Biosorption of textile dye effluents using marine red macroalgae Gracilaria corticata as a low-cost adsorbent and its potential was evaluated. The process parameters responsible for adsorption of dye from the aqueous solution, such as pH (4-8), temperature (30-40°C), initial dye concentration (10-20%), biomass concentrations (1-10 giL}, agitation/static and time (120-180 min) ware optimized using the statistical approach. Plackett-Burman design of josorption. The resultant decolourization of the effluent before and after biosorption were compared using UV spectrophotometer. The predicted values and the experimental values were found to be in good agreement (R*= 0.9794 and Adj- R? = 0.9547). Four out of six operational parameters, namely biomass concentration, pH, dye concentration and static-agitation (P<0.05) showed a very good sign in the decolourization process. KEYWORDS, Biosorption, Gracilaria corticata, Statistical approach, Plackett-Burman, decolourization, UV spectrophotom: eter 1. INTRODUCTION Chemical dyes are hazardous to all living beings and are released into the environment without any treat- ment by textile, leather, chemical, food and pharma- ceutical industries. Textile effluents are often a threat, to humanity in a densely populated urban city. Efflu- ents from the textile industries include various inor- ganic and organic chemicals of various quantities |1). ‘The textile industry is characterized into three differ- ent types, namely cellulose fibres, protein fibres and synthetic fibres. Direct dyes, such as direct brown 116, reactive yellow and congo red, reactive dyes, such as cibacron F, procion M, pricion H, livafix E and remazol, indigo dyes, such as indigo white and tyrian purple, and naphthol dyes, such as fast blue B and fast scar- let are some of the dyes used in the cellulose fibres dyeing process. Acid dyes, such as azo dyes, anthraquinone dyes, riaryl methane dyes, triphenyl methane and lanaset dyes, like Bordeaux B are used to dye protein fibres [2]. Synthetic dyes are dyed using direct dyes, dispersive dyes, like disperse navy 35, disperse yellow 218, basic dyes, such as basic red and basic orange 37 [3]. Textile industries also add chemicals, like sulphates, peroxides in order to fix the dyes onto the clothes. Textile effluents also contain heavy metal contaminants, such as arsenic, lead, zinc, chromium, copper and mercury [3]. These chemical dye effluents are released into the water bodies and lands. which accounts for the cause of rising environmental, pollution. The release of chemical dyes will increase the levels, of chemical oxygen demand (COD), biological oxygen demand (BOD) and total dissolved solids (TDS) in the water body. The presence of the dyes and heavy met- als released into the water bodies causes immense threat, such as toxicity, carcinogenicity to both aquatic, and human lives. The textile effluents are prone to cause ‘mutation in genes and inhibit DNA replication and pro- tein synthesis [4,5]. The presence of chemicals in water can cause nausea, skin problems, eye irritation, haemorrhage. The textile effluents undergo three types, of treatments, namely primary, secondary and tertiary treatment which includes filtration, sedimentation, co- agulation, flocculation, ozonation, electrocoagulation. Often textile dyes are difficult to treat since they have the potential resistance towards microorganisms, sta- bility against heat, light, stability against oxidizing agents [6]. Several low-cost technologies using bioremediation, phytoremediation, mycoremediation and, biosorption have come upto decolourize the dye in a more efficient and cost-effective manner. Adsorption of dyes on activated carbon is developing nowadays because of its high etficiency and low investment. Marine macro-algae are a very cheap source and have a great potential in adsorption of dyes due to its sur- face charge binding efficiency and large surface area Green algae (Cladophora fascicularis) have the poten- m2 INDIAN J. ENVIRONMENTAL PROTECTION, VOL. 39, NO. 8, AUGUST 2019 © 2019 - Kalpana Corporationtial to adsorb heavy metals, like lead [71. This research, work focuses on the use of mi 3d algae species (Gracilaria corticata) as effective biomass for adsorp- tion of dyes present in the effluent. The presence of, charged functional groups, such as sulphoxide, phos: phate, phenol and alcohol on the surface of Gracilaria corticata make them more suitable for the adsorption, process. 2. MATERIAL AND METHOD All the chemical reagents used for the experiment was of high grade and high purity. 0.11N HCI and 0.1M NaOH were prepared in distilled water to adjust the pH of the solution. 2.1 Collection of red macroalgae Gracilaria corticata ‘The Gracilaria corticata was harvested from the inter- tidal zone of Mandapam coast (latitude: 9°16'32.56"N and longitude: 79°7'25.03"E), Ramanathapuram dis- trict, Tamil Nadu. The sample was collected manually using hands. The harvested seaweed was washed with sea water initially to get rid of the sand and other epi- phytes and was collected in a polyethylene bag which was taken to the lab and washed with tap water fol- lowed by distilled water (Figure 1). 2.2 Species identification The seaweeds collected were authenticated by Dr. M. Ganesan, Scientist, CSIR-Central Salt and Marine Chemicals Research Institute (CSMCRI), Mandapam Camp, Ramanathapuram district, Tamil Nadu. 2.3 Preparation of adsorbent The red algae were shade dried at room temperature for a period of four days (Figures 2 and 3). The dried algae were crushed and ground coarsely using an elec- trical blender. Then the blended sample was stored in an air-tight polyethylene bags and was stored at room ‘temperature ina dry place. These blended algae wer used as an adsorbent. The biosorbent was sieved manu- ally to remove fine particles. Adsorbents with particle size ranging from 1000 to 1200 um were utilized (Fi INDIAN J. ENVIRONMENTAL PROTECTION, VOL. 39, NO. 8, AUGUST 2019 713 © 2019 - Kalpana CorporationTable 1. Experimental variables with thelr respective factor lavel ‘Sd order] Run ordor [Pr type [Blocks [pH |Temperature | Dye conc. [Biomass [Stalls or | Time (min) i) 1%) cone. (git) | agitation 1 1 1 1 a _[20 20 1 State 120 2 1 ht 2 [ao 10 10 Static 120 a 1 ih fa [ao 20 1 ‘Agitation [120 4 1 1 2 (so 20 10 Static 180 5 1 1 e [ao 10 10 ‘Agitation [120 6 8 1 1 e (40 20 Agitation [180 7 7 1 1 4 _[40 20 10 ‘Static 180 @ 8 1 hfe [so 20 10 ‘agitation [120 ® a 1 1 4 [20 10 10 Agration [180 10 10 1 hfe [20 10. 1 Agitation [180 in 1 1 ih fa [ao 10 1 Static 180 12 2 1 h 4 [so 10 1 Static 120 ure 4), 2.4 Pretreatment of the biosorbent In order for efficient biosorption, the surface of the biosorbent must be activated. The pretreatment was carried using physical and chemical methods. Physical treatment using boiling water was preferred. The dried biomass was added to 500 ml of 150°C boiling water and stirred for 15 min in a glass bowl. The biomass was then filtered using Whatman filter paper (size no. 40) and a funnel and dried in a hot air oven at 60°C for, 24 hr (8) 2.4.1 Chemical method (a) Calcium chloride treatment : 50 g of dried biomass was taken in a 1 L standard Erlenmeyer flask and was treated with 500 mL of 0.2 M CaCl, solution and the pH was set to 5. The mixture was kept in an orbital shaker at 150 rpm at 25°C for 24 hr. This was done to ensure complete cross-linking and the ion exchange of alginate present in the algae. Once the equilibrium time was achieved, the mixture was filtered using Whatman filter paper and a glass funnel. The filtrate was discarded and the biomass was washed with dis- tilled water in order to remove the excess calcium chlo- ride and dried in hot air oven at 60°C for 24 hr [9, 101. (b) H.CO, treatment: 50 g of biomass was treated 170 ml of 36% formaldehyde and 330 mL of 0.1N HCl in a 1 L standard Erlenmeyer flask and shaken in an orbital shaker for 1 hr and complete cross-linking of alginate materials and polymerization was achieved. The mix- ture was then filtered using a funnel and Whatman fil m4 ter paper, then the biomass was washed with distilled water to remove the traces of formaldehyde and HCl and washed with 0.2 MNa,CO, and again with dis- tilled water. The biomass was then dried overnight at, 60°C and at 110°C for 2 hr in hot air oven [11] (Figure 5). 2.5 Collection of textile dye effluent The textile dye effluent was collected from the local area in Murugampalayam, Tiruppur, Tamil Nadu. The dye was collected in a clean 1 L plastic bottle. The dye was collected from the sewage treatment plant of the dustry. The dye was dark in colour with foul smell due to several chemicals. The spectrum for the crude effluent dye was obtained using Shimadzu UV-1800, UV-visible spectrophotometer (Tokyo, Japan). The 2 gax WAS found to be at 578.8 nm with an absorbance vale of 0.7. The textile effluent was diluted in the ratio of 10% (1:10) to 20% (2:10) with sterile distilled water and was used for the batch adsorption studies, respectively. 2.6 Batch adsorption studies using Plackett-Burman dk ‘The batch adsorption experiments were performed in 250 mL standard Erlenmeyer flasks with 100 mL of the diluted dye solution. The pH (4-8), temperature (30 40°C), dye concentration (10 -20%}, biomass con. centration (1-10 gil), statics/agitation, time (120-180 min) were preferred for batch adsorption studies. The adsorption studies were performed using the Minitab18 trial version software. The significance of Plackett- INDIAN J. ENVIRONMENTAL PROTECTION, VOL. 39, NO. 8, AUGUST 2019 © 2019- Kalpana CorporationTable 2. Experimental value and predicted value for 12 tals in Plackett-Burman methods Run] ak pH Tome Dye cone. [Biomass | Static or | Time [exp (m) | Pred) cane agitation 1 Hi 20.01 90.28 2 1 + + + 26.72 27.08 3 1 i A ena 27.69 4 1 + + + + 28.20 27.94 5 1 A i Hi 86.13, 86.70 8 1 + + + : + + 29.14 29.66 7 1 i A 29.71 09.69 a 1 : + + + 9.67 29.63 2 1 * i 7.21 67.60 1 [a + : : + + 8.07 28.21 mn + loo.as. 20.20 2 : : : : : : lo0.98 20.47 Table 3. Regression stetstics and ANOVA table for the Plackett Burman desion Term Effect Coctfcient | SE coefficient | T value P value VF ‘Constant 28.980 0.102, 268.05, (0.000) pH 1.747 0.873 0.102 8.53 0.000" 1.00 Temperacure 0.110 0.055) 0.102, 0.54 0.614 1.00) bye 1.450 0.728 0.102 7.08 0.001" 1.00 Biomass 2.047 1.028 0.102, 9.99) (0.000° 1.00) StaticiAgitation 0.785 0.377, 0.102 3.68 0.014 1.00 Time 0.243) 0.122, 0.102, 119 0.288, 1.00) + The significant velues (P-<0.08) Burman design was to determine the essential signifi cant parameters among a large number of parameters. that affect the process of biosorption of textile efflu tent onto the biomass [121. In the current study, Plackett- Burman design with 12 trails for 6 different variables ‘was performed. The factor levels were set from lower, limits (-) to the upper limits (++). Table 1 denotes the experimental design with their actual variable levels for the six variables. ‘The experiments were performed in an incubator for static condition and in an orbital shaker for agitation. ‘The speed of agitation was fixed to150 rpm for agita- tion. Once the equilibrium conditions were attained, the contents were centrifuged at 2500 rpm for 5 min to remove the biomass. The pellets were discarded and the supernatant was filtered using Whatman filter pa- per and a funnel and then the filtrates were analyzed in the Shimadzu UV-1800 UV-vis spectrophotometer at 878.8 nm. Control was set up for the same param- INDIAN J. ENVIRONMENTAL PROTECTION, VOL. 39, NO. 8, AUGUST 2019 eters without the algae biosorbent in order to know whether there was an influence of abiotic parameters in the degradation. The blank used in the spectropho- tometer was sterile distilled water. The percentage of decolourization was obtained using the below-mentioned formula 3. RESULT AND DISCUSSION Biosorbent act as a promising remedy for the adsorp- tion of toxic dye. Biosorption is based on the ability of, 2 biomaterial, such as seaweed, activated carbon, plant, bacterial or fungal biomass, to uptake the dyes from the aqueous solution through the metabolically mediated pathway or physico-chemical absorption 113, 14], Biosorption of textile dye effluents has largely relied on equilibrium which is dependent upon various process parameters, such as pH, temperature, dye con: 715 © 2019 - Kalpana CorporationFigure 8. Biomass ready 10 be used as 2 biosorbent timated effects of Independent variables on dye effluent biosorption Figure 6. Estimated effect of independent variebles on textile dye effluents using Gracilerio corticata centration, biomass concentration, static/agitation, time and their interaction with different cationic and. anionic dyes [15,16]. Apart from the process param- ters, the availabilty of the binding site on the surface of the biomass is also an essential factor that pro- vokes the biosorption process, since the cationic and, the anionic dyes bind to the surface of the biosorbent due to their respective complementary charges [17] ‘The pH is one of the most significant factors as it en- sures the availability of the binding site to the sorbate ‘onto the biosorbent. The type of biomass and the con- centration which determine the binding of the dyes onto, the sorbate. The increase in the concentration of the biomass increases the quantity of dye biosorbed per unit weight of biomass (17]. Increase in the dye con. centration may sometime reduce the decolourization and biosorption efficiency. Temperature also has a posi- tive impact on biosorption process, thereby enhancing the kinetic energy of the adsorbate [4]. 3.1 Sereening for significant variables using Plackett- Burman design ‘Six process parameters, namely pH, temperature, dye concentration, biomass concentration, static/agitation and time were analyzed for the adsorption of dyes on biomass using statistical approaches (Plackett-Burman method). Four out of six process parameters (biomass concentration, pH, dye concentration and static/agita- tion) were found to be significant {P<0.05) in the present study. Table 2 shows the levels of predicted and the experimental values of the tested variables and the percentage decolourization due to adsorption ineach run. The variation in the percentage decolouriza- tion was between 86.70-91.59. The highest decolouri- zation was attained in the third run with 91.59% decolourization efficiency. Table 3 indicates the regres- sion statistics and analysis of variance for the dye adsorption using Plackett-Burman design. The coeffi cient values of respective factor denote the effect of that factor on dye adsorption. The coefficient values of pH, temperature, biomass concentration, staticagi- tation and time were found to be -1.747, -0.110, - 2.047, -0.783 and -0,243, The negative coefficient effects conclude that the decrease in the pH, tempera- ture, biomass concentration, static/agitation and time could exert positive effects on the adsorption of dyes onto the biomass. Similarly, the coefficient value of dye concentration is + 1.450. The positive value de notes that increase in the dye concentration could ex- ert positive effects on the dye uptake by the biomass hence contributing to efficient decolourization. Higher the positive or the negative value, greater the rate of, the influence of the respective parameter on the adsorption efficiency of the dye onto the biomass [4] Table 3 denotes the significant factors. The variables with P value less than 0.05, are considered as signifi- cant that affect the biosorption process. Figure 6 de- notes the significant variables affecting the biosorption process. The significant variables are denoted in the Pareto chart (Figure 7). The Plackett-Burman design experimental results were fitted onto the first-order polynomial equation and is denoted as response (Fig ure 8), Response = 91.669-0.4367 pH, - 0.0110 tempera- ture, + 0.1450 dye, - 0.2274 biomass, -0.377 a tion, - 0.00406 time Adsorption of basic dye crystal violet by Gracilaria corticata reported by Vijayaraghavan reported that the maximum biosorption was attained at pH 8, with the adsorbent dosage of 5 g/l. and the initial dye concen: tration of 1000 mg/L [18]. The equilibrium adsor isotherms were fitted onto the Langmuir, Redlich- Peterson isotherms, Toth and Sips and Freundlich mod- els. The results reported that the marine red algae (Graciiaria corticata) could potentially adsorb 193.2 mg/ @ for crystal violet using Langmuir model. This was 716 INDIAN J. ENVIRONMENTAL PROTECTION, VOL. 39, NO. 8, AUGUST 2019 © 2019- Kalpana CorporationFigure 7. Pareto chart for six independent variables Nem rebity Pet vrpone o Figure 8. Normal probabity plot due to the presence of negatively charged functional, groups, like carboxyl, phenol, alcohol and sulphoxide groups [19]. Vijayaraghavan reported the potential of, Gracilaria corticata in biosorption of cationic toxic methylene blue dye. The experiment involved three parameters, namely pH, initial dye concentration and biomass concentration. The optimum adsorption was at the initial dye concentration of 150 mg/L, pH 8 and, the biomass concentration of 6 g/L. Langmuir model, was best fitted with the biosorption potential of, 95.41mgig for methylene blue [20, 21) 4. CONCLUSION ‘The marine red algae Gracilaria corticata was found to be a low-cost alternative for the biosorption of textile dye effiuent. The algae were pretreated in order to enhance the process of biosorption of dyes, this was. achieved using boiling water, calcium chloride, formal- dehyde, HCI and sodium carbonate. The presence of, charged functional groups on the surface of algae had, a significant impact on dye adsorption. The statistical approach using Plackett-Burman design was used to screen the essential significant variables that affect the biosorption process. The four out of six variables, that had a significant impact on the adsorption, namely biomass concentration, pH, dye concentration and static/agitation (P<0,05) which represents strong evi- dence against the null hypothesis. The maximum decolourization of textile dye effluent was 91.59% with R’=0.9794 and Adj-R’ =0.9547, was obtained in the third run of the 12 run Plackett-Burman design. FUTURE WORK The future work will focus on using the obtained sig- nificant parameters for effective biosorption of textile dye effluents onto the Gracilaria corticata using Box- Behnken method in response surface methodology. Using of desorbing chemical such as solvents can be used to desorb the adsorbed dye from the biomass, the biomass can be filtered and the desorbed dyes can be reused by the textile industry. This desorbed biom- ass can be safely used as landfills or as soll amend- ments. ACKNOWLEDGEMENT ‘The authors sincerely thank Vels Institute of Science Technology and Advanced Studies management for their support to carry out the research work. REFERENCES Kromm DE. 1973. Response to air pollution in Ljubljana, Yugoslavia. Ann. Am. Assoc. Geogr, 63(2) : 208.217. 2. Schmidt, A., E, Bach and E, Schollmeyer. 2003. 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