Sophorolipid Production Research Articles

Production and characterization of sophorolipids from stearic acid by solid-state fermentation, a cleaner alternative to chemical surfactants

In this manuscript, we approach the production of biosurfactants as a cleaner alternative to the chemically-produced surfactants currently used in a wide range of industries. Sophorolipids are microbially produced biosurfactants of the glycolipid type that have entered the market in select applications such as detergent or cosmetic formulation ingredients. This study focuses on sophorolipid production by the yeast Starmerella bombicola from stearic acid (C18:0), a low-cost carbon source that is difficult to work with in submerged fermentation since it remains a solid due to its high melting temperature. Consequently, optimizations of solid-state fermentation inoculated with Starmerella bombicola were studied for conversions of stearic acid and molasses to sophorolipids. Polyurethane foam functioned as the inert support. The effect of polyurethane foam density and water holding capacity was assessed and the process was optimized in terms of substrate and inoculum ratio. The best conditions were: foam with a density of 32 kg m−3 at 75% water holding capacity, 1.17:1 molasses/stearic acid (w/w) and 5% (v/w) inoculum, to obtain a yield of 0.211 g sophorolipids per g of substrates. Mass spectrometry revealed that the sophorolipids produced herein had high concentrations of diacetylated acidic and lactonic C18:0 forms. The results of interfacial properties studies revealed that C18:0 sophorolipids had promising surface tension lowering capacity and emulsification behavior. This study describes a new strategy to produce biosurfactants using low environmental impact technologies as an alternative to traditional ways to produce chemical detergents.

Anti-biofilm activity of a sophorolipid-amphotericin B niosomal formulation against Candida albicans

Sophorolipids (SLs) have gained interest in the pharmaceutical industries due to their anti-microbial, anti-adhesive and anti-biofilm properties. In the present study, the production of SL was increased by using low-cost media components. The potential of a SL-based niosomal formulation of amphotericin B (AmB) was determined against biofilm of the opportunistic fungal pathogen Candida albicans. In-house prepared SL-AmB niosomes were characterized by different microscopic techniques. The mean entrapment efficiency of AmB within SL-AmB niosome was 63.20% ± 3.86. The cytotoxicity of SL-AmB on mature C. albicans biofilm was compared with an expensive, marketed drug, viz. phosome (a liposomal formulation of AmB). Fewer hyphae were observed in C. albicans biofilm treated with SL-AmB niosome whereas more budding cells were found in phosome treated biofilm. The present study has established the affordable production of SL and the suitability of this approach for delivery of poorly soluble drugs such as AmB against candidiasis infections.

Sophorolipid production by yeasts: a critical review of the literature and suggestions for future research.

Sophorolipids are well-known biosurfactants produced by yeasts, having potential applications ranging from nanomaterials, medicine, and cosmetics to large-volume applications such as cleaning and soil remediation. Because of their environmentally friendly nature, they attained much interest during the past decades as a sustainable and ecological alternative to petroleum-derived surfactants. Stronger yet, research activities and scientific publications on the topic are ever increasing. However, often these studies lack proper producer strain identification and detailed structural product characterization. Flaws regarding strain identity can have huge consequences when moving to valorization and moreover tend to persist quite long in scientific literature. Furthermore, too often sophorolipid production is proposed where other biosurfactant structures cannot be ruled out based on the chemical analysis. Finally, absolute quantitative yield determination frequently occurs with variable product purity and without proper calibration standards. This review aims to highlight and discuss these discrepancies and proposes some guidelines for good practice in future sophorolipid research.

Production of sophorolipids by Starmerella bombicola yeast using new hydrophobic substrates

Sophorolipids (SLs) are surface active compounds that have excellent surface-lowering properties. Typically, SLs are produced using different hydrophobic substrates, such as alkanes, vegetable oils, and industrial effluents. The properties of the SLs are highly dependent on the hydrophobic substrate used for their production. The aim of the present study is to investigate the properties of SLs produced using three new hydrophobic substrates: Tapis oil, Melita oil, and Ratawi oil. The structures of the SLs were determined by Fourier transform infrared spectroscopy. The SLs yields using Tapis, Melita, and Ratawi oil were 26, 21, and 19gL−1, respectively. The SLs produced using Tapis, Melita, and Ratawi oil reduced the surface tension of pure water to 36.38, 37.84, and 38.92 mN/m, respectively, corresponding to critical micelle concentrations (CMCs) of 54.39, 55.68, and 58.34mgL−1. These values are comparable with SLs produced using palm oil (surface tension 35.38 mN/m and CMC 48.76mgL−1). The SLs produced using Tapis oil show a maximum emulsification activity of 71.2%. The produced SLs are active over the pH range 2–10 and for salinity up to 20% (w/v) NaCl. Thus, they show potential for various applications, including microbial enhanced oil recovery and oil spill remediation.

Sweet sorghum bagasse and corn stover serving as substrates for producing sophorolipids.

To make the process of producing sophorolipids by Candida bombicola truly sustainable, we investigated production of these biosurfactants on biomass hydrolysates. This study revealed: (1) yield of sophorolipds on bagasse hydrolysate decreased from 0.56 to 0.54 and to 0.37g/g carbon source when yellow grease was dosed at 10, 40 and 60g/L, respectively. In the same order, concentration of sophorolipids was 35.9, 41.9, and 39.3g/L; (2) under similar conditions, sophorolipid yield was 0.12, 0.05 and 0.04g/g carbon source when corn stover hydrolysate was mixed with soybean oil at 10, 20 and 40g/L. Sophorolipid concentration was 11.6, 4.9, and 3.9g/L for the three oil doses from low to high; and (3) when corn stover hydrolysate and yellow grease served as the substrates for cultivating the yeast in a fermentor, sophorolipid concentration reached 52.1g/L. Upon further optimization, sophorolipids production from ligocellulose will be indeed sustainable.

Selective Production of Acid-form Sophorolipids from Glycerol by Candida floricola.

Biosurfactants (BSs) are produced in abundance from various feedstocks by diverse microorganisms, and are used in various applications. In this paper, we describe a new yeast isolate that produces glycolipid-BSs from glycerol, with the aim of enhancing the utilization of the surplus glycerol produced by the oleo-chemical industry. As a result of the screening, strain ZM1502 was obtained as a potential producer of BS from glycerol. Based on TLC analysis, the strain produced glycolipid BSs. According to structural analyses (NMR, MALDI-TOF MS, and GC-MS), the main component of the glycolipids was 6',6"-di-O-acetylated acid-form sophorolipid (SL). Interestingly, the strain produced only acid-form SL, without lactone-form SLs, although the conventional SL-producing yeast, Starmerella bombicola, produces lactone-form SLs with small amounts of the acid-form. Based on taxonomy, the strain was identified as Candida floricola. It produced 3.5 g L-1 of acid-form SLs in 20% (w/v) glycerol. In addition, C. floricola CBS7290 and NBRC10700T also produced only acid-form SLs from glycerol. These results suggest that C. floricola would enhance the utilization of waste glycerol as a fermentation feedstock and facilitate a broad range of applications for SLs.

Production and characterization of sophorolipids produced by Candida bombicola from coconut oil

The biosurfactants from microbial origin increasingly gained interests because of their application in many field and excellent properties compared to surfactants from chemical origin, such as the higher biodegradability, lower toxicity and environmentally friendly. Sophorolipids, biosurfactants of glycolipid groups are produced through the fermentation by nonpathogenic yeasts such as Candida bombicola. In this study, we investigated the production, surveyed properties of sophorolipids through fermentation by C. bombicola from coconut oil. The results showed that the yield of sophorolipid obtained after 7 days of culture was 14.6 g/L, the surface tension was 40 mN/m. The obtained sophorolipid showed ability to be resistant to some bacteria such as E. coli, B. subtilis, P. aeruginosa, and S. aureus. Through DPPH experiment, sophorolipids showed the scavenging acitivity with IC50 = 1.4063 mg/mL. These results showed that sophorolipids could be applied in cosmetics.

Integrated sophorolipid production and gravity separation

Abstract A novel method for the integrated gravity separation of sophorolipid from a fermentation broth has been developed, enabling removal of a sophorolipid phase of either higher or lower density than the bulk fermentation broth, while cells and other media components are recirculated and returned to the bioreactor. The capability of the separation system to recover an enriched sophorolipid product phase was demonstrated on three sophorolipid producing fed batch fermentations using Candida bombicola, giving an 11% reduction in fermenter volume required whilst maintaining sophorolipid production. Sophorolipid recoveries of up to 86% (280 g) of the total produced over a whole fermentation were achieved at an enrichment of up to 9. Furthermore, the broth viscosity reduction achieved by removal of the sophorolipid phase enabled a 34% reduction in mixing power to maintain the same dissolved oxygen level by the end of the fermentation, with a 9% average reduction over the course of the fermentation. Fermentation duration could be extended to 1023 h, allowing production of 623 g sophorolipid from 1 l initial batch volume. These benefits could lead to a substantial decrease in the cost of sophorolipid production, making high volume applications such as enhanced oil recovery economically feasible.

Computational fluid dynamics simulation of a novel bioreactor for sophorolipid production

This paper describes three-dimensional computational fluid dynamics (CFD) simulations of gas–liquid flow in a novel laboratory-scale bioreactor contained dual ventilation-pipe and double sieve-plate bioreactor (DVDSB) used for sophorolipid (SL) production. To evaluate the role of hydrodynamics in reactor design, the comparisons between conventional fed-batch fermenter and DVDSB on the hydrodynamic behavior are predicted by the CFD methods. Important hydrodynamic parameters of the gas–liquid two-phase system such as the liquid phase velocity field, turbulent kinetic energy and volume-averaged overall and time-averaged local gas holdups were simulated and analyzed in detail. The numerical results were also validated by experimental measurements of overall gas holdups. The yield of sophorolipids was significantly improved to 484g·L−1 with a 320h fermentation period in the new reactor.

Isolation of yeast candidates for efficient sophorolipids production: their production potentials associate to their lineage.

Eleven biosurfactant-producing strains were newly isolated from environmental samples using a drop-collapse assay and thin-layer chromatography (TLC). According to the TLC analysis, the separation patterns of the glycolipid spots of nine dominant strains corresponded to that of the sophorolipids produced by a Starmerella bombicola type strain. The retention factor values of the spot patterns of two strains were less than those of the others. Two representative major products were purified, and their molecular structures were determined. The major products were identified as diacetylated lactonic and acidic sophorolipids. The fatty acid moieties of both compounds were estimated to be 17-hydroxymethyl hexadecenoic acid. The amounts of glycolipids ranged from 5.0 to 22.9g/L after 4 d of cultivation. According to a phylogenetic analysis, the strains were identified as Starmerella bombicola and Candida floricola.

Sophorolipids production from rice straw via SO3 micro-thermal explosion by Wickerhamiella domercqiae var. sophorolipid CGMCC 1576.

A novel lignocellulose material, holocellulose from rice straw via the pretreatment of SO3 micro-thermal explosion, was developed to produce sophorolipids (SLs) with Wickerhamiella domercqiae var. sophorolipid CGMCC 1576. The influence factors of inoculum dose, yeast extract concentration and pH regulators (chemical regents used for adjusting/influencing pH) was investigated and discussed. Results showed that W. domercqiae can grow in the rice straw holocellulose hydrolysate, and acquire relative high SL yield of 53.70 ± 2.61 g/L in shake flask culture. Inoculum dose, yeast extract concentration and pH regulator made obvious influence on fermentation parameters, especially on final broth pH and SLs production. Furthermore, there is a strong negative linear correlation existing between final broth pH and lactonic SL or ratio of lac SL/tot SL. Additionally, comparison between SL production and non-glucose carbon sources, culture methods, microbes in previous reports was carried out. These results will be benefit for acquiring SL mixture with suitable lac SL/tot SL ratio for specific purpose and scope economically.

Production of sophorolipids from winterization oil cake by solid-state fermentation: Optimization, monitoring and effect of mixing

Sophorolipids (SLs) are a group of extracellular biosurfactants produced by the yeast Starmerella bombicola. The present study explored the use of winterization oil cake (WOC), a residual oil cake that comes from the oil refining industry, as a substrate for the production of SLs by solid-state fermentation (SSF). Sugar beet molasses (MOL) was used as a co-substrate and S. bombicola ATCC 22214 as the inoculum. Fermentation was performed on the 100-g scale and was optimized in terms of the ratio of substrates and the aeration rate using response surface methodology. The optimized SSF process (1:4 MOL:WOC mass ratio and 0.30Lkg−1min−1 aeration rate), carried out under static conditions, was monitored for 10days with a maximum SL yield of 0.179g per g DM (dry matter). The effect of intermittent mixing on the process was also investigated. Mixing caused a 31% increase in SL production, with a total yield of 0.235g per g DM. The Oxygen Uptake Rate (OUR) and the Cumulative Oxygen Consumption (COC) were used to monitor the biological activity of the fermentation processes. There were significant correlations between the SL yield and the oxygen and fats consumed. The SLs were characterized by FTIR and 1H NMR analysis.

Sophorolipid Amine Oxide Production by a Combination of Fermentation Scale-up and Chemical Modification

Production scale-up of high-purity diacetylated C18:1 sophorolipid lactone was demonstrated from lab to pilot scale with the Starmerella bombicola lactone esterase overexpression strain (oe sble) as production organism. The 150 L fermentation using oleic acid and yeast extract, characterized by a titer of 199 g/L and a volumetric productivity of 0.9 g/L·h, was most successful in obtaining a highly pure (>98%) and uniform (96% C18:1 SL lactone) sophorolipid product suitable for chemical derivatization. The fermentation product was subsequently modified to produce sophorolipid amine oxides, which cannot be produced enzymatically. First, the fermentation product was transformed into an intermediate sophorolipid aldehyde via methanolysis and protection of the sugar head through acetylation and ozonolysis. This aldehyde intermediate was then used for the synthesis of the sophorolipid amine oxides via reductive amination, oxidation, and deprotection of the sugar head. The total yield of this synthetic pathway a...

Sophorolipids Production by Candida bombicola ATCC 22214 and Its Potential Application in Soil Bioremediation

The sophorolipids are biosurfactants composed by a disaccharide sophorose linked by a hydroxyl group of a fatty acid. Due to their surface-active properties, sophorolipids have a great potential for using in different industrial sectors. They are synthesized by various strains of the genus Candida in the presence of hydrophilic and hydrophobic sources. In this study, it was evaluated the production of sophorolipids by Candida bombicola ATCC 22214 using sugar cane molasses, sugar cane juice, sucrose or glucose as hydrophilic source and chicken fat or sunflower oil as hydrophobic source. The sophorolipids were characterized and applied in bioremediation of soils contaminated with lubricating oil. The production of sophorolipids was 39.81 g L−1 in the optimized condition of 75 g L−1 of chicken fat, 77.5 g L−1 of glucose, 2.5 g L−1 of yeast extract and without urea. The sophorolipids produced present the diacetylated lactonic form, exhibiting the surface tension properties of 35 mN/m and of CMC 65 mg L−1. The sophorolipids showed great emulsification index in lubricant oil, toluene and n-heptane, suggesting the potential application in bioremediation of lubricating-oil-contaminated soils.

Experimental Investigation of Sophorolipid Biosurfactants Produced by Candida and Pleurotus Species Using Waste Oils and Rice Bran and Their Oilfield Benefits

Conventional chemical surfactants applications usually linked with environmental unfriendliness and toxicity are associated with high production costs resulting from fluctuations in oil prices and thermal energy requirements. Sophorolipid biosurfactants can potentially be implemented with a remarkably low operating cost. Besides economic interest, sophorolipids and their derivatives have shown promise as emulsifiers, antimicrobials, surfactants and a source of specialty chemicals reduction capacities, thus, facilitating microbial hydrocarbon and heavy metal emulsification and uptake. In this study, sophorolipids produced by Candida and Pleurotus species respectively isolated and harvested from oil contaminated soils from Ukwa West LGA of Abia State were investigated. Mineral salt media supplemented with different hydrophilic (rice bran, spent mushroom substrate and cassava peels) and hydrophobic (food industry waste oil) renewable wastes as sources of carbon were tested on nine (9) of the potential isolates to select the best medium and organism that maximized sophorolipids production. This was supported by the emulsification index after 24 hours (E24), FT-IR and GC-MS analysis. All isolates were subjected to biosurfactants production screening, to find the best sophorolipid producer among the available strains. Results showed that isolate CP1SP6c, a hydrocarbon utilizing fungi (HUF) exhibiting β- haemolysis and 92.4% microbial adhesion to hydrocarbon (MATH), gave the maximum E24 of 69.2%, within 6 days of incubation on media fortified with rice bran and food industry waste oil in a rotary shaker. Further studies on CP1SP6c were carried out to assess the interaction of the produced sophorolipids with porous media in core flooding experiments as a tertiary recovery technique. The results showed high promises of using this bio-product in hydrocarbon recovery, where 12.3% of crude oil was recovered after injecting the culture supernatant. An additional recovery of 15.7% of residual oil was observed after concentrating the sophorolipid solution.

Isolation of sophorose during sophorolipid production and studies of its stability in aqueous alkali: epimerisation of sophorose to 2-O-β-d-glucopyranosyl-d-mannose

NMR and anion exchange chromatography analysis of the waste streams generated during the commercial production of sophorolipids by the yeast Candida bombicola identified the presence of small but significant quantities (1% w/v) of free sophorose. Sophorose, a valuable disaccharide, was isolated from the aqueous wastes using a simple extraction procedure and was purified by chromatography on a carbon celite column providing easy access to large quantities of the disaccharide. Experiments were undertaken to identify the origin of sophorose and it is likely that acetylated sophorose derivatives were produced by an enzyme catalysed hydrolysis of the glucosyl-lipid bond of sophorolipids; the acetylated sophorose derivatives then undergo hydrolysis to release the parent disaccharide.Treatment of sophorose with aqueous alkali at elevated temperatures (0.1M NaOH at 50 °C) resulted in C2-epimerisation of the terminal reducing sugar and its conversion to the corresponding 2-O-β-d-glucopyranosyl-d-mannose which was isolated and characterised. In aqueous alkaline solution β-(1,2)-linked glycosidic bonds do not undergo either hydrolysis or peeling reactions.

Sophorolipids Production by Candida bombicola ATCC 22214 and its Potential Application in Microbial Enhanced Oil Recovery.

Biosurfactant production using Candida bombicola ATCC 22214, its characterization and potential applications in enhancing oil recovery were studied at laboratory scale. The seed media and the production media were standardized for optimal growth and biosurfactant production. The production media were tested with different carbon sources: glucose (2%w/v) and corn oil (10%v/v) added separately or concurrently. The samples were collected at 24 h interval up to 120 h and checked for growth (OD660), and biosurfactant production [surface tension (ST) and interfacial tension (IFT)]. The medium with both glucose and corn oil gave better biosurfactant production and reduced both ST and IFT to 28.56 + 0.42mN/m and 2.13 + 0.09mN/m, respectively within 72 h. The produced biosurfactant was quite stable at 13–15% salinity, pH range of 2–12, and at temperature up to 100°C. It also produced stable emulsions (%E24) with different hydrocarbons (pentane, hexane, heptane, tridecane, tetradecane, hexadecane, 1-methylnaphthalene, 2,2,4,4,6,8-heptamethylnonane, light and heavy crude oil). The produced biosurfactant was extracted using ethyl acetate and characterized as a mixture of sophorolipids (SPLs). The potential of SPLs in enhancing oil recovery was tested using core-flooding experiments under reservoir conditions, where additional 27.27% of residual oil (Sor) was recovered. This confirmed the potential of SPLs for applications in microbial enhanced oil recovery.

Towards the industrialization of new biosurfactants: Biotechnological opportunities for the lactone esterase gene from Starmerella bombicola.

Although sophorolipids (SLs) produced by S. bombicola are a real showcase for the industrialization of microbial biosurfactants, some important drawbacks are associated with this efficient biological process, e.g., the simultaneous production of acidic and lactonic SLs. Depending on the application, there is a requirement for the naturally produced mixture to be manipulated to give defined ratios of the components. Recently, the enzyme responsible for the lactonization of SLs was discovered. The discovery of the gene encoding this lactone esterase (sble) enabled the development of promising S. bombicola strains producing either solely lactonic (using a sble overexpression strain described in this paper: oe sble) or solely acidic SLs (using a sble deletion strain, which was recently described, but not characterized yet: Δsble). The new S. bombicola strains were used to investigate the production processes (fermentation and purification) of either lactonic or acidic SLs. The strains maintain the high inherent productivities of the wild-type or even perform slightly better and thus represent a realistic industrial opportunity. 100% acidic SLs with a mixed acetylation pattern were obtained for the Δsble strain, while the inherent capacity to selectively produce lactonic SLs was significantly increased (+42%) for the oe sble strain (99% lactonic SLs). Moreover, the regulatory effect of citrate on lactone SL formation for the wild-type was absent in this new strain, which indicates that it is more robust and better suited for the industrial production of lactonic SLs. Basic parameters were determined for the purified SLs, which confirm that the two new strains produce molecules with distinctive properties of which the application potential can now easily be investigated independently.

Production of Sophorolipid from an Identified Current Yeast, Lachancea thermotolerans BBMCZ7FA20, Isolated from Honey Bee.

Biosurfactants are a family of diverse amphipathic molecules that are produced by several microorganisms such as bacteria, molds, and yeasts. These surface active agents have several applications in agriculture, oil processing, food, and pharmaceutical industries. In this research using YMG and YUG culture media, a native yeast strain, HG5, was isolated from honey bee. The oil spread test as a screening method was used to evaluate biosurfactant production by the yeast HG5 isolate. The 5.8s-rDNA analysis confirmed that the isolated yeast was related to Lachancea thermotolerans. We named this strain Lachancea thermotolerans strain BBMCZ7FA20 and its 5.8s-rDNA sequence was deposited in GenBank, NCBI under accession number of KM042082.1. The best precursor of biosurfactant production was canola oil and the sophorolipid amount was measured for 24.2g/l. The thin layer chromatography and Fourier Transform Infrared Spectroscopy analysis showed that the extracted biosurfactant from Lachancea thermotolerans was sophorolipid. In conclusion, this is the first report of sophorolipid production by a native yeast Lachancea thermotolerans BBMCZ7FA20 we isolated from the honey bee gut collected from an apiary farm in Saman, Chaharmahal Bakhtiari province, Iran. We suggested that some cost-effective supplements such as canola oil, sunflower oil, and corn oils could be applied for increasing the sophorolipid production by this native yeast strain. According to several applications of biosurfactants in today world, the production of sophorolipid by Lachancea thermotolerans could be considered as a potential in the current industrial microbiology and modern microbial biotechnology.

High‐titer production and strong antimicrobial activity of sophorolipids from Rhodotorula bogoriensis

Rhodotorula bogoriensis produces sophorolipids (SLs) that contain 13-hydroxydocosanoic acid (OH-C22 ) as the lipid moiety. A systematic study was conducted to further understand the fermentative production of SLs containing OH-C22 (C22 -SL) by R. bogoriensis. Shake-flask studies showed that R. bogoriensis consumed glucose at a slow pace. HPLC analysis of the C22 -SL products from shake-flask fermentations at different glucose concentrations showed a correlation between glucose depletion and the extent of C22 -SL deacetylation. A large-scale bioreactor fermentation resulted in the isolation of C22 -SL at a volumetric product yield of 51 g/L. HPLC analysis of C22 -SL product from the bioreactor fermentation corroborated the finding that glucose depletion correlated with extensive deacetylation of C22 -SL. The antimicrobial activity of C22 -SL was established for the first time to be stronger than the C18 -SL from Candida bombicola against Propionibacterium acnes in a plate assay.