black liquor (BL) was added. Cell concentration was 2.5%. pH of diluted BL was as per policy decided. It is kept at 28⁰C, 220rpm for 24 h. Sugar, phenol and colour was determined.

Experimental:

Screening of cultures

We have used 17 bacterial and 19 yeast cultures for screening study. List of cultures used is given in table 1. Study was performed using diluted (1:5) BL with different pH i.e. 8.0, 6.5 and 4.5. Results obtained were screened. We have selected 3 cultures showing which show maximum sugar utilization, phenol degradation and decolorization from each pH study. Results are shown in table 2-4. Use fermentation residue Fruits were fermented. Banana, grape, guava, sapota, fig, papaya, pineapple, and tamarind were used. Sugar concentration was 10-15%. Volume was 25mL. 2.5% baker’s yeast was used. Fermentation mixture was centrifuged after 24h and residue was collected. Residue contains yeast and fruit debris. This is used for removal of colour. Residue equivalent to 2.5% baker’s yeast was used for decolorization of. 20mL 1:5 diluted black liquor pH 4.5. It was stirred till decolorization and decolorization was measured.

RESULTS AND DISCUSSION:

We have done study at different pH i.e. 8.0, 6.5 and 4.5. Cultures having maximum results are discussed.

Biodegradation at pH 8.0:

We found Candida catenula NCIM 3337 has ulitized maximum sugar i.e. 58.33%. Candida brumptii 3402 degraded 16.14% phenol. Pseudomonas fluorescens 2653 has shown 37.81% decolorization (table 2).

Table 1: List of cultures

Genus	Culture and NCIM no.	Numbers of cultures
Arthrobactrer	Arthrobacter sp. 2934, 2935, 2937, 2938	4
Candida	C. lambica 3532, C. epicola 3367, C. glabrata 3237, C. catenulata 3337, C. bombi 3531, C. brumptii 3402	6
Cellulomonas	C. flavigena 2481	1
Chainia	Chainia sp. 2980	1
Gluconobacter	G. roseus 2524	1
Kluyveromyces	K. marxianus 3232, 3465	2
Pachysolen	P. tannophilus 3502, 3445	2
Pichia	P. guilliermondii 3251, P. farinosa 3461, P. stipitis 3498	3
Pseudomonus	P. fluorescens (5164, 2653, 2099, 2100, 2141, 2173); P. pictorum 2077; P. cruciviae 2004; P. sp. 2223, 2303	10
Rhodotorula	R. glutinis 3170	1
Saccharomyces	S. cerevisiae 3523, 3524, 3262	3
Yarrowia	Y. lipolytica 3472, 3589	2

Table 2: Biodegradation of BL at pH 8.0

Culture	NCIM no	Biodegradation %
Culture	NCIM no	Sugar utilization %	Phenol degradation %	Decolorization %
Candida catenulata	3337	58.33	12.23	0
Saccharomyces cerevisiae	3524	54.21	0	7.17
Pachysolen tannophilus	3445	50.60	0	0
Candida brumptii	3402	29.16	16.14	0
Candida lambica	3532	0	15.62	0
Candida bombii	3531	20.83	14.06	9.58
Pseudomonus fluorescens	5164	31.81	0	19.53
Pseudomonus fluorescens	2653	0	0	37.81
Saccharomyces cerevisiae	3523	36.14	0	19.55

Table 3: Biodegradation of BL at pH 6.5

Culture	NCIM no	Biodegradation %
Culture	NCIM no	Sugar utilization %	Phenol degradation %	Decolorization %
Arthrobacter sp.	2934	40	0	0
Pichia guilliermondii	3251	42.71	11.46	4.13
Yerrowia lipolytica	3472	45.63	12.64	0
Pseudomonus fluorescens	2173	0	29.48	0
Candida bombii	3531	0	19.26	0
Candida epicola	3367	30.09	17.78	0
Yerrowia lipolytica	3589	40	0	20.97
Pseudomonus fluorescens	2653	0	0	9.27
Pseudomonus fluorescens	5164	5.77	0	6.7

Figure 1: Comparison of biodegradation at different pH: Parameters on X-axis are S-sugar utilization; P-phenol degradation ; D- decolorization.

Sugar utilization(S): B- Candida catenula 3337-pH 8.0; C-Pseudomonas fluorescens 2141-pH- 4.5

Phenol degradation (P): D-Candida brumptii 3402-pH 8.0; E- Pseudomonas fluorescens 2173-

pH 6.5; F-Pachysolan tannophilus 3445-pH4.5

Decolorization (D): G- Pseudomonas fluorescens 2653-pH 8.0; H-Candida lambica 3532-pH 4.5

Table 4: Biodegradation of BL at pH 4.5

Culture	NCIM no	Biodegrdation %
Culture	NCIM no	Sugar utilization %	Phenol degradation %	Decolorization %
Pseudomonus fluorescens	2141	63.49	15.59	18.10
Pseudomonus sp.	2303	57.14	0	53.44
Arthrobacter sp.	2934	55.55	0	50
Pachysolen tannophilus	3445	0	28.85	39.79
Saccharomyces cerevisiae	3262	0	24.47	57.14
Candida glabrata	3237	0	24.16	5.43
Candida lambica	3532	0	9.52	63.26
Candida catenulata	3337	28.57	0	59.48
Pichia guilliermondii	3251	27.27	0	62.93

Results at lower pH are higher. Maximum decolorization was observed at pH 4.5. Candida lambica 3532 has shown highest decolorization. 79% decolorization was observed in 144h using bacteria by Chandra and Abhishek (2011)². Comamonas sp. B-9 has done 54% color reduction in 7days [Chai et al. (2014)]¹. Zheng et. al. (2014)⁸ has reported 35.3% decolorization by Comamonas sp. B-9. Bacillus pumilus CBMAI 0008 has shown 41.87% decolorization, whereas Paenibacillus sp. CBMAI 868 has shown 42.3% decolorization [Oliveira et al. (2009)]³. Chandra et al. (2007)⁵ have studied decolorization by mixed bacterial cultures. They reported 69% decolorization. Jin (2008)⁴ removed 93.15% color of BL using Aspergillus fumigatus JX-5.

Fermentation of fruits was carried out. Fermentation residue was used decolorization of 1:5 diluted black liquor pH 4.5. Results are presented in figure 2. It shows banana and tamarind fermentation residues have more decolorization. Maximum decolorization (65.62%) was observed by using tamarind fermentation residue. Clear liquid is obtained by treatment. This is due to adsorption of colored matter on yeast and fruit debris. Wang and Guo (2011)⁹ used beer yeast sludge for removal of color of dye. Biomass from amino acid fermentation was used for removal of color of dye by some researchers [Won et al. (2006)¹⁰; Vijayraghavan and Yun (2007)¹¹]. In our study we have used biomass and fruit debris to adsorb the coloring constituents. As some fruits contain pectin, which is sticky in nature, may help in attaching colouring matter. This will help in removing colour more effectively than only cells itself. The present work will be useful in developing feasible technology at higher scale where patent can be developed.

CONCLUSION:

Candida lambica NCIM 3532 has shown 63.26 % decolorization of 1:5 diluted black liquor at pH 4.5. This culture is best culture. Among fermentation residue study, we observed banana and tamarind are good. Tamarind has shown 65.62 % decolorization which is highest among the fruits used. Clear liquid is obtained by these two approaches.

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Received on 29.12.2014 Modified on 10.01.2015

Research J. Science and Tech. 7(1): Jan.-Mar. 2015; Page 47-51

DOI: 10.5958/2349-2988.2015.00008.X