*Article* **In Vitro Bioadsorption of Cd2**<sup>+</sup> **Ions: Adsorption Isotherms, Mechanism, and an Insight to Mycoremediation**

**Raman Kumar 1,\*, Priyanka Sharma <sup>2</sup> , Ahmad Umar 3,4,\*, Rajeev Kumar <sup>2</sup> , Namita Singh <sup>5</sup> , P. K. Joshi 6,7, Fahad A. Alharthi <sup>8</sup> , Abdulaziz Ali Alghamdi <sup>8</sup> and Nabil Al-Zaqri <sup>8</sup>**


Received: 1 August 2020; Accepted: 29 August 2020; Published: 2 September 2020

**Abstract:** The objective of this paper is to establish the significance of the mycoremediation of contaminants such as Cd2<sup>+</sup> to achieve sustainable and eco-friendly remediation methods. Industries such as electroplating, paint, leather tanning, etc. release an enormous amount of Cd2<sup>+</sup> in wastewater, which can drastically affect our flora and fauna. Herein, we report on the in vitro bioadsorption of Cd2<sup>+</sup> ions using fungal isolates obtained from different contaminated industrial sites. The detailed studies revealed that two fungal species, i.e., *Trichoderma fasciculatum* and *Trichoderma longibrachiatum*, were found to be most effective against the removal of Cd2<sup>+</sup> when screened for Cd2<sup>+</sup> tolerance on potato dextrose agar (PDA) in different concentrations. Detailed adsorption studies were conducted by exploring various experimental factors such as incubation time, temperature, pH, inoculum size, and Cd2<sup>+</sup> salt concentrations. Based on optimum experimental conditions, *T. fasciculatum* exhibited approximately 67.10% removal, while *T. longibrachiatum* shows 76.25% removal of Cd2<sup>+</sup> ions at pH 5.0, 120 h incubation time, at 30◦C. The inoculum sizes for *T. fasciculatum* and *T. longibrachiatum* were 2.5% and 2.0%, respectively. Finally, the morphological changes due to Cd2<sup>+</sup> accumulation were examined using scanning electron microscopy (SEM). Further, Fourier transform infrared spectroscopy (FTIR) spectroscopy reveals the presence of various functional groups (-CH, –C=O, NH and –OH), which seem to be responsible for the efficient binding of Cd2<sup>+</sup> ions over the fungal surfaces.

**Keywords:** *T. longibrachiatum*; *T. fasciculatum*; bioadsorption; cadmium; heavy metals; isotherms; bioadsorption mechanism; mycoremediation

#### **1. Introduction**

The emerging worldwide environmental problems of the past decades are mostly related to the superfluous increase in metallic contaminants in the environment [1–6]. Inorganic contaminants such as heavy metals released into the environment through various industrial, agricultural, and domestic practices [5,6]. Heavy metals may also enter into the environment by natural sources, but the extent of such exposures is non-significant. Heavy metals are often present together with organic pollutants in industrial wastewater [7]. The industrial effluents are rich in dissolved salts of Cr, Cu, Ni, Zn, and Cd, etc., and can cause a serious problem to the surrounding environment. Heavy metals pollution causes serious problems due to their non-biodegradable nature [8,9]. Although these metals are vital for the proper functioning of biological systems but only in trace concentrations, higher concentrations may lead to disturbed functioning of bio-geo-chemical cycles [10–12]. In addition, serious health issues may arise if such metal ions can enter into the food chains and their corresponding products. Thus, even 1.0–10 mg/L can cause health problems in human beings such as jaundice, facial edema, blue lungs, kidney damage, hearing disorder, skin cancer, asthma, protein metabolism, and bronchial cancer, etc. [13,14]. Therefore, a number of traditional physiochemical treatment techniques have been reported to remove the heavy metals from wastewater, especially from the industrial wastewater, to name a few, electro-coagulation, solvent extraction, ion-exchange, electro-reduction, reverse osmosis, adsorption, membrane separation, chemical precipitation, and so on [15–29]. Even though used widely, these above-mentioned techniques exhibited several disadvantages which include the utilization of high-cost equipment and monitoring systems, the use of various expensive chemicals, the discharge of toxic sludge, long processing time, the production of toxic waste products that need further processing, and so on [23–31]. Thus, it is the necessity of the time to develop a new, simple, and inexpensive method for heavy metal removal from wastewater. Recently, by utilizing living/non-living microorganisms and their derivatives to remove the heavy metals from industrial wastewater is considered a promising technique [32–34]. Accordingly, the bioremediation can be considered as one of the most important and advantageous process that can efficiently remove the heavy metals from industrial wastewater. The entrapment of the heavy metal ions and their corresponding sorption on the binding cites of the cellular structures leads to the bioadsorption of heavy metal ions from the industrial wastewater [35,36]. As a result of the faster adsorption process, the microbial cells are considered as an important and advantageous scaffold for the biosorption of heavy metals and wastewater treatment.

In this study, different fungal isolates extracted from various contaminated sites (Haryana, India) were cultured and screened for tolerance to Cadmium (Cd2+). Various process factors, i.e., inoculum size, pH, initial metal ion concentration, temperature, and incubation time on the Cd2<sup>+</sup> removal by highly efficient Cd2<sup>+</sup> tolerant fungal isolates have been investigated and presented in this article. In addition, mechanisms of Cd2<sup>+</sup> removal by efficient Cd2<sup>+</sup> tolerant fungal isolates have also been studied.

#### **2. Materials and Methods**

#### *2.1. Chemicals Used*

Potato dextrose broth (PDB) and nutrient media, potato dextrose agar (PDA), were procured from Hi-Media, Mumbai India. Cd(NO3)2, was provided by Fine-Chem Limited, Mumbai, India. The solutions were prepared in triply distilled water (conductivity =0.5 µS cm−<sup>1</sup> ), and the reagents were used without any further purification.

#### *2.2. Sample Collection and Isolation of Fungi*

Samples from various sites such as industrial effluents, sewerage discharge, and sludge were collected from Karnal, Panipat, and Sonepat districts of Haryana (India). The samples were stored at 4 ◦C prior to further processing. Fungal isolates were isolated from samples by the PDA serial dilution method. The serial dilutions were made up to 10<sup>6</sup> and 1 mL of each dilution i.e., 10<sup>4</sup> and 10<sup>6</sup> were poured over PDA plates. An incubation of cultured petri plates was performed at 28 ◦C for 96 h. The prominently grown fungal colonies were regenerated and purified using the streak plate method.
