Efficacy of Cassava Peel Extracts for the Removal of Heavy Metals from Hospital Sewage Sludge in Nigeria

Background. The use of strain-specific microbial fermentation in the production of organic acids for the removal of heavy metals from sludge has been extensively studied. However, there is scarce information on the use of microflora for fermentation. Objectives. To assess the efficacy of cassava peel extracts for the removal of heavy metals from hospital sewage sludge in Nigeria. Methods. A composite sewage sludge sample was collected from the University College Hospital sewage treatment plant in Ibadan, Nigeria and analyzed for heavy metals using standard methods. Aspergillus niger fermentation and crude fermentation extract were obtained from the cassava peels strain of Aspergillus niger and indigenous microflora, respectively. The experiment was carried out by adding 10 ml of the treatment to 3 g of each sludge sample (extracts and controls) at varied temperatures (room and elevated) and pH (3–5). The mixture was centrifuged after a contact time of 1–12 days at 1000 rpm for 1 hour. The filtrate was analyzed for heavy metals concentrations and compared with the standards. Data were analyzed using descriptive statistics and adsorption models. Results. Mean heavy metal concentrations in the sludge were estimated for copper (2.22±0.2 mg/kg), zinc (52.3±0.1 mg/kg), chromium (1.46±0.1 mg/kg), nickel (5.6±0.01 mg/kg), and lead (1.9±0.1 mg/kg) and were below permissible limits. Optimum heavy metal removal for Aspergillus niger fermentation extract at room temperature was achieved on day 12 at pH 3.5 for zinc (74.5%), while optimum heavy metal removal at elevated temperature was achieved on day 9 at pH 3.0 for lead (79.3%). The optimum pH for crude fermentation extract lies between pH 3.0–4.5 for nickel (76.2%) at room temperature and chromium (76.6%) at elevated temperature. Conclusions. Crude fermentation extract of cassava peel was found to be effective in removing heavy metals from sewage sludge. Therefore, its use could be adopted and promoted for removing heavy metals from sewage sludge to achieve safe disposal. Competing Interests. The authors declare no competing financial interests.


Introduction
Sludge is a solid residue generated during sewage treatment in industrial treatment plants. It may contain organic and inorganic compounds, non-essential trace elements, microorganisms, and the eggs of parasitic organisms. 1,2 Increasing rates of urbanization and industrialization are responsible for the large volume of sludge being generated globally from domestic residences, industries, the agricultural and commercial sector, and institutions. 3,4 Hospitals produce a significant volume of wastewater per day, which contains microorganisms, heavy metals, toxic chemicals and radioactive elements. According to Ekhaise and Omavwoya, differences in hospital sludge compared to municipal sludge are due to patient therapeutics (e.g. antibiotics, chemotherapy) which increase the content of organic matter, chemicals, metals, and pathogenic organisms. 5 Heavy metals in hospital sludge originate from feces, paint, wear and tear of utensils and equipment, and radioactive materials from radiology departments. 5,6 Sewage sludge is often used for agricultural applications due to its abundance of organic matter and nutrients. 7 However, the presence of hazardous contents such as heavy metals, pathogenic organisms, and soluble salts limits its land application. 8, 9 Remediation of the heavy metals content of dewatered sludge is needed, since heavy metals are not degradable and once released into the soil environment, they have the potential to deteriorate the quality of soil, water (surface and underground), and human health and safety. 10-14 Methods of extraction such as bioremediation, composting, stabilization of metals, and acid treatment have been employed in Research the removal of heavy metals from sludge. 15- 18 However, due to the complex matrix of sewage sludge and bonding nature of metals in organic solids, metals can only be satisfactorily solubilized under extreme acidic conditions achieved in high redoxpotential environments using chemical leaching or bioleaching techniques. 7 Inorganic chelating agents have been productively used in heavy metal removal as they form stable complexes over a broad pH range. However, their consistent use tends to decrease soil productivity and impair the physic-chemical structure of soils, thereby rendering soil unfit for further use. 11,16 Other disadvantages of their use include persistence in the environment, high cost, and adverse health effects. For the bioleaching of metals from sewage sludge, organic acids are more promising than inorganic chelating agents, since extraction can occur in mildly acidic conditions (pH 3-4) and they are biodegradable and have low environmental impact. There is a need for a sustainable, environmentallyfriendly, readily available, inexpensive, and efficient extractant for the removal of heavy metals. [19][20][21][22][23][24] Nigeria is the largest cassava producer in the world, producing one-third more than Brazil and almost double the production capacity of Thailand and Indonesia, with an annual production of 49 million metric tons. 25-27 Cassava peels represent a unique renewable carbon source and millions of tons are generated every day. 28 Improper disposal of cassava peels includes littering the streets, dumping refuse in drains and causing blockages, or being discarded in bushy areas, becoming an eyesore, and creating breeding grounds for disease vectors and pathogens. The presence of cassava peels in the wet and heterogenous waste stream serves as an attractant and breeding site for houseflies, which are implicated in the transmission of typhoid and paratyphoid fevers, diarrhea, dysentery, cholera, gastroenteritis, amoebiasis, helminthic infestations, conjunctivitis, poliomyelitis, and other diseases that can spread by mechanical transmission. 29 The use of strain-specific microbial fermentation in the production of organic acids from agricultural wastes for bioleaching of heavy metals has Adeolu, Adewoye

Methods
University College Hospital sewage treatment plant is located in Ibadan, Oyo State, Nigeria (Figure 1). The processes of the plant involve the conversion of organic matter into inorganic matter before discharging the treated effluent into the recipient environment (the Olojuoro River), while the sludge is processed into organic manure for agricultural purposes.

Collection and analysis of dewatered sewage sludge
Composite sewage sludge was collected from the treatment plant, dried, homogenized, and analyzed for physico-chemical parameters (organic matter, total carbon, total nitrogen, and total phosphorus) using standards procedures. 9,33 The gradient concentrations of heavy metals in the sludge were determined using an atomic absorption spectrophotometer (Buck Scientific Model 210 VGP).

Isolation of bioleaching agent
The locally isolated citric acidproducing strain of Aspergillus niger was identified with the use of Czepak-Dox Agar. Aspergillus niger fermentation extract and crude fermentation extract were obtained from the fermentation of 3 grams of cassava peels for 13 days using the acids-producing strain of A. niger as inoculum and indigenous microflora in the cassava peels. 34-36 These extracts were then used for bioleaching heavy metals from the sewage sludge, while commercial citric acid served as a control. The fermented materials were extracted with distilled water. Fermented sample extracts were collected every 48 hours for an estimation of citric acid over a period of 12 days. Citric acid produced by fermentation was estimated by the pyridine acetic anhydride method. 37

Extraction experiments
Heavy metal removal experiments were carried out to determine the efficacy of extracts (leaching agents or extractants) at varied optimum conditions (pH, temperature and contact time) using batch experiments as modified by Okareh and Enesi. 2 The analyses were carried out at unadjusted and adjusted pH for all of the extractants. Adjustments of pH were carried out to determine the effect of progressive acidification on the removal of heavy metals from the sludge using 1 m hydrochloric acid. The extraction was carried out at varied pH levels (3)(4)(5) and contact times of 1, 3, 6, 9 and 12 days at ambient (28 o C) and elevated temperature (45 o C) for each extractant. For each extraction, the centrifuge tube containing 3 g of sieved sewage sludge was filled with 100 ml of the extractant. The tubes were stirred continuously on a rotary shaker at 150 rpm and centrifuged at

Data management
Analyses of heavy metals were carried out in triplicate and data was subjected to descriptive statistical analysis using the Statistical Package for the Social Sciences (SPSS 21) software.
In addition, the adsorption model was used to calculate the amount of metal ions leached (Equation 1) and percentage removal of heavy metals (removal efficiency) from the sewage sludge (Equation 2).

Equation 2 (%) = (C1 V1)/Cs Ms x 100
where Q e is the metal uptake (mg/kg); C 1 and C s are the concentrations of the metal in extractants (mg/l) and sludge (mg/l), respectively, M s is the mass of the sludge (kg) and V is the volume of the extractant (liter) (l).

Results
The pH of the sewage sludge sample was 6.20±0.2, organic matter content was measured at 68.13±0.5%, total carbon was 35.13±0.5%, and the heavy metal contents (mg/kg) are shown in Table 1.  Table 3).

Efficacy of fermentation extracts for heavy metal removal
Optimum A. niger fermentation extraction occurred at elevated temperature with Pb (79.3%) and the lowest percentage extraction occurred for Ni (74.1%) as shown in Table 3. For crude fermentation, optimum extraction occurred at elevated temperature with Cr (76.6%) and the lowest percentage extraction occurred for Zn (75.1%) ( Table 3).

Effect of pH on heavy metal removal
For some of the extractions, there was a corresponding increase in removal efficiency with an increase in pH from 3-4, followed by a decrease

Effects of contact time on heavy metal removal
Generally, there was a decrease of removal efficiency with increase in time from day 1-9. In a few instances, there was a corresponding increase in heavy metal removal efficiency as contact time increased and in others there was a downward trend in removal efficiency as contact time increased, as represented in Figure  7-11.

Discussion
The average pH of the sludge sample was 6.20, which is within the WHO permissible limit of 6.5-8.5 and NESREA limit of 6.0-9.0; therefore, the discharge of sludge at a pH of 6.20 is acceptable to the receiving environment. The mean pH indicated that the dewatered sewage sludge was slightly acidic, as shown in

Effects of treatment/extractant
Findings showed that A. niger fermentation extract was more Adeolu, Adewoye

Effects of pH on heavy metal removal
The optimum pH for A. niger lies between 3-4.0 for the removal of Pb (79.3%), while the crude fermentation extract lies between pH 3-4.5 for the removal of Ni (76.2%) at room temperature and Cr (76.6%) at elevated temperature (Figures 2-6). The removal of heavy metals by progressive acidification for all of the analyzed metals depended on pH. These findings were consistent with previous studies which reported that pH is a dominant factor which influences the cation exchange capacity of the sludge, hence altering the redistribution and exchangeability of heavy metals in the sludge. 7,12, 18 As pH increased, the onset of the metal hydrolysis and precipitation commenced. When the pH of the extractant was increased from 3 to 4, there was a corresponding increase in deprotonation of the extractant's surface, leading to a decrease in hydrogen ions on the surface. This creates more negative charges on the extractant's surface, which favors extraction of positively charged species.

Effects of contact time on heavy metal removal
Generally, there were initial increases in removal efficiency with corresponding increases in contact time, as depicted in Figure 7-11. The findings of the present study vary greatly from those of Stylianou et al., who reported that maximum heavy metal removal efficiency was attained in less than 24 hours using synthetic inorganic chelating agents. 7 It should be noted that the initial increased rate was due to the availability of the uncovered surface area of the extractants since the kinetics depends on the surface area of the extractants. Extraction takes place at the more Adeolu, Adewoye

Abbreviations: ANFE (Aspergillus niger fermentation extract); CFE (Crude fermentation extract); CCA (Commercial-grade citric acid (control)).
Research active binding sites. As these sites are progressively filled the more difficult the extraction becomes, as the extraction process tends to be more unfavorable.

Management of cassava peel residues and treated sludge
The biosol (solid fraction of the fermented cassava peel) was added to the soil as recommended by Ubalua. 46 The treated sludge (acidified) was dried, treated with lime, and applied to farmland as fertilizer and soil conditioner of high hygienic quality, as stated by Uriah et al. 47

Conclusions
Cassava peel wastes, which are often an environmental nuisance, could be used to produce resource materials (organic acids) which can be used to remove heavy metals of public health concern from industrial effluent/sewage sludge. Converting these wastes into organic acids has greatly helped to reduce its environmental risk and enhances effective waste management.

Abbreviations: ANFE (Aspergillus niger fermentation extract); CFE (Crude fermentation extract); CCA (Commercial-grade citric acid (control)).
Research peel wastes in removing heavy metals from sewage sludge indicated that crude fermentation extract of cassava peel waste competed favorably with strain specific of Aspergillus niger in removing heavy metals from sewage sludge. In addition, the fermentation extract showed appreciably better heavy metal removal efficiency compared with commercial grade organic acids. Cassava peel wastes represent a cheaply available substrate associated with low fermentation process costs for obtaining crude fermentation extract, making this an attractive method for the removal of heavy metals from hospital sewage sludge waste.