**2. Material and methods**

#### **2.1. Biosorbents used**

*Litchi chinensis Sonn, Citrus limonium, Mammea americana, Tamarindus indica, Citrus sinen‐ sisOsbeck, Citrus reticulate, Cucumis melo* L., and *Musa cavendishii* shells were obtained from the fruits harvested between the months of June-September 2010, in the marketplace Republic of San Luis Potosi, SLP. Mexico. To obtain the natural biomass, shells rind were washed with trideionized water 72 h under constant stirring, with water changes every 12 h. Subsequently, boiled for 1 h to remove traces of the fruit and was dried at 80°C, for 12 h in the oven, ground in blender and stored in amber vials until use.

#### **2.2. Preparation of stock solution**

An aqueous stock solution (1000 mg/L) of Cr (VI) ions was prepared using K2Cr2O7 salt. pH of the solution was adjusted using 0.1 N HCl or NaOH. Fresh dilutions were used for each study.

#### **2.3. Biosorption studies**

The biosorption capacity of shells biomasses was determined by contacting 100 mL of solution containing different concentration of Cr (VI) (100-1000 mg/L) in 250 mL Erlenmeyer glass flasks, with 1 g of biomass. The mixture was shaken in a rotary shaker at 120 rpm followed by filtration at different times (covering minutes, days and weeks). The filtrate containing the residual concentration of Cr (VI) was determined spectrophotometrically at 540 nm after complexation with 1, 5 Diphenylcarbazide, these method have a detection limit between 0.02-0.5 mg/L of Cr (VI) [19], Cr (III) with Chromazurol S [20], and Cr total by Electrothermal Atomic Absorption Spectroscopy [19]. For the determination of rate of metal biosorption by biomasses from 100 mL (at 100, 200, 300, 400, 500, and 1000 mg/L), the supernatant was analyzed for residual Cr (VI) after the contact period of 1-12 hours. The effect of pH and temperature on Cr (VI) sorption by natural biomass, was determined at pH values of 1, 2, 3, and 4, 28°C, 40°C, and 50°C, respectively. The effect of different doses of biomass ranging from 1 to 5 g/L, with 100 mg/L of Cr (VI) concentrations was determined. The values shown in the results section are the mean from three experiments carried out by triplicate.

#### **2.4. Bioremediation assay**

disposal and imply operational complexity [7]. In this context, considerable attention has been focused in recent years upon the field of biosorption for the removal of heavy metal ions from

The process of heavy metal removal by biological materials is known as biosorption. Biomass viability does not affect the metal uptake. Therefore any active metabolic uptake process is cur‐ rently considered to be a negligible part of biosorption. Various biosorbents have been tried, which include seaweeds, molds, yeast, bacteria, crab shells, agricultural products such modi‐ fied corn stalks, [9], hazelnut shell [10], orange waste [11] and tamarind peel [12]. It has also been reported that some of these biomass can reduce chromium (VI) to chromium (III), like *Litchii cinensis* Sonn peel [13], tea fungal biomass [14], Mesquite [15], Eucalyptus bark [16], red rose's waste biomass [17] and Yohimbe bark [18]. The present study is undertaken with follow‐ ing objective: Investigate the use of different natural biomasses for the biosorption and reduc‐ tion of Chromium (VI) in aqueous solution, and their elimination from contaminated sites.

*Litchi chinensis Sonn, Citrus limonium, Mammea americana, Tamarindus indica, Citrus sinen‐ sisOsbeck, Citrus reticulate, Cucumis melo* L., and *Musa cavendishii* shells were obtained from the fruits harvested between the months of June-September 2010, in the marketplace Republic of San Luis Potosi, SLP. Mexico. To obtain the natural biomass, shells rind were washed with trideionized water 72 h under constant stirring, with water changes every 12 h. Subsequently, boiled for 1 h to remove traces of the fruit and was dried at 80°C, for 12 h in the oven, ground

An aqueous stock solution (1000 mg/L) of Cr (VI) ions was prepared using K2Cr2O7 salt. pH of the solution was adjusted using 0.1 N HCl or NaOH. Fresh dilutions were used for each study.

The biosorption capacity of shells biomasses was determined by contacting 100 mL of solution containing different concentration of Cr (VI) (100-1000 mg/L) in 250 mL Erlenmeyer glass flasks, with 1 g of biomass. The mixture was shaken in a rotary shaker at 120 rpm followed by filtration at different times (covering minutes, days and weeks). The filtrate containing the residual concentration of Cr (VI) was determined spectrophotometrically at 540 nm after complexation with 1, 5 Diphenylcarbazide, these method have a detection limit between 0.02-0.5 mg/L of Cr (VI) [19], Cr (III) with Chromazurol S [20], and Cr total by Electrothermal Atomic Absorption Spectroscopy [19]. For the determination of rate of metal biosorption by biomasses from 100 mL (at 100, 200, 300, 400, 500, and 1000 mg/L), the supernatant was analyzed for residual Cr (VI) after the contact period of 1-12 hours. The effect of pH and

aqueous effluents [8].

208 Applied Bioremediation - Active and Passive Approaches

**2. Material and methods**

in blender and stored in amber vials until use.

**2.2. Preparation of stock solution**

**2.3. Biosorption studies**

**2.1. Biosorbents used**

Four 250 mL Erlenmeyer glass flasks, with 5 g of shell biomass, were added with 20 g of contaminated earth and water with 297 mg Cr (VI)/g earth or 373 mg Cr(VI)/L water, of tannery (Celaya, Guanajuato, México), and the volume was complete to 100 mL with trideionized water. The mixture was shaken in a rotary shaker at 120 rpm followed by filtration using Whatman filter paper No. 1. The filtrate containing the residual concentration of Cr (VI) was determined with 1, 5 diphenylcarbazide [19].

#### **2.5. Determination of hexavalent, trivalent and total Cr:**

Hexavalent and trivalent chromium were quantified by a spectrophotometric method employing diphenylcarbazide and chromazurol S, respectively [19, 20], and total Chromium by Electrothermal Atomic Absorption Spectroscopy [19].
