Sophorolipid Production Research Articles

Sophorolipid biosynthesis and production from diverse hydrophilic and hydrophobic carbon substrates.

Sophorolipids (SLs), mainly synthesized by yeasts, were a sort of biosurfactant with the highest fermentation level at present. In recent years, SLs have drawn extensive attention for their excellent physiochemical properties and physiological activities. Besides, issues such as economics, sustainability, and use of renewable resources also stimulate the shift from chemical surfactants towards green or microbial-derived biosurfactants. SLs' large-scale production and application were restricted by the relatively high production costs. Currently, waste streams from agriculture, food and oil refining industries, etc., have been exploited as low-cost renewable substrates for SL production. Advanced cultivation method, uncommonly used substrates, and new genetically modified SL-producing mutants were also designed and applied to improve the productivity or the special properties of SLs. In this review, a systematic and detailed description of primary and secondary metabolism pathways involved in SL biosynthesis was summarized firstly. Furthermore, based on the pathways of SL biosynthesis from different carbon substrates, we reviewed the current knowledge and advances in the exploration of cost-effective and infrequently used hydrophilic and hydrophobic substrates for large or specialized SL production.

Efficient in-situ separation design for long-term sophorolipids fermentation with high productivity

Until now, the production of microbial biosurfactants has been limited to the use of first-generation (food) substrates. Moreover, the results on sophorolipids yield and productivity reported by state-of-the-art fed-batch processes for sophorolipids production remain unsatisfactory and lead to a significantly high cost per kg of the product. With an aim to address these problems, this study demonstrates a long-term and high-efficiency sophorolipids production method using a semi-continuous integrated production-separation system which utilizes food waste as a substrate. An average volumetric productivity of 2.43 g L −1 h −1 and the overall sophorolipids yield on the substrate (i.e. the combination of glucose and oleic acid as carbon source) of 0.73 g g −1 was achieved within 240 h by fed-batch and separation fermentation. Moreover, the potential of sustaining high production efficiency during long-term fermentation times (480 h) was investigated. No reduction in process efficiency was observed, i.e. average volumetric productivity and an overall sophorolipids yield of 2.39 g L −1 h −1 and 0.73 g g −1 were obtained, respectively. Notably, a very high biomass concentration of 117.2 g L −1 was observed in fed-batch fermentation and separation, which is the highest cell density ever reported for Starmerella bombicola . The separation efficiency was also calculated to evaluate the in-situ separation performance of the developed process quantitatively. In 480-h fermentation, the average separation efficiency of 74.3% and overall separation efficiency of 93.0% was achieved after six separation cycles, suggesting that both separation design and process control were successful in laboratory-scale fermentation. These findings demonstrate the potential of the developed sophorolipids fermentation-separation system for application at industrial scale. • Food waste was used for sophorolipids (SL) fermentation. • Combination of fed-batch with an in-situ separation strategy was developed. • An average productivity of 2.4 g L −1 h −1 was obtained in 480 h fermentation. • A yield of 0.73 g g −1 of carbon added was achieved. • A cell density of 117.2 g L −1 and separation efficiency of 93.0% were achieved.

Production of long-chain hydroxy fatty acids by Starmerella bombicola.

To decrease our dependency for the diminishing source of fossils resources, bio-based alternatives are being explored for the synthesis of commodity and high-value molecules. One example in this ecological initiative is the microbial production of the biosurfactant sophorolipids by the yeast Starmerella bombicola. Sophorolipids are surface-active molecules mainly used as household and laundry detergents. Because S. bombicola is able to produce high titers of sophorolipids, the yeast is also used to increase the portfolio of lipophilic compounds through strain engineering. Here, the one-step microbial production of hydroxy fatty acids by S. bombicola was accomplished by the selective blockage of three catabolic pathways through metabolic engineering. Successful production of 17.39 g/l (ω-1) linked hydroxy fatty acids was obtained by the successive blockage of the sophorolipid biosynthesis, the β-oxidation and the ω-oxidation pathways. Minor contamination of dicarboxylic acids and fatty aldehydes were successfully removed using flash chromatography. This way, S. bombicola was further expanded into a flexible production platform of economical relevant compounds in the chemical, food and cosmetic industries.

Production and characterization of sophorolipids by Candida bombicola using catfish fat

Catfish fat is a high nutrition by-product of seafood processing industry. In this work, catfish fat has been used as a sustainable and economical raw material for sophorolipids production by Candida bombicola. Sophorolipids yield was maximum as 21.8g/L after 7 days of fermentation at 25oC, pH 6, 180 rpm. The obtained sophorolipids was to contained the main component as lactonic sophorolipids, which has been confirmed by Thin layer chromatography (TLC). Sophorolipids also exhibited the ability to resistant Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa and showed the ability of free radical scavenging the dose dependent manner with IC50 was 4.45 mg/ml. These results suggested that sophorolipids could be used in health care products and cosmetic. Catfish fat could be used as the low cost hydrophobic carbon source to replace fatty acid for sophorolipids production.

Production and antimicrobial activity of sophorolipid against Clostridium perfringens and Campylobacter jejuni and their additive interaction with lactic acid

Abstract The poultry industry experiences enormous economic losses due to microbial contamination, leading to searches for natural antimicrobials. Sophorolipid is a glycolipid produced by Starmerella bombicola, that acts as an emulsifier and antimicrobial agent. The aim of this research was to produce, characterize and apply sophorolipid from S. bombicola against the chicken pathogens Clostridium perfringens and Campylobacter jejuni, which are responsible for diseases that cause great damage to the poultry industry. The application of sophorolipid against these microorganisms has not yet been reported and to aggregate value to the product, sophorolipid was associated with lactic acid, a nontoxic molecule already utilized in the food industry. Sophorolipid production reached 69.83 g.L−1 with a productivity of 0.24 g.L−1.h−1 and a yield of 46.41% at 288 h in a bioreactor. The structural characterization analyses confirmed the predominance of the lactonic form. Sophorolipid demonstrated antimicrobial activity against the tested bacteria and the combined treatment of sophorolipid and lactic acid represented an additive interaction. Therefore, this combination could be an alternative to use as a new natural sanitizer in the poultry industry, reducing the microbial contamination of these main pathogens.

Use of wastes for sophorolipids production as a transition to circular economy: state of the art and perspectives

Chemical processes and petroleum-based chemicals are being substituted by biological processes and bioproducts. Surfactants and biosurfactants are an example of this trend. Among the biosurfactants, sophorolipids (SLs) have excellent surface and interfacial tension properties, which make them ideal to be used in a wide variety of applications. SLs are produced at full scale through submerged fermentation of pure substrates (glucose and oleic acid). However, research trends suggest that there is a lot of interest to produce SLs from waste effluents and other low-cost substrates, both in submerged and solid-state fermentation processes. This study reviews the current research in the production of SLs via fermentation processes, focusing on those using wastes, by-products, or low-cost substrates (liquids or solids). It details the substrates, process variables, microorganisms, and use of supplementary media for batch, fed-batch, and continuous submerged or solid-state fermentation processes. Sophorolipids production based on industrial by-products and waste effluents presents huge potential for its application at an industrial scale in a more economical and environmentally friendly process, boosting the necessary change to circular economy.

Efficient sophorolipids production using food waste

Recent sustainable development goals of food security, environmental protection, material and energy efficiency are the key drivers of the valorization of food waste. In the present work, the production of biosurfactant sophorolipids from several (food) waste streams was investigated, using the non-pathogenic yeast Starmerella bombicola. From a preliminary screening, restaurant food waste emerged as the most suitable feedstock compared to bakery waste, textile waste, used corn oil, animal fat and lipid fraction of hydrolyzed food waste. Restaurant food waste was subsequently used for sophorolipids production in a laboratory-scale bioreactor. Food waste obtained from a local restaurant was subjected to enzymatic hydrolysis for 16 h, yielding a hydrolysate containing about 100 g/L glucose and 2.4 g/L free amino nitrogen. High SL process efficiency was achieved by fed-batch fermentation using the restaurant food waste hydrolysate as the complete batch medium, i.e. without any supplementation of additional medium components such as vitamins, salts, nitrogen or phosphate. Controlled feeding of glucose and oleic acid to the culture was performed after the batch phase. A sophorolipids titer of 115.2 g/L was obtained in a fermentation time of 92 h resulting in an overall volumetric productivity of 1.25 g/L.h. These results achieved for sophorolipids productivity using hydrolyzed food waste are in the same order of magnitude as the reported values using traditional (complex) fermentation media. This indicates the suitability of the developed process using food waste for the advancement of waste-based bio-processes for the production of sophorolipids.

Rational high-throughput screening system for high sophorolipids production in Candida bombicola by co-utilizing glycerol and glucose capacity

BackgroundA rational and high-performance high-throughput screening system effectively improves the efficiency of strain screening.ResultsA rapid and efficient method was developed to detect carbon sources. And then by using glucose analog as screening pressure and two indexes for sophorolipids (SLs) production and glycerol consumption, a mutant of Candida bombicola L1.3 was successfully selected among 3000 mutants. It produces high SLs titer and co-utilizes glucose and glycerol simultaneously.ConclusionsCompared to the wild-type strain, L1.3 exhibited 15.06% higher SLs titer and 35.69% higher glycerol consumption capacity in a 5-L bioreactor. We believe that L1.3 can potentially be used for the efficient industrial production of SLs with simple downstream processing for separating SLs and glycerol.

Miniaturization of Starmerella bombicola fermentation for evaluation and increasing (novel) glycolipid production.

Both strain engineering and process optimization are intensively studied in microbial biosurfactant literature. However, screening of multiple strains and/or medium components in parallel is a very labor-intensive and timely process, considering the only applied technique nowadays is evaluation through shake flask and/or bioreactor experiments. Therefore, in this work, the development, optimization, and application of a more throughput technique-based on 24-deep well plates-are described for a new Starmerella bombicola strain producing bolaform sophorolipids. To develop an optimal setup, the influence of plate position and culture volume and the type of sandwich cover was investigated. Optimal parameters, which did not result in significant differences compared with shake flask experiments concerning growth, glucose consumption, and production of novel sophorolipids, were defined and validated. Next, the new method was applied to evaluate the influence of the use of alternative (commercial) nitrogen sources in comparison with the yeast extract currently applied in the production medium, aiming to increase production efficiency. Self-made yeast extracts from S. bombicola cells were also included to evaluate possible recycling of cells after fermentation. In conclusion, the designed method enabled the efficient and successful comparison of ten different nitrogen sources in varying concentrations (1, 4, and 10g/L) on bola sophorolipid production, which can now also be performed for other parameters important for growth and/or glycolipid production.

Metabolic profiling and flux distributions reveal a key role of acetyl-CoA in sophorolipid synthesis by Candida bombicola

Sophorolipids are important biosurfactants produced by the wild-type strain of Candida bombicola ATCC 22214, with antimicrobial, antiviral, and anticancer activities. Our previous study showed that a mutant strain lacking multifunctional enzyme type 2 displayed extremely low sophorolipid production capacity compared to the wild-type strain. In the present study, we aimed to reveal the possible reasons for this deficiency through a series of analytical methods. First, comparative metabolic profiling analysis between the wild-type and mutant strains was performed. Multivariate statistical analysis, including principal component analysis and orthogonal partial least squares discriminant analysis, showed that nine potential metabolites were closely associated with the metabolic differences between strains. Second, metabolic flux analysis revealed that an inadequate supply of intracellular acetyl-CoA in the mutant strain could significantly affect the biosynthesis of sophorolipids. In addition, supplementation of citric acid recovered the sophorolipid production in the mutant strain by approximately 83%, suggesting that citrate metabolism plays an important role in sophorolipid biosynthesis. These findings provide novel theoretical insights for further improving the production of sophorolipids in C. bombicola.

Evaluation of sunflower oil refinery waste as feedstock for production of sophorolipid

Abstract Sophorolipid, a glycolipid biosurfactant, was produced by submerged fermentation using Starmerella bombicola. Sunflower acid oil, waste from a vegetable oil refinery, has been studied as a newer feedstock with glucose for production of sophorolipid. Maximum sophorolipid production was found to be 41.6 g/L at shake flask and 51.5 g/L at fermenter level after 8 days of fermentation. Acidic and lactonic forms of sophorolipid were synthesized using acid oil, with the later being abundant. Structure of purified sophorolipid confirmed with the help of HPLC, LCMS and NMR analysis. The final product was characterized for its surface activity and exhibited equivalent performance when compared with the fresh substrate (oleic acid and sunflower oil) based sophorolipid. The sunflower acid oil based sophorolipid reduced the surface tension of water to 35.5 mN/m and interfacial tension (water/n-heptane) to 0.923 mN/m whereas it reduced to 30.69 mN/m and 5.8 mN/m respectively for synthetic surfactant polysorbate 20. The sunflower acid oil based sophorolipid was also shown to have better emulsification as well as wetting property but less foamability comparatively.

Investigation of fermentation conditions for Candida bombicola ACTT22214 from molasses and soybean oil for sophorolipid production

Sophorolipid (SL) is a biosurfactant belonging to the glycolipids group and was produced by harmless Candida bombicola ACTT22214 and has been widely used in many fields in our life. In order to search for appropriate condition for C. bombicola fermentation producing SL with high efficiency, this study focused on the investigation of the soy oil concentration, molasses concentration, fermentation time, pH and temperature. SL products were tested for antimicrobial activity, antioxidant, emulsifier, foaming ability. The highest content of SL was 43.27 ± 0.30 g/L under conditions of: soybean oil content 5%, molasses content 150 g/L, fermentation time 7 days, pH = 5, 28oC fermentation temperature. The antibacterial activity of SL was good: the highest resistance to Candida albicans (16.33 ± 1.15 mm), good resistance to Bacillus spizizenii (13.67 ± 0.58 mm), resistance to Staphylococcus aureus (12.67 ± 1.15 mm), relatively weak resistance to Pseudomonas aeruginosa (11.33 ± 0.58 mm) and Escherichia coli (9.67 ± 0.58 mm). The antioxidant capacity of SL was quite high with an IC50 value of 6.024 mg/mL. The emulsifying capacity of SL was equivalent to the emulsification of the tween 20 at a concentration of 5 - 10 mg/mL. SL had the ability to foam evenly from concentrations of 5 to 20 mg/mL but not higher than the corresponding concentrations of tween 20, SL was smooth, even, stable longer than tween 20

Real-time dynamic analysis with low-field nuclear magnetic resonance of residual oil and sophorolipids concentrations in the fermentation process of Starmerella bombicola

The yeast Starmerella bombicola is known to produce sophorolipids (SLs) by fermentation method. In the fermentative production of SLs, at least three phases i.e. hydrophilic phase of glucose, ions and acidic-form SLs; hydrophobic phase of oil and lactone-form SLs; solid phase of cell biomass and SLs crystals are present in the broth. Therefore, a rapid and real-time detection of residual oil and SLs can provide valuable information in the regulation and optimization of the process to better produce SLs effectively. Looking into the importance, in this study, a rapid, accurate and precise method to quantify the concentrations of oil and SLs using low-field nuclear magnetic resonance (LF-NMR) was developed. Compared to the traditional weighing method, all the parameters for the evaluation of accuracy, precision and relative recovery presented better performances. Moreover, the proposed LF-NMR method could completely avoid using organic solvents that are commonly employed in the weighing method, besides shortening the pre-treatment and detection times from 24 h to just <20 min. Finally, LF-NMR method has been successfully used to monitor residual oil and SLs during the fermentation of Starmerella bombicola to obtain SLs and it has been observed that the production of SLs would be effectively improved by appropriately controlling the concentration of oil in terms of titer, productivity and yield.

Efficient sophorolipids production via a novel in situ separation technology by Starmerella bombicola

Abstract Sophorolipids are environmentally friendly biosurfactants and hence their demand has greatly increased during last decade, especially in cleaning and cosmetic applications. However, their higher production cost becomes a major barrier for wider applications compared to their counterparts i.e. petrol-derived surfactants. In this investigation, a novel and efficient technology for the in situ separation of sophorolipids has been proposed, including froth flotation followed by recovery of entrained cells through washing. First, independent cascade separators having a pre-separator and a froth separator were designed. 75% of sophorolipids were separated in the froth separator within 12 min after adjusting oil to sophorolipids ratio (0.08–0.012) in the bioreactor. Washing was then applied to recover over 90% of entrained cells in the froth. Finally, using 5 L bioreactor three rounds of in situ separation were carried out to achieve a final production of 342 g/L (normalized to initial volume) for 221 h, an improvement of 41% compared to fed-batch fermentation. Besides, the overall productivity reached 1.55 g/L/h. Overall, this investigation provides an in situ separation technology with controllable frothing and is applicable to an industrial scale. Also it maximizes the production potential of strain along with lowering the production cost of sophorolipids significantly.

Starmerella bombicola: recent advances on sophorolipid production and prospects of waste stream utilization

AbstractSophorolipids are among the most extensively studied microbial biosurfactants. Starmerella bombicola is the most productive strain known for sophorolipid production, with volumetric productivity of up to 3.7 g L−1 h−1. This review focuses on the two most important aspects that have an influence on sophorolipid commercialization. Firstly, the metabolic engineering achievements of S. bombicola in the last decade are summarized. Secondly, three improvements of the bioprocess are described, including alternative feedstock, fermentation strategy and specially designed bioreactor. Discussion is made on the waste sources that have been used as feedstock for sophorolipid production, and the review also emphasizes the potential of food waste as nutrient source. Fermentation strategies that correlate with the specially designed bioreactors for commercialization are also discussed in detail. © 2018 Society of Chemical Industry

Valorisation of digestate from biowaste through solid-state fermentation to obtain value added bioproducts: A first approach

Digestate from biowaste was assessed as a potential source of bioproducts of commercial and industrial interest through solid-state fermentation. The targeted bioproducts were hydrolytic enzymes (cellulases and proteases from autochthonous microbiome), biosurfactants (sophorolipids produced from Starmella bombicola) and biopesticides (produced from Bacillus thuringiensis). Low cellulase production was observed within the range of 0.5–1.5 FPU g−1 DM while protease production showed two discrete peaks of 66 ± 8 and 65 ± 3 U g−1 DM at 3.5 and 48 h, respectively. Low sophorolipids production was also obtained, with a maximum yield of 0.02 g g−1 DM using hygienised digestate supplemented with external sugar and fat sources. Biopesticides produced by B. thuringiensis were successfully at 72 h of operation, reaching a maximum spore production of 8.15 ± 0.04 (107) CFU g−1 DM and 2.85 ± 0.22 (107) CFU g−1 DM using sterile and hygienised digestate, respectively. These biopesticides could contribute to the substitution of chemically produced pesticides, moving towards a sustainable digestate management in a circular economy scheme.

Process optimization concepts for cost‐efficient production of sophorolipids

Chemie Ingenieur TechnikVolume 90, Issue 9 p. 1273-1273 PosterFree Access Process optimization concepts for cost-efficient production of sophorolipids C. Zerhusen, C. Zerhusen TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanySearch for more papers by this authorJ. Brotsmann, J. Brotsmann TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanySearch for more papers by this authorS. Barbe, S. Barbe TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanySearch for more papers by this authorU. Schörken, Corresponding Author U. Schörken ulrich.schoerken@th-koeln.de TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanyCorrespondence: U. Schörken (ulrich.schoerken@th-koeln.de), TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanySearch for more papers by this author C. Zerhusen, C. Zerhusen TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanySearch for more papers by this authorJ. Brotsmann, J. Brotsmann TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanySearch for more papers by this authorS. Barbe, S. Barbe TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanySearch for more papers by this authorU. Schörken, Corresponding Author U. Schörken ulrich.schoerken@th-koeln.de TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanyCorrespondence: U. Schörken (ulrich.schoerken@th-koeln.de), TH Köln, Faculty of Applied Natural Sciences, Kaiser-Wilhelm-Allee, 51368 Leverkusen, GermanySearch for more papers by this author First published: 24 August 2018 https://doi.org/10.1002/cite.201855310Citations: 1AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. References 1 R. Ashby et al., J. Surfact. Deterg. 2013, 16, 683– 691. 2 S. Roelants et al., Biotechnol. Bioeng. 2016, 113, 550– 559. Citing Literature Volume90, Issue9Special Issue: ProcessNet-Jahrestagung und 33. DECHEMA-Jahrestagung der Biotechnologen 2018September 2018Pages 1273-1273 ReferencesRelatedInformation

Gate‐to‐gate life cycle assessment of biosurfactants and bioplasticizers production via biotechnological exploitation of fats and waste oils

AbstractBACKGROUNDThis study investigated the biotransformation of fats and waste oils to glycolipid biosurfactants and bioplasticizers. The ecological performance and environmental impacts of the bioprocesses were evaluated aiming to assess their present environmental status and thus suggest future improvements using life cycle assessment (LCA) methodology.RESULTSBiosurfactants, namely rhamnolipids and sophorolipids, were obtained via fermentation. Bioplasticizers, fatty acid ethyl esters (FAEE) and monoglycerides (MAG), were developed via enzymatic catalysis with selected enzymes in mesophilic temperatures via ethanolysis and glycerolysis, respectively. The study revealed that air emissions, electricity and thermal energy requirements are the key contributors to the potential environmental impacts in the life cycle impact assessment (LCIA). More specifically, rhamnolipids production has less energetic needs compared with sophorolipids manufacturing, resulting in lower environmental impacts. The increased thermal requirements of the MAG production phase is the main contributor to their negative environmental performance, with the overall energy consumption for MAG production being 3‐fold higher than for the FAEE formation phase.CONCLUSIONSThe assessment identified that among the biosurfactant production processes, sophorolipids production resulted in 22.7% higher environmental impact compared with rhamnolipids. Similarly, FAEE production can be classified as a more environment‐ friendly process compared with MAG, resulting in 67% lower environmental impact based on the environmental indicators assessed. © 2018 Society of Chemical Industry

From lab to market: An integrated bioprocess design approach for new-to-nature biosurfactants produced by Starmerella bombicola.

Glycolipid microbial biosurfactants, such as sophorolipids (SLs), generate high industrial interest as 100% biobased alternatives for traditional surfactants. A well-known success story is the efficient SL producer Starmerella bombicola, which reaches titers well above 200 g/L. Recent engineering attempts have enabled the production of completely new types of molecules by S. bombicola, e.g. the bolaform SLs. Scale-up of bolaform SL production was performed at 150 L scale. The purified product was evaluated in detergent applications, as classic SLs are mostly applied in eco-friendly detergents. In this paper, we show that they can be used as green and non-irritant surfactants in for example (automatic) dishwashing applications. However, due to the presence of an ester function in the biosurfactant molecule a limited chemical stability at higher pH values (>6.5) was noticed, (therefore called 'non-symmetrical' (nsBola)) which, is a major drawback that will most likely inhibit market introduction. An integrated bioprocess design (IBPD) strategy was thus applied to resolve this issue. The strategy was to replace the fed fatty acids with fatty alcohols, to generate so-called "symmetrical bolaform (sBola) sophorosides (SSs)," containing two instead of one glycosidic bond. Next to a change in feeding strategy, the blocking of the fatty alcohols from metabolizing/oxidizing through the suggested ω-oxidation pathway was necessary. For the latter, two putative fatty alcohol oxidase genes (fao1 and fao2) were identified in the S. bombicola genome and deleted in the bolaform SL producing strain (ΔatΔsble). Shake flask experiments for these new strains (ΔatΔsbleΔfao1 and ΔatΔsbleΔfao2) were performed to evaluate if the fed fatty alcohols were directly implemented into the SL biosynthesis pathway. Indeed, sBola sophorosides (SSs) production up to 20 g/L was observed for the ΔatΔsbleΔfao1 strain. Unexpectedly, the ΔatΔsbleΔfao2 strain only produced minor amounts of sBola sophorosides (SSs), and mainly nsBola SLs (alike the parental ΔatΔsble strain). The sBola sophorosides (SSs) were purified and their symmetrical structure was confirmed by NMR. They were found to be significantly more stable at higher pH, opening up the application potential of the biosurfactant by enhancing its stability properties.

Efficient Production of Acid-Form Sophorolipids from Waste Glycerol and Fatty Acid Methyl Esters by &lt;i&gt;Candida floricola&lt;/i&gt;

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.