www.els-journal.com Jenny Gustavsson 1, * Sepehr Shakeri Yekta 1, * Anna Karlsson 2 Ulf Skyllberg 3 Bo H. Svensson 1 1 Department of Thematic Studies-Water and Environment, Linköping University, Linköping, Sweden 2 Scandinavian Biogas Fuels AB, Stockholm, Sweden 3 Department of Forest Ecology and Management, Swedish University of Agricultural Sciences, Ume ˚ a, Sweden Research Article Potential bioavailability and chemical forms of Co and Ni in the biogas process—An evaluation based on sequential and acid volatile sulfide extractions Several previous studies reported stimulatory effects on biogas process performance after trace metal supplementation. However, the regulation of the bioavailability in relation to chemical speciation, e.g. the role of sulfide is not fully understood. The objective of the present study was to determine the effect of sulfide on chemical speciation and bioavailability of Co and Ni in lab-scale semicontinuous stirred bio- gas tank reactors treating stillage. The chemical forms and potential bioavailability of Co and Ni were studied by sequential extraction, analysis of acid-volatile sul- fide (AVS), and simultaneously extracted metals. The results demonstrated that Ni was completely associated to the organic matter/sulfide fraction and AVS, suggest- ing low potential bioavailability. Cobalt was predominantly associated to organic matter/sulfide and AVS, but also to more soluble fractions, which are considered to be more bioavailable. Process data showed that both Co and Ni were available for microbial uptake. Although the actual bioavailability of Co could be explained by association to more bioavailable chemical fractions, the complete association of Ni with organic matter/sulfides and AVS implies that Ni was taken up despite its expected low bioavailability. It was concluded that extensive Co- and Ni-sulfide precipitation did not inhibit microbial uptake of Co and Ni in the reactors. Keywords: Biogas production / Cobalt / Nickel / Potential bioavailability / Speciation Received: September 27, 2012; revised: December 19, 2012; accepted: February 23, 2013 DOI: 10.1002/elsc.201200162 1 Introduction The importance of trace metal availability for microbial activities as a prerequisite to obtain efficient and stable biogas processes is reported in several studies [1–13]. In the listed reports, Co and Ni stand out as they are often shown to have stimulatory effects on process performance. However, the factors regulating their bioavailability and chemical speciation need to be further un- derstood in order to determine proper Co and Ni dosage in biogas reactors. Particularly the role of sulfide on Co and Ni bioavailabi- lity is still unclear. Sulfur-containing organic matter and sulfate Correspondence: Sepehr Shakeri Yekta (sepehr.shakeri.yekta@liu. se), Department of Thematic Studies-Water and Environment, Link¨ oping University, SE-581 83 Link¨ oping, Sweden Abbreviations: AAS, atomic absorption spectroscopy; AVS, acid-volatile sulfide; AVS-Me, simultaneously extracted metals; HRT, hydraulic reten- tion time; SCSTR, semicontinuous stirred tank reactors; SE, sequential extraction; TS, total solids; VFA, volatile fatty acid; VS, volatile solids in substrates are converted to sulfide during anaerobic digestion [14], and several studies reported the importance of sulfide as a regulator for metal bioavailability due to high affinity of sulfide for binding metal ions [9, 15–22]. It is reasonable to believe that the bioavailability of Co and Ni will be limited by the precipita- tion of Co- and Ni-sulfides, due to their low solubility products [15–17]. Nevertheless, the precipitation/dissolution rates of Co- and Ni-sulfides and the competition of metal-binding organic ligands have also been suggested to affect Co and Ni bioavail- ability [9, 19]. The starting point for the present study was the observations that digestion of stillage (containing 12 g total S per kg total solids [TS] –1 ) in semicontinuous stirred tank reactors (SCSTRs) was improved by supplementation of Co and Ni (0.5 and 0.2 mg L –1 , respectively) [13]. It was demonstrated that after a period of un- stable process performance, the processes responded to daily Co and Ni supplementation by an increase in methane production ∗ Both authors contributed equally to this work. 572 Eng. Life Sci. 2013, 13, 572–579 C  2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim