A comprehensive methodology for parameter assessment in solid-state fermentation modelling

Sophorolipids (SLs), currently one of the most promising biosurfactants, are secondary metabolites produced by many non-pathogenic yeasts, among which Candida bombicola ATCC 22214 is the main sophorolipid-producing strain. SLs have gained much attention since they exhibit anti-tumor, anti-bacterial, anti-inflammatory, and other beneficial biological activities. In addition, as biosurfactants, SLs have a low toxicity level and are easily degradable without polluting the environment. However, the production cost of SLs remains high, which hinders the industrialization process of SL production. This paper describes SL structure and the metabolic pathway of SL synthesis firstly. Furthermore, we analyze factors that contribute to the higher production cost of SLs and summarize current research status on the advancement of SL production based on two aspects: (1) the improvement of strain performance and (2) the optimization of fermentation process. Further prospects of lowering the cost of SL production are also discussed in order to achieve larger-scale SL production with a high yield at a low cost. KEY POINTS: • Review of advances in strain performance improvement and fermentation optimization. • High-throughput screening and metabolic engineering for high-performance strains. • Low-cost substrates and semi-continuous strategies for efficient SL production.

We discovered that Candida floricola ZM1502 is capable of selectively producing the promising hydrophilic biosurfactants, acid-form sophorolipids (SLs), from glycerol. However, productivity was very low (approximately 3.5 g L-1) under the initial culture conditions. Here, we describe the design of culture medium for abundant production of acid-form SLs by C. floricola ZM1502 using waste glycerol and hydrophobic substrates in order to develop a method for SL production and disposal of waste glycerol produced by oleo-chemical industries. Urea provided the best nitrogen source for acid-form SL production from glycerol among four nitrogen sources tested [urea, NaNO3, NH4NO3, and (NH4)2SO4]. Among carbon sources we compared, hydrophobic substrates (soybean oil and oleic acid) led to productivities of approximately 20 g L-1, indicating that hydrophobic substrates provided fatty acid moieties for SL production. Addition of olive oil and oleic acid to waste glycerol enhanced acid-form SL production to 42.1 ± 0.9 and 37.5 ± 3.4 g L-1, respectively. To develop a potential industrial process, we explored other suitable hydrophobic substrates for SL production, which were obtained on site from oleo-chemical industries. Alkyl C18 esters (Pastell M-182), along with waste glycerol, increased acid-form SL production to 48.0 ± 3.4 g L-1 over a 7-d period. Furthermore, we demonstrated abundant production of acidic SLs at the mini-jar fermenter scale, obtaining 169 g L-1 over 180 h using a fed-batch cultivation technique. Efficient acid-form SL production by C. floricola could have a great impact on the development of bio-industrial processes using waste glycerol as a substrate.

The production of sophorolipids by Candida bombicola NRRL Y- 17069 grown in a mixture of sunflower oil cake and crude soybean oil as economic substrates with different fermentation techniques was studied. The highest yield (49.5 g·100 g−1 substrates) was obtained from solid state fermentation after employing a new concept for extraction by methanol (E I) followed by ethyl acetate (E II), then partially purified with hexane (E III). The course of time of fermentation was also studied, and E I extracted of the 12th day showed the minimum surface tension (45 mN·m−1) at a critical micelle dilution (CMD) of 10% concentration. The produced sophorolipids were characterized and confirmed by FTIR and 1H NMR spectroscopy. The anticancer activity of the produced compounds was assessed against MCF-7, HepG2, A549, HCT116 cancer cell lines and the results revealed that E III and E IV (a mixture of E I & E III) act as promising anticancer agents in HepG2 and A549 by inhibiting urokinase and histone deacetylase activities.

To facilitate the development of bio-based chemicals from renewable and inexpensive natural resources, we sought to produce biosurfactants using non-edible jatropha oil. Twenty yeasts known to produce biosurfactants were tested in this study, and Stamerella bombicola NBRC 10243 was found to use jatropha oil efficiently to produce sophorolipids (SLs) as a mixture of lactone-form SL (L-SL) and acid-form SL (A-SL). Under culture conditions using rice bran as the source of organic nutrients, the yield of SLs reached 122.6 g/L in 5-L jar fermentors after 9 d in culture. HPLC analysis of the culture medium showed that the levels of phorbol esters (PEs), major toxic components of the oil, decreased markedly with an increase in culture time, suggesting that the yeast degrades PEs. Although the SLs obtained by solvent extraction of the culture medium contained a small amount of PEs, the sodium salt of A-SL (A-SL-Na) obtained by alkaline treatment (5N NaOH, 80°C) showed no PE peaks upon HPLC analysis. A-SL-Na had excellent surface activity with low CMC (9.0×10⁻⁴ M) and γ(CMC) (29.6 mN/m), which are lower than that of sodium dodecyl sulfate (SDS). The solubilizing ability of A-SL-Na toward for octanoic acid ([octanoic acid]/[A-SL-Na]) was found to be 2.0, which is half that of SDS. Our findings should help improve SL production from non-edible feedstock and broaden the use of promising bio-based surfactants.

Chapter 23 - Solid-State Fermentation Strategy for Microbial Metabolites Production: An Overview

Microflora for improving the Auricularia auricula spent mushroom substrate for Protaetia brevitarsis production

In luminescence thermochronometry, the thermal stability of feldspar minerals is conventionally constrained from isothermal decay experiments. However, despite recent refinement of the measurement protocol, measurements take several days and are routinely done for each individual sample. Following that most other thermochronometric methods usually use only a single reference set of thermal kinetic parameters, and that recent studies on direct physical probing of feldspar sample properties have shown that trap depth and band-tail width are broadly similar despite large variations in chemical composition, we sought to optimise luminescence thermochronometry measurements by exploring whether a single set of thermal kinetic parameters can describe luminescence thermal decay in feldspar. We explored the effect of using averaged thermal kinetic parameters rather than sample-specific thermal kinetic parameters to model luminescence signal accumulation under different thermal conditions. A set of K- and Na-feldspar minerals extracted from all over the world were analysed after being measured with a multi-elevated temperature protocol, comprising four different IRSL signals at 50, 100, 150, and 225 °C. Comparisons were done between the thermal kinetic parameters of each IRSL signal depending on different variables such as geographic region, transect, lithology, or mineralogy of the analysed feldspar grains. Even though it is not possible to generalise the thermal kinetic parameters between IRSL signals measured at different temperatures, the variance between the thermal kinetic parameters of different samples measured at the same IRSL temperature is consistent with the uncertainties on the individual parameters (i.e., <2–10%), suggesting that averaged, rather than sample-specific values may be appropriate. We then explored the effect of using these averaged parameters to model luminescence signal accumulation under different synthetic and natural thermal conditions. For our dataset, results show minimal impact on the obtained cooling histories and exhumation rates. We therefore propose the use of averaged rather than sample-specific thermal kinetic parameters for rapid investigation of luminescence thermochronometry samples. Based on careful initial characterisation of a few samples to verify the validity of using averaged thermal kinetic parameters, this would reduce measurement times by ca. 50% (i.e., 3–4 days per sample), allowing higher resolution sampling and measurement.

Techno-economic evaluation of a biorefinery applying food waste for sophorolipid production – A case study for Hong Kong

Solid-state fermentation (SSF) technology has been rapidly developed for the past 10 years as a production platform for secondary metabolites, biofuels, food, and pharmaceuticals. Yet, the main drawback of SSF is the local temperature rise of up to 20 K, which potentially reduces the strain activity and inactivates heat sensible products. Due to the low heat capacity and thermal conductivity of mixtures of air with plant material, in comparison to aqueous suspensions in submerged fermentations, heat from metabolic processes is less efficiently dissipated. The exact knowledge of the metabolic heat generation during SSF processes is crucial to guide strategies against overheating. In this work, a simple method using a cost-efficient multichannel instrument is proposed, which allows the determination of heat generation during SSF processes. This method was successfully tested and validated with Blakeslea trispora producing β-carotene during growth on barley. Additionally, the consequences of the generated metabolic heat during SSF on temperature rise and water evaporation were discussed. Finally, changes in growth and product concentration could also be detected by the heat signal, implying the potential as a timesaving screening method.

Filamentous fungi, especially basidiomycetes, produce a wide range of metabolites, many of which have potential biotechnological and industrial applications. Solid-state fermentation (SSF) is very suitable for the cultivation of basidiomycetes since it mimics the natural habitat of these fungi. Some of the major advantages of SSF are the robustness of the process, the use of low-cost residual materials as substrates, and the reduced usage of water. However, monitoring key variables is difficult, which makes process control a challenge. Specifically, it is very difficult to determine the biomass during SSF process involving basidiomycetes. This is problematic, as the biomass is normally a key variable in mass and energy balance equations. Further, the success of fungal SSF processes is often evaluated, in part, based on the growth of the fungus. Direct determination of the dry weight of biomass is impossible and indirect quantification techniques must be used. Over the years, various determination techniques have been developed for the quantification of fungal biomass in SSF processes. The current review gives an overview of various direct and indirect biomass determination methods, discussing their advantages and disadvantages.

The increasing growth of agroindustrial activity resulting in excessive amounts of agriwaste has led to the accumulation of a large quantity of lignocellulosic residues all over the world, in particular in deforestation initiatives for the removal of invasive trees in South Africa. These lignocellulosic residues are rich in energy resources and consist of a mixture of natural polymers based on lignin, cellulose, and hemicellulose. The use of lignolytic fungi such as mushrooms in solid-state fermentation could sufficiently degrade the indigestible lignocellulosic components and add medicinal and nutritional value to otherwise unusable, high-energy waste material, which in turn could yield a new method of producing energy-rich fodder for ruminant animals. The digestive type of animal for which the potential feed is developed must be identified and considered before deciding on the bioconversion method and process, as the outcomes for obtaining potentially high-quality feeds for nonruminant and ruminant animals are different. The current study presents data on the bioconversion of lignocellulosic substrate using solid-state fermentation with edible and medicinal mushrooms, Ganoderma lucidumand Pleurotus ostreatus, and a possible new species, to increase digestibility and nutritional value to be applied as ruminant animal feed. The solid-state fermentation process was optimized and the resulting product was analyzed for the degradation of the lignocellulosic components. Results indicated that the solid-state fermentation duration and mushroom species were key components in achieving significant degradation. Data obtained after 18 weeks of degradation indicated a significant (p < 0.05) reduction in the acid detergent fiber, acid detergent lignin, and neutral detergent fiber fractions of the biomass, with up to a 20% reduction in indigestible components. This increase in digestibility could contribute to increased energy availability for ruminant animals.

Recent process developments in solid-state fermentation

It is well known that the solid-state fermentation (SSF) is a new technology with which antibiotics, enzymes, fine chemicals can be obtained by new bioprocesses with improved efficiency (Giovannozzi-Sermanni & Porri 1989; Pandey et al., 2000) if compared to the classical submerged fermentations. SSF processes can be defined as “the growth of microorganisms, mainly fungi, on moist solid materials in the absence of free-flowing water”(Cannel & Moo-Young 1980). In the last few decades SSF has grown quickly in interest and importance and has been used for the production of antibiotics, alkaloids, aroma compounds, plant growth factors, enzymes, biofuel, and also for the bioremediation of polluting compounds.

&lt;p&gt;Solid-state fermentation (SSF) process appears to be an efficient way to recover and valorise organic residues from industries and to generate other products of industrial significance, such as enzymes. As the reproducibility of SSF is one of the main drawbacks in the development of this process, a complete study on this point was carried out. Materials with high nitrogen content, soy waste (SW) and hair waste (HW), were treated by SSF to obtain both proteases for different uses and a stabilized material for soil amendment. The bench scale reactors used in the experiments had a 4.5 l volume. Eight and five experiments were performed with the SW and HW, respectively, over a period of 2.5 years with different batches of each material. The highest production of proteases was observed after 3 days and after 14 days of fermentation, for soy and hair wastes, respectively. In the case of hair waste (HW), the overall CV was close to 4.0% for both the maximum protease activity (PA) and the maximum specific protease activity (sPA). In the case of SW, the overall CVs recorded were 43% and 18.6% for the maximum PA and the maximum sPA, respectively. This demonstrates that SSF can be a reproducible process in bench scale reactors. This is the first study and a help to researchers to develop SSF experiments in a consistent and conclusive way.Solid-state fermentation (SSF) process appears to be an efficient way to recover and valorise organic residues from industries and to generate other products of industrial significance, such as enzymes. As the reproducibility of SSF is one of the main drawbacks in the development of this process, a complete study on this point was carried out. Materials with high nitrogen content, soy waste (SW) and hair waste (HW), were treated by SSF to obtain both proteases for different uses and a stabilized material for soil amendment. The bench scale reactors used in the experiments had a 4.5 l volume. Eight and five experiments were performed with the SW and HW, respectively, over a period of 2.5 years with different batches of each material. The highest production of proteases was observed after 3 days and after 14 days of fermentation, for soy and hair wastes, respectively. In the case of hair waste (HW), the overall CV was close to 4.0% for both the maximum protease activity (PA) and the maximum specific protease activity (sPA). In the case of SW, the overall CVs recorded were 43% and 18.6% for the maximum PA and the maximum sPA, respectively. This demonstrates that SSF can be a reproducible process in bench scale reactors. This is the first study and a help to researchers to develop SSF experiments in a consistent and conclusive way.&lt;/p&gt;

Modeling conversion and transport phenomena in solid-state fermentation: A review and perspectives