sustainability Article Immobilization of Metanil Yellow Decolorizing Mixed Culture FN3 Using Gelling Gum as Matrix for Bioremediation Application Fatin Natasha Amira Muliadi 1 , Mohd Izuan Effendi Halmi 1, *, Samsuri Bin Abdul Wahid 1 , Siti Salwa Abd Gani 2 , Khairil Mahmud 3 and Mohd Yunus Abd Shukor 4   Citation: Muliadi, F.N.A.; Halmi, M.I.E.; Wahid, S.B.A.; Gani, S.S.A.; Mahmud, K.; Shukor, M.Y.A. . Sustainability 2021, 13, 36. https://dx.doi.org/10.3390 /su13010036 Received: 30 September 2020 Accepted: 18 November 2020 Published: 22 December 2020 Publisher’s Note: MDPI stays neu- tral with regard to jurisdictional claims in published maps and institutional affiliations. Copyright: © 2020 by the authors. Li- censee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/ licenses/by/4.0/). 1 Department of Land Management, Faculty of Agriculture, Universiti Putra Malaysia, Serdang 43400, Malaysia; fatin.muliadi@gmail.com (F.N.A.M.); samsuriaw@upm.edu.my (S.B.A.W.) 2 Department of Agricultural Technology, Faculty of Agriculture, Universiti Putra Malaysia, Serdang 43400, Malaysia; ssalwaag@upm.edu.my 3 Department of Crop Science, Faculty of Agriculture, Universiti Putra Malaysia, Serdang 43400, Malaysia; khairilmahmud@upm.edu.my 4 Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang 43400, Malaysia; mohdyunus@upm.edu.my * Correspondence: m_izuaneffendi@upm.edu.my; Tel.: +603-9769-4958 Abstract: In this study, the Metanil Yellow (MY) decolorizing mixed culture, namely FN3, has been isolated from agriculture soil. The mixed culture was immobilized using gellan gum. In order to optimize the immobilization process for maximal dye decolorization, Response Surface Methodology (RSM) was performed. The optimal conditions for immobilization predicted by desirability function are 130 mg/L of MY dye concentration, 1.478% of gellan gum concentration, 50 beads and 0.6 cm of beads size with the percentage of decolorization of 90.378%. The correlation coefficients of the model (R 2 and R 2 adj) are 0.9767 and 0.9533, respectively. This indicates that the established model is suitable to predict the effectiveness of dye decolorization under the investigated condition. The immobilized beads of mixed culture FN3 were able to be reused up to 15 batches of decolorization. The immobilized cells also have high tolerance towards heavy metals. This was proven by higher dye decolorization rate by the immobilized cells even with the addition of heavy metals in the media. The decolorization potential of the mixed culture indicates that it could be useful for future bioremediation of soil contaminated sites and treatment solutions of water bodies polluted with MY dye. Keywords: immobilization; mixed culture; Metanil Yellow; response surface methodology 1. Introduction Azo dyes are extensively used in various industries such as textile industry, food, paper-making and cosmetic industries [1,2]. With the growing demand of the textile industry, approximately 40,000 different dyes and pigments are being use and 2000 azo dyes are currently in use. The production of azo dyes annually worldwide is around 7 × 10 5 tonnes [3]. Azo dyes are highly water-soluble. They include one or more azo (-N=N-) groups and sulfonic (SO 3− ) groups [4]. Due to the dyeing process, the textile industry releases an enormous amount of wastewater into the environment. Typically, about 10–15% of the dye is lost in effluents, since they do not bind to the fibres [5]. This has led to water pollution. Metanil Yellow (MY) (3-(4-Anilinophenylazo) benzene sulfonic acid sodium salt is a type of acidic azo dyes [6]. MY is commonly used as soap colouring, spirit lacquer, shoe pol- ish, bloom sheep dip, for preparation of food stains, leather dyeing, for manufacturing of pigment lakes and paper staining [7]. MY is categorized as nonpermitted food colourants. It is widely used as alimentary dye adulterant in India [8]. However, it was found out Sustainability 2021, 13, 36. https://dx.doi.org/10.3390/su13010036 https://www.mdpi.com/journal/sustainability