Elsevier

Chemical Engineering Journal

Volume 337, 1 April 2018, Pages 764-771
Chemical Engineering Journal

Continuous fermentation of xylose to short chain fatty acids by Lactobacillus buchneri under low pH conditions

https://doi.org/10.1016/j.cej.2017.12.100Get rights and content

Highlights

  • First report on continuous low pH bacterial fermentation on short chain fatty acid production.

  • Batch fermentation kinetics of L. buchneri using xylose was established.

  • High lactic (67 g L−1) and acetic acid (32 g L−1) levels were achieved in fed-batch fermentation.

  • High acid productivities were achieved in immobilized cell fermentation systems.

  • A mathematical model could describe changes in productivity with different dilution rates.

Abstract

Short chain fatty acid fermentation was carried out using Lactobacillus buchneri ATCC 4005 under anoxic conditions at low pH values using xylose as substrate. Fermentation studies were conducted with free cells and with cells immobilized on alumina fibers in batch, fed-batch, and continuous reactors. The results indicate that xylose is readily utilized by L. buchneri as a carbon source with an estimated maximum specific growth rate (µmax) of 0.15 h−1 for free cell and immobilized cell fermentations. Lactic and acetic acids were produced at high concentration levels of 67 g L−1 and 32 g L−1, respectively in fed-batch fermentation without any apparent cell inhibition. Volumetric productivities of 6.4 g L−1h−1 for lactic acid, and 4.3 g L−1h−1 for acetic acid were obtained at a dilution rate of 0.5 h−1. The Luedeking-Piret model that relates cell growth to product formation is utilized to develop a mathematical model of the continuous fermentation system. The model predictions for volumetric acid productivities as a function of dilution rates are presented, and are shown to agree well with experimental data. This study is the first demonstration of a continuous fermentation process for the conversion of xylose to lactic and acetic acids using L. buchneri at a pH value of 4.0.

Introduction

The utilization of biomass and biomass wastes to produce useful chemicals and fuels is a sustainable approach to reduce our dependence on petroleum resources, and for the effective management of wastes. A majority of bioconversion studies in the past have focused on the use of biomass for the production of biofuels such as ethanol, methane and hydrogen [1], [2]. However, the theoretical yields of ethanol and hydrogen via bioconversion processes are only about 51% and 33% of the total sugar consumed, respectively, and this leaves substantial amounts of residuals that need to be disposed off. Often these processes face challenges and limitations due to product and/or substrate inhibition at moderate to high loading concentrations [3]. Anaerobic bioconversion of wastes to methane is a widely used technology worldwide, and the methane generated is typically used as a fuel. The capital and operating costs of anaerobic digestion systems are high due to the need to operate at thermophilic temperatures, and from the high reactor volumes required due to the high retention times needed for substrate utilization [4]. Moreover the product value of methane is quite low at present due to the abundant supply of natural gas. Therefore, production of high-value chemicals such as carboxylic acids is more attractive compared to methane production. The biobased renewable chemicals market is expected to increase globally from $57 billion currently to $83 billion by 2018 [5].

In the past two decades, there have been substantial advancement in development of pretreatment processes for lignocellulosic biomass to overcome the recalcitrance [2], [3]. Xylose is the major sugar obtained from the pre-treatment of hemicellulosic biomass, but is often underutilized during fermentations due to limited ability of microorganisms to utilize pentose sugars [3], [6]. A few organisms have been found to use xylose to some extent along with glucose. These include Bacillus coagulans for lactic acid production, Candida shehatae, Pichia stipitis or Pachysolen tannophilus for ethanol production, Actinobacillus succinogenes for succinic acid production, and Geobacillus or Klebsiella oxytoca for acetoin and 2,3-butanediol production with ≤85% of theoretical yield [7], [8], [9].

The production of platform chemicals belonging to carboxylic acids such as lactic, acetic, propionic and butyric acids from biomass has been gaining significant attention in recent years [5]. Among carboxylic acids, lactic and acetic acids have a wide range of industrial applications, and the demands for these acids are increasing each year. While the food industry is a major market for lactic and acetic acids, new markets in the production of polymers such as polyethylene terephthalate (PET), ploylactic acid (PLA), and polyvinyl acetate (PVA) are expanding the demand for these chemicals [5], [6]. In addition, there is a large potential market for acetic acid in the production of biodegradable deicing salts such as calcium magnesium acetate and potassium acetate [10].

In general, most of the microorganisms producing carboxylic acids via fermentation of lignocellulosic materials have lower growth rates or less affinity towards xylose, low tolerance to inhibitors or high product concentrations, and they typically operate at pH values ≥5.0 [11], [12]. This tends to lower product yields when compared with other sugars, and increases the downstream processing costs, as the acid separation is more efficient at low pH values. Selective separation of an organic acid from fermentation broth containing biomass hydrolysate is often associated with a high processing cost. Therefore, economic feasibility of using biomass or low value biomass wastes largely depends on the downstream processing efficiency [13]. Maximizing the product yield and substrate conversion efficiency in the upstream step and increasing the recovery efficiency in the downstream step will help enable the production of high-value biochemicals at lower costs. Lee, et al. [14] have studied the separation of lactic and acetic acids using poly(4-vinylpyridine) resin (PVP) in a simulated moving bed (SMB). They achieved good separation efficiency for lactic and acetic acids present in a synthetic broth at pH 2.2, and they obtained a lactic acid recovery yield of over 93% at 99.9% purity.

The upstream process efficiency depends on the choice of microbial specie that is able to overcome the limitations discussed earlier. Lactic acid bacteria (LAB) are widely used in agricultural and food industries as nutritional or food-supplements, inoculants and preservatives. LAB can be divided into homolactic and heterolactic groups depending on the ability to produce lactic acid alone or produce other products in addition to lactic acid. Heterolactic bacteria are preferred due to the ability of these bacteria to provide increased aerobic stability [15] and for the production of variety of byproducts in addition to lactic acid [16], [17], [18]. Among, the heterofermentative bacteria Lactobacillus sp., Lactobacillus buchneri has been reported in a number of studies on its ability to produce a variety of byproducts and to increase the aerobic stability of silage [17], [18]. The FDA has approved the use of silage isolates of L. buchneri as bacterial inoculants for ensiling various grains since 2001 [19]. The reports from previous ensiling and fermentation studies suggest that L. buchneri is able to tolerate low pH conditions [16]. The tolerance of L. buchneri to volatile fatty acids, high product levels and low pH has been reported in various studies involving silage preservation [15], [19], [20]. This tolerance to high acid concentrations along with their ability to ferment under anoxic or microaerophilic conditions makes L. buchneri a good candidate for investigation for bioprocessing applications [16], [20]. In LABs, the microaerophilic conditions or presence of oxygen helps in more conversion of pyruvate to acetic acid which in turn benefits catabolic activities of the cell [21].

There have been no studies reported in the literature on the kinetics of xylose fermentation by L. buchneri to produce organic acids. Liu, et al. [17] have shown that L. buchneri NRRL-B-30929 can utilize xylose at pH 5.0 to 6.0 and generate lactate at 60% yield. Liu, et al. [22] also studied in flask the ability of NRRL-B-30929, NRRL-1837 and DSM 5987 to utilize corn stover and wheat straw hydrolysate sugars at pH 5.0. Only one study was reported at pH 4.0 and this study utilized glucose as substrate, and showed a conversion of approx. 50% to produce lactate and ethanol at a molar ratio of 1.03 to 1.0 [17]. These exploratory studies were more focused on bioethanol production, and lack kinetic data or data on continuous fermentations operations that would be required for commercialization. Liu et al. [17] and Jung and Lovitt [23] conducted studies with xylose and glucose as substrate respectively, at pH ≥ 5.0, and reported the acid extraction and separation efficiencies to be low at such pH values. This paper examines the kinetics of xylose fermentation by L. buchneri at pH ∼ 4 using free and immobilized cells in batch and continuous operation modes.

Section snippets

Microorganisms and growth conditions

Lactobacillus buchneri (ATCC, 4005) obtained from American Type Culture Collection (Manassas, VA, USA) was used in this work. The culture was maintained in the De Mann- Rogosa-Sharpe (MRS) media supplemented with 20% glycerol (v/v) at −25 °C. The culture was routinely tested for viability and cultured in MRS medium and incubated at 36 °C under static aerobic conditions.

Seed inoculum preparation

The fermentation studies were initially conducted in batch serum bottles with a working volume of 0.05 L. The composition used

Batch fermentation kinetics

The results from the batch fermentation experiments shown in Fig. 1a indicate that L. buchneri, is able to ferment up to 100% the xylose substrate within 35 h of fermentation time. The lactic acid and acetic acid concentration reached up to 52 and 33 g L−1, respectively, while the biomass yield reached up to 0.1 g dry cells g−1 xylose. The optimal parameter values determined from the Luedeking-Piret equation for both lactate and acetate resulted in an α value of 5.6 and 3.6, respectively, while

Discussion

The major costs in bioprocessing of biomass to produce value added chemicals stem from (1) upstream processing of raw materials and fermentation; and (2) downstream processing costs in the separation and purification of the products from the fermentation broth. The cost of upstream processing is mainly driven by raw material costs. The use of refined sugars poses a major obstacle in the commercialization of processes to produce biobased chemicals due to the high cost of refined sugars [6]. Use

Conclusions

In this study, the effect of dilution rate and cell immobilization on continuous fermentation of xylose to lactic and acetic acids was investigated. The process was carried out at different dilution rates varying from 0.01 to 0.08 h−1 in the case of free cell fermentation and 0.04 to 0.5 h−1 in the case of immobilized cell fermentation at pH ∼ 4. The volumetric productivities obtained at a dilution rate of 0.5 h−1 were, 6.4 g L−1h−1 for lactic acid and 4.3 g L−1h−1 for acetic acid with the

Acknowledgments

The authors are grateful to the NSF Environmental Engineering program for funding the project (CBET Award no: 1600075). The authors are also grateful to Department of Civil Engineering Kansas State University, for instrumentation funding.

References (40)

  • P.L.H. Yeh et al.

    An improved kinetic-model for lactic-acid fermentation

    J. Fermen. Bioeng.

    (1991)
  • A. Garde et al.

    Lactic acid production from wheat straw hemicellulose hydrolysate by Lactobacillus pentosus and Lactobacillus brevis

    Bioresour. Technol.

    (2002)
  • A. Yousuf et al.

    Recovery of carboxylic acids produced during dark fermentation of food waste by adsorption on Amberlite IRA-67 and activated carbon

    Bioresour. Technol.

    (2016)
  • Y.X. Zhang et al.

    Enhanced D-lactic acid production from renewable resources using engineered Lactobacillus plantarum

    Appl. Microbiol. Biotechnol.

    (2016)
  • Y.J. Wee et al.

    Lactic acid production by Lactobacillus sp RKY2 in a cell-recycle continuous fermentation using lignocellulosic hydrolyzates as inexpensive raw materials

    Bioresour. Technol.

    (2009)
  • S.R. Parekh et al.

    Continuous production of acetate by Clostridium thermoaceticum in a cell-recycle membrane bioreactor

    Enz. Microb. Technol.

    (1994)
  • G.D. Gebreeyessus et al.

    Thermophilic versus mesophilic anaerobic digestion of sewage sludge: a comparative review

    Bioeng.

    (2016)
  • I. Baumann et al.

    Microbial production of short chain fatty acids from lignocellulosic biomass: current processes and market

    Biomed. Res. Int.

    (2016)
  • S. Sanchez et al.

    The fermentation of mixtures of D-glucose and D-xylose by Candida shehatae, Pichia stipitis or Pachysolen tannophilus to produce ethanol

    J. Chem. Technol. Biotechnol.

    (2002)
  • D. Salvachua et al.

    Succinic acid production on xylose-enriched biorefinery streams by Actinobacillus succinogenes in batch fermentation

    Biotechnol. Biofuels.

    (2016)
  • Cited by (16)

    • “Fish gill” -shaped ordered porous PVA@CNNS hybrid hydrogels with fast charge separation and low resistance for effectively photocatalytic synthesis of lactic acid from biomass-derived sugars

      2022, Molecular Catalysis
      Citation Excerpt :

      Consequently, increasing attention has been focused on the lactic acid production via an efficiently, economically, and environmentally friendly strategy. Currently, over 70% of lactic acid in the world is produced by microbial fermentation of sugar-rich biomass (e.g., potatoes, sweet potatoes, or corn) [11,12]. Nevertheless, the aforementioned strategy continues to pose drawbacks such as harsh fermentation conditions and low efficiency [13].

    • Dietary bile acids reduce liver lipid deposition via activating farnesoid X receptor, and improve gut health by regulating gut microbiota in Chinese perch (Siniperca chuatsi)

      2022, Fish and Shellfish Immunology
      Citation Excerpt :

      Dietary BAs supplementation increased the abundance of Lactobacilli in Firmicutes, showing that Lactobacilli might play a key role in the effect of dietary BAs supplementation on the gut health. Lactic acid bacteria are a common acid producing species and produce lactic acid by fermentation [60]. In this study, the increase of Lactobacillus might cause the increase of the lactic acid level and the decrease of pH. Previous studies have shown that acidic gut physiological environment is conducive to the growth of gut villi, which may be the reason for the gut villus length and gut wall high in this study [61,62].

    • Reasonable regulation of carbon/nitride ratio in carbon nitride for efficient photocatalytic reforming of biomass-derived feedstocks to lactic acid

      2021, Applied Catalysis B: Environmental
      Citation Excerpt :

      Over the past decades, lactic acid production has shown great potential in food, beverage, pharmaceutical, cosmetic and chemical industries [14,15]. Currently, commercial lactic acid is produced mainly by fermentation of carbohydrates [16]. However, most of these processes have several drawbacks such as slow reaction kinetics, high nutrient cost, low space-time yield and complicated post-processing [17,18].

    • Combined proteomics and transcriptomics analysis of Lactococcus lactis under different culture conditions

      2021, Journal of Dairy Science
      Citation Excerpt :

      Results showed that fructose metabolism was accelerated, but pyruvate metabolism was inhibited, besides, alpha-acetolactate decarboxylase were significantly downregulated, physiological role of decarboxylation was related to the production of metabolic energy (Romano et al., 2014). The LAB gradually acidify their environment through the conversion of pyruvate to lactate, an essential process to regenerate NAD (+) used during glycolysis and conversion of pyruvate to acetic acid, which in turn benefits catabolic activities of the cell (Fernandez et al., 2008; Veeravalli and Mathews, 2018). This study had reported that Lactococcus may produce less acid and flavor substances when lactose was replaced by fructose.

    • Microwaves in Chemistry Applications: Fundamentals, Methods, and Future Trends

      2021, Microwaves in Chemistry Applications: Fundamentals, Methods and Future Trends
    • Microwave-assisted selective oxidation of sugars to carboxylic acids derivatives in water over zinc-vanadium mixed oxide

      2020, Chemical Engineering Journal
      Citation Excerpt :

      These monosaccharides, which contain two appropriate functional sites (anomeric hydroxyl and carboxylic acid), are suitable starting material for reactions with nucleophilic agents (alcohols, amines, or thiols) [14,16]. Organic acids are generally produced by fermentation of carbohydrate substrates [17,18]. This is presently the most used route [19].

    View all citing articles on Scopus
    View full text