Remediation and recycling of chromium from tannery wastewater using combined chemical–biological treatment system

Process Safety and Environmental Protection 1 0 4 ( 2 0 1 6 ) 1–10

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Remediation and recycling of chromium from tannery wastewater using combined chemical–biological treatment system Engy Ahmed a,b,∗ , Hesham M. Abdulla c , Amr H. Mohamed d , Ahmed D. El-Bassuony c a

Department of Geological Sciences, Stockholm University, Stockholm, Sweden Science for Life Laboratory, Stockholm University, Solna, Sweden c Botany Department, Faculty of Science, Suez Canal University, Ismailia, Egypt d Housing and Building National Research Center (HBNRC), Cairo, Egypt b

a r t i c l e

i n f o

a b s t r a c t

Article history:

Tannery wastewater containing chromium (Cr) is one of the most serious problems in leather

Received 14 March 2016

industry. In order to develop an effective and eco-friendly treatment technology, a combined

Received in revised form 2 August

chemical–biological treatment system was performed for Cr remediation and recycling. The

2016

aim of the present study is to design a laboratory scale system using chemical precipita-

Accepted 3 August 2016

tion of Cr(III) combined with biological removal of Cr(VI) from tannery wastewater, and to

Available online 11 August 2016

investigate the possibility of recycling the recovered Cr(III) in the tanning industry.

Keywords:

nomycete strain Kitasatosporia sp. was used in microcosm studies for Cr(VI) bioremoval.

Chemical precipitation of Cr(III) was carried out using lime and cement dust. The actiActinomycetes

Moreover, parameters such as type of porous medium, inoculum size, flow rate and culture

Cr(VI)

conditions were investigated. The precipitated Cr(III) that was recovered from the chemical

Cr(III)

precipitation stage was recycled in the leather tanning industry.

Kitasatosporia Microcosm

Our findings indicate that the maximum Cr(III) precipitation (98%) was achieved using 2 g/100 mL of lime and 2 h of settling rate. On the other hand, microcosm columns using sand that was inoculated with induced culture (OD600 = 2.43) and flow rate (2 mL/min) gave

Reuse

the maximum recovery (99%) of Cr(VI). The experimental Cr(III) was successfully recycled in the tanning process and the experimental leathers showed comparable properties as same as the leathers tanned with commercial Cr(III). Thus, we concluded that using combined chemical–biological treatment system for Cr remediation from tanning wastewater together with recycling process for the recovered Cr(III) is a promising strategy for economic and environmental friendly tanning industry. © 2016 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

1.

Introduction

Chromium (Cr) is one of the major environmental pollutants coming from various industrial activities, such as leather tanning, electrolytic plating, metal finishing, petroleum refining and etc. (Acosta-Rodríguez

et al., 2015). There are two stable forms of chromium in the environment, trivalent (Cr(III)) and hexavalent (Cr(VI)) chromium. However, Cr(VI) is more toxic than Cr(III) due to its high solubility and mobility in soil and aquatic environments, and high permeability through biological membranes (Marsh and McInerney, 2001; Shukla and Rai, 2006). Thus, it is considered as a risk pollutant by the United States Environmental Protection Agency (EPA: www.epa.gov).

∗

Corresponding author at: Department of Geological Sciences, Stockholm University, SE-10691 Stockholm, Sweden. E-mail address: [email protected] (E. Ahmed). http://dx.doi.org/10.1016/j.psep.2016.08.004 0957-5820/© 2016 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

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Tanning industry worldwide generates approximately 40 million L of wastewater containing Cr every year (Saranraj and Sujitha, 2013). In most of the countries, the tanning wastewater is discharged without proper treatment into the sewerage system causing serious environmental impact. Treatment of Cr polluted wastewater is mostly dependent on physico-chemical methods, such as chemical precipitation, reverse osmosis, membrane processes and adsorption (Ludvik, 2000; Kocaoba and Akin, 2002). However, chemical precipitation has been considered to be one of the most effective techniques to separate heavy metals from industrial wastewater (Barakat, 2011). Chemical precipitation is a process in which soluble metals and inorganics are converted to relatively insoluble forms (precipitates) by the addition of precipitating agents (Bennett et al., 1981). Substances used habitually to promote the precipitation are: calcium hydroxide, sodium hydroxide, magnesium oxide or calcium magnesium carbonate (Tsugita and Ellis, 1981; Hintermeyer et al., 2008). Subsequently the precipitated chromium hydroxide can be re-dissolved by acidification and be easily reused in the tanning process (Kanagaraj et al., 2008). Supernatant from Cr precipitation process is relatively free of Cr(III). The biological treatment of such supernatant solution is one of the successful approaches to remove Cr(VI) (Lovely and Coates, 1997; Langerwerf, 1999; Rittmann et al., 2004). However, various fungal and bacterial species are reported for Cr(VI) bioremoval from industrial wastewater (e.g., Congeevarama et al., 2007; Das and Santra, 2012; Vermaa et al., 2015), there are a few studies concern the abilities of actinomycetes (More et al., 2001; Abdulla et al., 2010, 2011). The metabolic diversity and genomic characteristics of actinomycetes make them significant agents for bioremoval of metals from contaminated environments (Polti et al., 2007). Combined chemical–biological treatment of Cr polluted wastewater became more economic and environmental friendly strategy than either physico-chemical or biological treatment alone (Abdulla et al., 2010, 2011). Such combined treatment leaves a lower concentration of Cr in the effluent than the other methods. Combined chemical–biological treatment of Cr has been approached from three directions (Ayres et al., 1994; Goswami and Mazumder, 2014): (a) biological treatment followed by chemical treatment as a polishing step, (b) chemical treatment at levels where the stoichiometry is effective and economic for treatment, followed by biological treatment as a polishing step, (c) chemical precipitation followed by biological treatment was also approached in a staged reactor system. The present study aims to design a laboratory scale system using a combined chemical–biological treatment to remove Cr(III) and Cr(VI) from tannery wastewater, and to investigate the possibility of recycling the recovered Cr(III) in the tanning industry.

2.

Materials and methods

2.1. Sampling and characterization of tanning wastewater Wastewater samples were collected in clean 1000 mL polyethylene bottles from the outlet point of the chromium tanning stage of selected tanneries in Old Cairo, Egypt. The bottles were kept immediately in 4 ◦ C until further analysis. The characteristics of the water samples, such as pH, biochemical oxygen demand (BOD), chemical oxygen demand (COD), total suspended solids (TSS), sulfate, turbidity, salinity and electric conductivity (EC) were determined according to standard methods for the examination of water and wastewater (APHA, 1998). The other measurements were carried out as following: a) Analytical estimation of Cr(VI) and Cr(III) The concentration of Cr(VI) was determined spectrophotometrically using 1,5-diphenylcarbazide as complexing agent

(Bartlett and James, 1996). Diphenylcarbazide stock solution (0.25% w/v in 50% acetone) was prepared. A volume of 15 mL of each sample was pipetted out into 25 mL standard flasks. Then 2 mL of H2 SO4 (3 M) was added to the sample followed by 1 mL of diphenylcarbazide and the total volume was made up to 25 mL using distilled water. The solution was allowed to stand for 10 min, after which the absorbance of the purple-colored solution was measured using spectrophotometer (CECIL, CE 393, Series 2, UK) at 540 nm. Cr(VI) concentration was extrapolated from a standard curve prepared from standard solutions of potassium dichromate. In order to estimate the concentration of total chromium, Cr(III) in samples was completely oxidized to Cr(VI) (Chandrachekhara et al., 2015; Onchoke and Sasu, 2016). Methyl orange was used as an indicator. A volume of 15 mL of each sample was pipetted out into 25 mL standard flasks. Then 1 mL of H2 SO4 was added and the total volume was made up to 40 mL using distilled water. The mixture was heated till boiling. Then 2 drops of KMnO4 were added to give a dark red color followed by 1 mL of NaN3 and continue boiling gently for 30 s. The total chromium concentration was estimated using the same colorimetric method as for Cr(VI) estimation. The concentration of Cr(III) was obtained by calculating the difference between the values of total Cr concentration and Cr(VI) concentration as estimated by the above procedures (Bartlett, 1991).

b) Enumeration of microbial cells in tanning wastewater

One milliliter of wastewater samples were serially diluted in phosphate buffer and 0.1 mL of the suitable dilutions were plated onto duplicates of the appropriate media using spread plate technique. Dilutions up to 10−3 and 10−4 were used for enumeration of actinomycetes on starch casein agar amended with cyclohexamide (0.05 g/L) to inhibit fungal growth. Plates were incubated at 28 ◦ C for 10–14 days. Bacteria were enumerated using nutrient agar; plating was performed from dilutions 10−3 and 10−4 and was incubated at 37 ◦ C for 24–36 h. Fungi were enumerated using Czapek–Dox agar; plating was performed from dilutions 10−3 and 10−4 and plates were incubated at 28 ◦ C for 4 days.

2.2. Set-up of combined chemical–biological treatment system for tannery wastewater The treatment system was designed in two stages, (a) chemical precipitation of Cr(III) from raw tannery wastewater, and (b) biological removal of Cr(VI) from pre-treated tanning wastewater (the supernatant of the precipitation stage).

2.2.1.

Chemical precipitation of Cr(III)

The chemical precipitation was performed according to Abdulla et al. (2010). Thirteen glass jars were filled with 100 mL tannery wastewater. Lime and cement dust were added with different concentrations from 0.5 g to 3 g per 100 mL. The stirring was continued for 10 min with rapid mixing of 100 rpm by orbital shaking, followed by slow mixing for 5 min at 40 rpm. The jars were allowed to settle then samples were taken from the supernatant for Cr(III) analysis (according to the above method) at intervals of 30 min for 3 h.

Process Safety and Environmental Protection 1 0 4 ( 2 0 1 6 ) 1–10

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2.2.2. Construction of laboratory-scale microcosms for Cr(VI) bioremoval

2.3.2. Analyses of physical and organoleptic properties of leather

Microcosms were designed as vertical glass columns (4 × 25 cm) filled with porous media (sand or gravel). The percolating gravel microcosm contained 274 g of gravel (2–5 mm diameter), while the sand microcosm columns contained 350 g of sand (1.25 mm diameter). Each column was connected to a polyethylene container contained 250 mL of pre-treated tanning wastewater and the flow rate regulated by using a stopper. The actinomycete Kitasatosporia sp. that was used in this study was the most effective Cr resistant species isolated from Cr contaminated industrial wastewater in Egypt (Ahmed, 2010). The actinomycete isolate was grown overnight in water peptone broth with and without 100 mg/L chromate (Cr induced and non-induced), then the harvested biomass was washed and re-suspended in distilled water to be inoculated in microcosm columns. The inoculation was performed by circulating the suspended culture through the microcosm columns for 3–4 times, and then the inoculated columns were kept for 24 h to assure the adsorption of the biofilm on the porous medium. To optimize the experimental conditions of the microcosms, parameters like type of porous medium (sand and gravel), flow rate (between 1 and 3 mL/min), inoculum size between OD = 0.406 and 2.43 equivalent to (693–4161 cfu/mg) and culture conditions (Cr induced and non-induced) have been studied. Cr(VI) bioremoval efficiency was determined for zero time (the first drop of treated wastewater obtained from the column), 30, 60, 90, 120 and 150 min intervals and was calculated on the basis of residual Cr(VI) concentration in the wastewater as estimated by the diphenylcarbazide method. All the microcosm studies were designed in duplicates and control columns (un-inoculated) were included. In addition, the characteristics of the treated wastewater from the combined chemical–biological treatment system were measured according to standard methods for the examination of water and wastewater (APHA, 1998).

Tanning quality of the leather samples with dimensions of 30 × 30 cm were assessed by testing the leather physical and organoleptic properties according to Kanagaraj et al. (2008) including: shrinkage temperature of leather, tensile strength, percentage of elongation at break, tear strength and permeability test. General appearance of the leathers was also examined by experienced tanners. The quality of experimental and control leathers were assessed according to ISO and Egyptian standards.

2.3.

Reuse of recovered Cr(III) in tanning industry

The precipitated Cr(III) that was recovered from the chemical precipitation stage was re-dissolved with sulfuric acid (pH 2.7–3.0) and reused in the leather tanning industry. Cow skins were used for tanning studies. The standard Cr tanning process was applied for all experiments. The cow skins were tanned with the following types of chromium, (a) experimental chromium (recovered Cr(III) from the present study), (b) commercial chromium sulfate (control), (c) 50% of the recovered chromium mixed with 50% of commercial chromium sulfate (mixed). The properties of the tanned leathers were assessed in the wet blue stage at Leather Technology Centre Lab (LTC Lab), Cairo as follows.

2.3.1.

Estimation of chromium concentration in the leather

Leather samples from the three different Cr tanning experiments were analyzed for Cr content. One gram of wet blue leather was cut into small pieces. The small leather pieces were taken in a crucible and dried in air oven at 80–100 ◦ C for 5–6 h. After cooling in desiccator for 30 min, the pieces were weighed. Then the dried leather pieces were digested using acid mixture (perchloric acid, nitric acid and sulfuric acid in the ratio of 11.5:5.0:3.5). The Cr content in the oxidized mixture was estimated by using the diphenylcarbazide detection method according to Bartlett and James (1996).

2.4.

Statistical analysis

The data were normalized, and the single/multifactor analysis of variance (ANOVA) and correlation were analyzed.

3.

Results

3.1.

Characterization of tannery wastewater

Wastewater samples were collected from Cr tanning stream in intervals from February to December. There was no significant difference between the characteristics of the wastewater through the year. In general, the color of tanning wastewater was dark blue and very acidic. The chemical, physical and microbiological properties of wastewater showed that the tannery effluent exceeded the standard pollution limits that were established by the Egyptian regulations for industrial wastewater (Table 1).

3.2. Chemical precipitation of Cr(III) from tannery wastewater The chemical precipitation of Cr(III) has been performed by two precipitating agents, lime and cement dust. Our finding showed that the type, dose and settling time of the precipitating agent affect the precipitation efficiency of Cr(III) (Fig. 1). The maximum Cr(III) precipitation (98–99%) was observed by using high dose of lime (2–3 g/100 mL), while the precipitation efficiency was decreased (95.5–97%) by decreasing the lime dose to 0.5–1.5 g/100 mL. Moreover, the maximum Cr(III) precipitation (98%) was observed after 120 min of settling. Settling time higher than 120 min did not show any significant difference (P-value > 0.05) with regard to Cr(III) precipitation. On the other hand, Cr(III) precipitation by cement dust was very low (between 1.5 and 2.6% at 2–3 g/100 mL) and settling time had no effect on the precipitation efficiency.

3.3. Microcosm study for Cr(VI) bioremoval from pre-treated tanning wastewater The actinomycete strain Kitasatosporia sp. was used in the microcosm studies for Cr(VI) bioremoval from pre-treated tannery wastewater containing 364 mg/L of Cr(VI). Impact of parameters such as type of porous medium, inoculum size, flow rate and culture conditions on Cr(VI) bioremoval efficiency was studied.

3.3.1.

Type of porous medium

Two different porous medium (sand and gravel) have been used in the microcosms. Sand columns showed high Cr(VI) bioremoval with time (Fig. 2b). At zero time, 30% of Cr(VI) was removed in the inoculated columns compared to only

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Table 1 – Chemical, physical and biological characterization of tannery wastewater.

Color COD (mg/L) BOD (mg/L) TSS (mg/L) Salinity (mg/L) EC (S/cm) pH Sulfate (mg/L) Cr(III) (mg/L) Cr(VI) (mg/L) Fungi (cfu/mL) Bacteria (cfu/mL) Actinomycetes (cfu/mL)

February

May

August

December

Mean value

Maximum limita

Dark blue 4538 1.12 40 35 55.6 4 265 2350 900 105 160 35

Dark blue 3920 2.2 37 29 47.8 3.5 235 2141 814 100 153 32

Dark blue 3538 2 35 26 41 3.1 198 1625 625 98 146 29

Dark blue 4765 1.5 42 37 57 3.8 301 2410 945 115 162 41

Dark blue 4190 ± 20 1.7 ± 0.03 38 ± 2.1 31 ± 2.7 50 ± 4.4 3.6 ± 0.07 249 ± 3.9 2131 ± 8.7 821 ± 5.8 418 ± 2.3 621 ± 5.3 137 ± 2.1

Free of color 100 60 60 NA NA 6–9 1 1 0.5 NA NA NA

NA: not applicable. a

Maximum limit of criteria, Annex 1 of the Egyptian Law 4/94: waste limits.

Fig. 1 – Cr(III) precipitation from tannery wastewater by (A) cement dust and (B) lime. 3% removal of Cr(VI) in the control columns. During the experiment, cycles of Cr(VI) adsorption and desorption were observed in both inoculated and control sand columns. After 150 min, the inoculated sand columns achieved 100% removal of Cr(VI) compared to 60% removal of Cr(VI) in the control columns. On the other hand, the bioremoval of Cr(VI) using gravel as a porous medium was relatively low (Fig. 2a). After 150 min, the inoculated gravel columns reached 60% removal of Cr(VI) compared to 49% Cr(VI) removal in the control columns.

sity equivalent to OD600 = 0.406, Cr(VI) removal reached 87% after 150 min. However, at OD600 = 0.812, 1.21, 1.62 and 2.03, the chromium removal reached 100% after 150, 120, 90 and 30 min, respectively. At OD600 = 2.43, Cr(VI) was removed completely at zero time. The results showed significant difference (P-value = 0.0241) between the different inoculum sizes.

3.3.3. 3.3.2.

Inoculum size

Inoculum size of the microbial culture is one of the most important factors affecting bioremediation experiments. Fig. 3 shows that the increasing of inoculum size was positively correlated to Cr(VI) removal efficiency (R = 0.909). At cell den-

Flow rate

Three flow rates (1 mL/min, 2 mL/min and 3 mL/min) of the pre-treated tannery wastewater were used. It was found that higher flow rate led to lower Cr(VI) removal efficiency (Fig. 4). However, the statistical analysis indicated no significant difference between Cr(VI) removal at the investigated flow rates.

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Fig. 2 – Effect of porous medium type (A) gravel and (B) sand on Cr(VI) bioremoval efficiency.

Fig. 3 – Effect of inoculum size of Kitasatosporia sp. on Cr(VI) bioremoval efficiency in sand columns.

3.3.4.

Culture condition

Two forms of culture conditions were used in this study, induced culture (which was grown in medium containing Cr) and non-induced culture (which was grown in normal medium without Cr). It was found that induced culture was more efficient in Cr(VI) removal than non-induced culture (Fig. 5). Cr(VI) was removed completely after 60 min by using induced culture and after 90 min by using non-induced culture. There was a high positive correlation between the culture condition and Cr(VI) removal (R = 0.942), in addition to a significant difference (P-value = 0.004) between the two different culture conditions.

3.4. Efficiency of combined chemical–biological technique in tannery wastewater treatment The final design of the combined chemical–biological treatment system was performed according to the parameters that gave the maximum Cr(III) precipitation and Cr(VI) bioremoval. Table 2 shows that the combination between chemical precipitation of Cr(III) and biological removal of Cr(VI) from tanning wastewater removed 99.3% of total Cr, 98.4% of Cr(VI), 77% of COD and 81% of turbidity. Thus, the combined chemical–biological treatment system improved the tanning wastewater to meet the limits of environmental regulations.

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Fig. 4 – Effect of wastewater flow rate on Cr(VI) bioremoval efficiency in sand columns.

Fig. 5 – Effect of culture conditions of Kitasatosporia sp. on Cr(VI) bioremoval efficiency in sand columns. Table 2 – Efficiency of the combined chemical–biological treatment of tannery wastewater.

pH Color COD Turbidity (NTU) Cr(VI) (mg/L) Total Cr (mg/L) a

Raw tanning wastewater

After the combined treatment

Treatment efficiency (%)

a

3.1 ± 0.03 Dark green 3538 ± 13.4 83.5 ± 4.5 625 ± 7.8 2250 ± 9.2

6.8–7 ± 0.1 Free of coloring materials 813 ± 6.9 16 ± 1.5 10 ± 0.5 15 ± 1.2

– – 77% 81% 98.4% 99.3%

6–9 Free of coloring materials 100 50 0.5 1

Maximum limit

Max limit of criteria, Annex 1 of the Egyptian Law 4/94: waste limits.

3.5. Recycling of recovered Cr(III) in the tanning process The recycling of Cr(III) recovered from the chemical precipitation stage in tanning process was investigated. Three tanning experiments were carried out by using experimental Cr, commercial Cr and mixed Cr. Table 3 shows the physical and organoleptic of the leathers produced from the three tanning experiments. The leathers produced by the experimental Cr met the stipulated norms for the physical properties. The shrinkage temperature of the leather produced from experimental Cr was 102 ◦ C, while for both the commercial Cr and mixed Cr were 120 ◦ C and 111 ◦ C, respectively. The Cr content of the leather tanned by commercial Cr was slightly higher (3550 mg/L) than the other leathers. The general appearance of the leathers produced from the three tanning experiments was relatively similar. The characteristics of the leathers such

as tear strength, permeability test, leather flex resistance and thickness were not altered by using the experimental Cr in the tanning process (Table 3).

4.

Discussion

Due to the toxic impact of Cr on environment, it is recommended to develop new treatment techniques in order to remediate Cr from wastewater and reuse it back in industry. Here we developed an approach based on combined chemical–biological treatment for tannery wastewater to recover Cr(III) chemically and reuse it in tanning process, and to remove Cr(VI) biologically. Chemical precipitation is the most widely used method for heavy metal removal from industrial wastewater (Barakat, 2011). In the present study, the chemical precipitation of Cr(III) was conducted using two precipitating agents, lime and cement dust. However, our findings showed that the suitable

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Table 3 – Characteristics of leathers tanned by experimental and commercial Cr(III). Parameters

Color Shrinkage temperature Chromium content (mg/L) Tear Strength (km/mm) Tensile strength and Leather flex resistance (%) of elongation Thickness (mm) Permeability test

Commercial Cr(III)

Experimental Cr(III)

Mixed Cr(III)

Bluish white 120 ◦ C 3550

Greenish white 102 ◦ C 3150

Bluish white 111 ◦ C 3370

92 No damage or cracking 1.9 Good permeability during 8 h

85 No damage or cracking 1.2 Good permeability during 8 h

90.1 No damage or cracking 1.5 Good permeability during 8 h

The no. of standard used – E.S 3571/2006 E.S 3571/2006 ISO 5402&5403 E.S 3571/2006 E.S 3571/2006 ISO 5402&5403 E.S 3571/2006 E.S 3571/2006

Limit of standard

Bluish white >100 ◦ C 3550 70 km/mm No cracking after 20.000 cycles 2 Good permeability during 8 h

E.S: Egyptian standard.

precipitating agent was lime. The maximum Cr(III) removal efficiency (98–99%) was observed by using lime, while cement dust had a much lower removal efficiency ranged between 1.5 and 2.6%. Possible explanation could be that the presence of lime maintains a conductive pH for the formation of Cr(OH)3 precipitate (Sharma and Goyal, 2009). By increasing the concentration of lime to 2–3 g/100 mL and settling time to 120–180 min, the pH values reached 7.7–8.2 (Abdulla et al., 2010), which was close to the optimum value (pH 8) for Cr(III) precipitation as suggested by Patterson and Minear (1975). On the other hand, cement dust increased slightly the pH value of the wastewater to 3.5 (Abdulla et al., 2010). The tannery wastewater tends to be acidic and it is known that there is a competition between protons and metal ions under acid conditions (Sutherland et al., 2002). Thus, lime addition was necessary for turning the pH of tannery wastewater from acidic to alkaline in order to achieve maximum Cr(III) removal (Bailey and Tunick, 1982; Cetin et al., 2013). Previous studies have also reported that lime was one of the most efficient precipitant agents due to their low-cost in most countries, simplicity of the process and safe operations (Mirbagherp and Hosseini, 2004; Aziz et al., 2008). Chemical precipitation using lime could also improve the pH of tannery wastewater to meet the standard discharge requirements (pH 6–9). Biological remediation of Cr(VI) from industrial wastewater has recently received much attention (e.g., Qian et al., 2016). Biotransformation (reduction of toxic, mobile Cr(VI) to less toxic, less mobile Cr(III)) and biosorption (accumulation of Cr(VI) on biomass) are the most common mechanisms used by microorganisms to remediate Cr(VI) (Kanmani et al., 2012; Wang, 2000). Most of the studies on microbial Cr(VI) removal have been conducted in batch experiments (Chen and Hao, 1998; Mahbub, 2004; Qian et al., 2016) and these studies often cannot be directly applied to bioremoval of Cr(VI) in situ. However, Cr(VI) transport through saturated porous media in microcosm columns is a highly dynamic process that cannot be fully defined through batch reactor experiments (Molokwane and Chirwa, 2013). In the present study, to adjust the microcosm experimental parameters for Cr(VI) bioremoval, factors such as types of porous medium, inoculum size, flow rate and culture conditions were studied. The type of porous medium determines the working volume of the wastewater to be treated and the surface area in which microorganisms can be attached (Shen and Wang, 1995). The present investigation showed that sand had higher Cr(VI) removal efficiency than gravel. Several explanations could be possible. Firstly, the saturation system of the gravel columns led to anaerobic conditions that inhibit the biore-

moval ability of the actinomycete strain. This observation was in agreement with the finding of Dermou and Vayenasa (2007) that the Cr(VI) bioremoval by microorganisms in microcosms using gravel as a supporting medium had a low efficiency for industrial wastewater treatment. Secondly, the surface area is another parameter affected drastically by the type of porous medium. Gravel offers lower surface area for microbial adhesion than sand (Nkhalambayausi-Chirwa and Wang, 2004; Verhagen et al., 2011). Thirdly, the macro-pores between the gravel in the column could reduce the contact time between microbial culture and gravel during the inoculation, thus the microbial attachment efficiency will be extremely low and will affect negatively the Cr(VI) bioremoval (Unc and Goss, 2003). Higher inoculum size of the actinomycete strain Kitasatosporia sp. was positively correlated to higher Cr(VI) removal from tannery wastewater. Previous studies found that the inoculum size had a significant effect on Cr(VI) removal efficiency by various bacterial and fungal isolates in batch and microcosm studies (e.g., Salunkhe et al., 1998; Rahman et al., 2000). That could be explained by that higher inoculum size led to higher biofilm formation in the treatment system, as a consequence higher metabolic activities could be contributed in the Cr(VI) reduction (Paul et al., 2012). That was in agreement with earlier studies showed that the maximum rate of Cr(VI) reduction was achieved at high inoculum size of 9.6 × 107 cells/mL with compared to a low inoculum size of 2.4 × 107 cells/mL (Mohammad and Shahida, 2003; Ezaka, 2012). Sepehr et al. (2005) have also reported that high inoculum size could affect the hydrodynamic characteristics of the medium and as a consequence the Cr(VI) removal rate could increase. Moreover, some studies found that increasing the inoculum size led to an increase in metal biosorption sites since the amount of added biosorbent determined the number of binding sites available for Cr(VI) biosorption and that would increase the removal of Cr(VI) from solution (Cervantes et al., 2001; Cárdenas and Acosta, 2011). The other two factors (flow rate and culture conditions) had a lower effect on Cr(VI) removal efficiency than the type of porous medium and inoculum size. For instance, using different flow rates of the wastewater affect slightly the Cr(VI) removal, in which higher flow rate led to lower Cr(VI) removal efficiency. Possible explanation is that low flow rate increase the contact time between the microbial culture and Cr(VI), thus increase the bioremoval efficiency. Baig et al. (2003) had a similar conclusion that the bioremoval efficiency of Cr(VI) in sand column was effective when the flow rate of wastewater was low. On the other hand, for the culture conditions, we found that induced culture was relatively more efficient in

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Cr(VI) removal than non-induced culture. Our findings agreed with previous studies that suggested that the microbial cultures used in bioremediation studies need to be cultivated in Cr(VI) containing medium to enhance the expression of the genes responsible for biosynthesis of Cr(VI) reductase enzymes (e.g., Focardi et al., 2013). On the other hand, we have noticed a serial of adsorption and desorption of Cr(VI) in both inoculated and control columns by time. That may be due to the negatively charged surfaces of both sand particles and microbial cell wall tend to attract Cr(VI), although a weak electrostatic bond was formed and led to Cr(VI) desorption by time (Klein and Hurlbut, 1993; Alam et al., 2014). The present study showed that the combined chemical–biological treatment not only removed 99.3% of total Cr and 98.4% of Cr(VI), but also recovered 77% of COD and 81% of turbidity. Sharma and Malaviya (2014) have reported similar finding is that using combined chemical (precipitation) and biological (by Fusarium chlamydosporium) treatment method reduced 71.80% of COD, 64.69% of turbidity and 62.33% of total Cr. Moreover, Goswami and Mazumder (2014) showed that the combination between physico-chemical and biological methods was very efficient in treating tannery wastewater by reducing Cr, other heavy metals, COD and turbidity. The recycling of precipitated Cr(OH)3 in the tanning process was very successful and almost gave the same results as the commercial Cr. There are two main ways of Cr(III) recycling (Belay, 2010): (a) direct recycling of the Cr containing effluent back into Cr tanning processing, (b) indirect recycling of precipitated Cr(III) from Cr containing effluent, and then redissolving it in acid for reuse. However, it has been suggested that the indirect recycling of Cr is more efficient in leather tanning and environmental friendly (Awan et al., 2003; Cetin et al., 2013). One important factor in Cr recycling in tanning industry is adjusting the basicity to avoid the precipitation of Cr during the tanning process (Buba, 2004; Gutiérrez-Gutiérrez et al., 2013). In the present study, the recovered Cr(III) was re-dissolved in sulfuric acid (pH 2.7–3.0) to be reused in the leather tanning. That was in line with a recent study showed that the optimum parameters for reusing Cr in the tanning industry were re-dissolving it in H2 SO4 (pH 2.5) (Yassen et al., 2015). Moreover, Ibrahim (2003) found that dissolving the recovered Cr in H2 SO4 (pH 2.5–3.0) was not only efficient in avoiding Cr precipitation but also had a remarkable impact on the leather tanning since those conditions increased the replacement of the OH groups by Cr(III) compounds during the tanning process and thus increased the quality of the tanned leather. Exhaustion of Cr in the recycled stream could mainly depend on the number of tanning cycles. In each cycle, 60% of Cr reacts with the hides and the rest amount (40% of Cr) remains in the liquid wastes (Sreeram and Ramasami, 2003). According to the extent of Cr reuse, Cr utilisation would be increased from 70% up to 95% and Cr discharge in the stream would be decreased (Ludvík, 2000). Thus, reusing the Cr mixture (50% of the recovered Cr mixed with 50% of commercial Cr sulfate) in the tanning process could be an efficient choice since it would increase the lifetime of the recycling process (Fathima et al., 2012).

5.

Conclusion

A combined chemical–biological treatment system was carried out for Cr remediation and recycling from tannery

wastewater. The present study demonstrated that the chemical precipitation of Cr(III) was extremely efficient by using 2 g/100 mL of lime and 2 h of settling rate. Furthermore, the maximum removal of Cr(VI) was achieved by using sand as a porous medium in the microcosm columns that were inoculated with induced culture of Kitasatosporia sp. (OD600 = 2.43) and the flow rate of wastewater was 2 mL/min. The present combined chemical–biological treatment method improved the characteristics of tanning wastewater by removing 99.3% of total Cr, 98.4% of Cr(VI), 77% of COD and 81% of turbidity. Moreover, the precipitated Cr(III) was successfully recycled in the tanning process and gave a comparable quality of leather tanning as the commercial Cr(III). Thus the present study succeeded in designing an environmentally safe and costeffective treatment system for tannery wastewater which was efficient enough to meet the standards of industrial wastewater disposal.

Acknowledgments We would like to thank Dr. Sahar El-Shatoury (Suez Canal University, Egypt) for her valuable suggestions. The present study was supported by grants from the Faculty of Science, Stockholm University, Sweden.

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