Two stage biodiesel and hydrogen production from molasses by oleaginous fungi and Clostridium acetobutylicum ATCC 824

https://doi.org/10.1016/j.ijhydene.2013.12.106Get rights and content

Highlights

  • Oleaginous fungi are promising source for biodiesel production.

  • Epicoccum nigrum and Alternaria alternata are the best for biodiesel production from molasses.

  • Molasses spent media were used for hydrogen production by Clostridium acetobutylicum.

  • Exploit sugars of spent media would increase the economic feasibility of bioenergy.

  • Interlinking the biodiesel and hydrogen production improve biofuel technology.

Abstract

In the present study biodiesel was produced by various fungal species isolated from Egypt using sugarcane molasses as substrate. In the first stage 6 oleaginous fungi, namely, Alternaria alternata, Cladosporium cladosporioides, Epicoccum nigrum, Fusarium oxysporum, Aspergillus parasiticus and Emericella nidulans var. lata were used for lipid production. Subsequent to fungal cultivation on sugarcane molasses the cultures were filtered and biodiesel was prepared by direct esterification of dry fungal biomass. Methyl esters of palmitic, stearic, linoleic and elaidic represented the major components while palmitoleic represented a minor component of biodiesel produced from tested oleaginous fungi. In the second stage, the spent medium of fungal culture was used as the fermentation medium for hydrogen production by Clostridium acetobutylicum ATCC 824. The maximum total H2 yield was obtained with the spent medium of E. nigrum and A. alternata. The results presented in this study suggest a possibility of interlinking the biodiesel production technology by fungi with hydrogen production by C. acetobutylicum ATCC 824 to exploit the residual sugars in the spent media and therefore increase the economic feasibility of the biofuel production from molasses.

Introduction

Fossil fuels are not renewable and will be exhausted sooner or later [1]. The increase in prices of petroleum based fuels, future depletion of worldwide petroleum reserves and environmental polices to reduce CO2 emissions have stimulated research into the development of biotechnology to produce clean energy from renewable resources [2]. Thus, it is necessary to find alternative energy sources that are renewable and environmentally friendly [3], [4], [5], [6], [7], [8]. Hydrogen and biodiesel are very promising alternative energy sources and have received more attention all over the world in recent years.

Biodiesel fuel is fatty acid mono alkyl (methyl, propyl or ethyl) esters produced from natural, renewable sources such as vegetable oils or animal fats by transesterification of such triglycerides [9], [10]. Biodiesel is rather an attractive alternative to the conventional petroleum diesel fuels because of its biodegradable, nontoxic and clean renewable characteristics [10], [11]. One way to increase the world bio oil production that would cause low ecosystem impact is to use lipids from single-cell oil microorganisms [12] for biodiesel production. Oleaginous yeasts and fungi have also been considered as potential oil sources for biodiesel production because they accumulate large amounts of lipids. Among these microorganisms, particular attention has been dedicated to Epicoccum purpurascens [13] and Mortierella isabelina [14].

Hydrogen is one of the most abundant elements in the universe. It is an odorless, colorless, tasteless and non-poisonous gas. Hydrogen gas has been proposed as the ultimate transport fuel for vehicles and vessels because of its non-polluting characteristics. When hydrogen is used as a fuel, it generates no pollutants but produces water. Hydrogen also enables the use of highly efficient fuel cells to convert chemical energy to electricity [15]. Another major use of hydrogen is in reduction reactions to produce industrial chemicals.

Hydrogen is produced mainly from natural gas, a finite resource, through steam reforming, a process that generates large quantities of carbon dioxide (CO2) which is a principal cause of global warming. Hydrogen production through dark or photofermentative conversion of organic substrates is of great interest due to its dual function of waste reduction and clean energy production, thereby acting as a promising option for biohydrogen production [16], [17], [18]. Anaerobic bacteria capable of hydrogen production can use various renewable biomass and numerous agriculture, municipal and food processing waste and wastewater sources [19], [20].

Molasses is the by-product from sugar industry and is considered as one of the most promising feedstocks in the production of biofuel. Black strap molasses is another byproduct of the sugar industry, obtained during sugarcane refining and, like beet molasses, has a high sugar content, around 50% (36% sucrose, 6% fructose, 3% glucose), making it a promising substrate for biohydrogen production [21]. Sugarcane molasses is also an important organic waste due to its high sugar content (55%) and high volume of production. It is even more viscous than beet molasses and the total sugar content is higher. The availability and cost of sugarcane molasses make it an attractive feedstock for use in many countries.

The generation of lipid by fermentative oleaginous fungi accompanies the formation of organic acids as metabolic products [22]. The accumulation of these acid results in a sharp drop in culture pH and subsequent inhibition of fungal growth. But, it is somewhat difficult to achieve the complete utilization of molasses sugar to lipid through fungal fermentation. This appears to be one of the major bottlenecks in the molasses fermentation process for lipid production. So, the process outlined in the present paper takes into consideration the non-utilization of sugar and the acids produced therein (spent media). Combining fermentative oleaginous fungi with hydrogen producing Clostridium could provide an integrated system for maximizing the biofuel yield from sugarcane molasses. In such a system, the fermentation of spent medium containing residual sugar and organic acids generated by fungi, which are then converted into hydrogen by Clostridium in the second step in a bioreactor. In the present study, sugarcane molasses was considered as a preliminary substrate for biodiesel production by various fungal species isolated from Egypt and the spent medium from this process was used as a substrate for hydrogen production by the strictly anaerobic Clostridium acetobutylicum ATCC 824 that also produces an acetone–butanol–ethanol mixture as well as hydrogen.

Section snippets

Fungal inoculums

Forty-five fungal isolates recovered from various sources in Upper Egypt and maintained in potato dextrose agar (PDA) medium at 4 °C were used in this study. Identification of fungal isolates was carried out based on the morphological features using the keys of Pitt [23], Raper and Fennell [24], Ellis [25], Kendrick [26] and Domsch et al. [27]. Fungi were grown on PDA plates at 25 ± 1 °C. Seven day old culture PDA plates of fungi grown at 25 ± 1 °C were used for inoculum preparation. Fungal

Lipid accumulation by fungi

Sugarcane molasses is an agro-industrial by-product often used in fermentation due to the presence of fermentative sugars, being an optimal for the microbial metabolism. In this study, our rationale is to investigate the potency of forty-five fungal isolates to produce oils in a remarkable manner for biodiesel production from low cost waste molasses. Furthermore, the fatty acid methyl ester composition of biodiesel from the detected oleaginous fungi was investigated. In this study 6 fungal

Conclusion

The current study indicated that the six fungal species were found to be oleaginous; containing more than 20% lipids per dry mass. A. alternata was the highest lipid producer from sugarcane molasses. Methyl esters of palmitic, stearic, linoleic and elaidic acids represented the major components of biodiesel produced from tested oleaginous fungi. The spent medium from first stage of sugarcane molasses fermentation by oleaginous fungal species is found to have the capacity for the dark-production

Transparency declaration

The authors declare no conflicts of interest.

Acknowledgments

The authors would like to thank Prof. Dr. George Bennett, Rice University, USA, for providing Clostridium acetobutylicum ATCC 824, precious comments and critical reading of the manuscript. The authors are very grateful to the precious suggestions, constructive comments and careful corrections made by three anonymous reviewers for further improvements of this paper. This research was supported by Assiut University Fund.

References (72)

  • C.W. Forsberg

    Future hydrogen markets for large-scale hydrogen production systems

    Int J Hydrogen Energy

    (2007)
  • D.B. Lata et al.

    Effect of light on generation of hydrogen by Halobacterium halobium NCIM 2852

    Int J Hydrogen Energy

    (2007)
  • V. Gadhamshetty et al.

    Photofermentation of malate for biohydrogen production – a modeling approach

    Int J Hydrogen Energy

    (2008)
  • Y. Tao et al.

    Characteristics of a new photosynthetic bacterial strain for hydrogen production and its application in wastewater treatment

    Int J Hydrogen Energy

    (2008)
  • C.C. Wang et al.

    Producing hydrogen from wastewater sludge by Clostridium bifermentans

    J Biotechnol

    (2003)
  • S.-K. Han et al.

    Biohydrogen production by anaerobic fermentation of food waste

    Int J Hydrogen Energy

    (2004)
  • A. Beopoulos et al.

    Yarrowia lipolytica as a model for bio-oil production

    Prog Lipid Res

    (2009)
  • J. Folch et al.

    A simple method for the isolation and purification of total lipids from animal tissues

    J Biol Chem

    (1957)
  • M.M. Bradford

    A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding

    Anal Biochem

    (1976)
  • G. Vicente et al.

    Biodiesel production from biomass of oleaginous fungus

    Biochem Eng J

    (2009)
  • D.W. Penfold et al.

    Increased hydrogen production by Escherichia coli strain HD701 in comparison with the wild-type parent strain MC4100

    Enzyme Microb Technol

    (2003)
  • C. Angerbauer et al.

    Conversion of sewage sludge into lipids by Lipomyces starkeyi for biodiesel production

    Bioresour Technol

    (2008)
  • S. Papanikolaou et al.

    Single cell oil (SCO) production by Mortierella isabellina grown on high-sugar content media

    Bioresour Technol

    (2004)
  • S. Fakas et al.

    Evaluating renewable carbon sources as substrates for single cell oil production by Cunninghamella echinulata and Mortierella isabellina

    Biomass Bioenergy

    (2009)
  • C.N. Economou et al.

    Single cell oil production from rice hulls hydrolysate

    Bioresour Technol

    (2011)
  • Z. Ruan et al.

    Evaluation of lipid accumulation from lignocellulosic sugars by Mortierella isabellina for biodiesel production

    Bioresour Technol

    (2012)
  • P. Dey et al.

    Comparative lipid profiling of two endophytic fungal isolates Colletotrichum sp. and Alternaria sp. having potential utilities as biodiesel feedstock

    Bioresour Technol

    (2011)
  • X.W. Peng et al.

    Single cell oil production in solid-state fermentation by Microsphaeropsis sp. from steam-exploded wheat straw mixed with wheat bran

    Bioresour Technol

    (2008)
  • O. Tehlivets et al.

    Fatty acid synthesis and elongation in yeast

    Biochim Biophys Acta

    (2007)
  • Y.H. Li et al.

    High-density cultivation of oleaginous yeast Rhodosporidium toruloides Y4 in fed-batch culture

    Enzyme Microbiol Technol

    (2007)
  • H. Malaoui et al.

    Purification and characterization of the 1,3-propanediol dehydrogenase of Clostridium butyricum E5

    Enzyme Microb Technol

    (2000)
  • X. Chen et al.

    Stoichiometric analysis of biological hydrogen production by fermentative bacteria

    Int J Hydrogen Energy

    (2006)
  • T. Lütke-Eversloh et al.

    Metabolic engineering of Clostridium acetobutylicum: recent advances to improve butanol production

    Curr Opin Biotechnol

    (2011)
  • L.T. Angenent et al.

    Production of bioenergy and biochemicals from industrial and agricultural wastewater

    Trends Biotechnol

    (2004)
  • C.-L. Cheng et al.

    Hydrolysis of lignocellulosic feedstock by novel cellulases originating from Pseudomonas sp. CL3 for fermentative hydrogen production

    Bioresour Technol

    (2011)
  • Y.-C. Lo et al.

    Fermentative hydrogen production from hydrolyzed cellulosic feedstock prepared with a thermophilic anaerobic bacterial isolate

    Int J Hydrogen Energy

    (2009)
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