To determine the performance of a sophorolipid biosurfactant production process, it is important to have accurate and specific analytical techniques in place. Among the most popular are the anthrone assay, gravimetric quantification (hexane:ethyl acetate extraction), and high-performance liquid chromatography (HPLC). The choice of analytical tool varies depending on cost, availability, and ease of use; however, these techniques have never been compared directly against one another. In this work, 75 fermentation broths with varying product/substrate concentrations were comprehensively tested with the 3 techniques and compared. HPLC-ultraviolet detection (198 nm) was capable of quantifying C18:1 subterminal hydroxyl diacetylated lactonic sophorolipid down to a lower limit of 0.3 g/L with low variability (<3.21%). Gravimetric quantification of the broths following liquid:liquid extraction with hexane and ethyl acetate showed some linearity (R2 = .658) when compared to HPLC but could not quantify lower than 11.06 g/L, even when no sophorolipids were detected in the sample, highlighting the non-specificity of the method to co-extract non-sophorolipid components in the final gravimetric measure. The anthrone assay showed no linearity (R2 = .129) and was found to cross-react with media components (rapeseed oil, corn steep liquor, glucose), leading to consistent overestimation of sophorolipid concentration. The appearance of poor biomass separation during sample preparation with centrifugation was noted and resolved with a novel sample preparation method with pure ethanol. Extensive analysis and comparisons of the most common sophorolipid quantification techniques are explored and the limitations/advantages are highlighted. The findings provide a guide for scientists to make an informed decision on the suitable quantification tool that meets their needs, exploring all aspects of the analysis process from harvest, sample preparation, and analysis.

Hydrogen, a clean and sustainable energy source, faces challenges from energy-intensive pre-processing technologies. This study explores the synergistic enhancement of active electric fields on enzymolysis of wheat straw and hydrogen production through dark fermentation. The active electric field enzymolysis system improved the adsorption capacity of wheat straw to cellulase, increasing cellulase activity by 18.0 %, causing a 39.1 % increase in reducing sugar content. In the active fermentation system, Clostridium_sensu_stricto_1 activity was enhanced in the first stage, increasing hydrogenase activity by 23.0 %, prolonging the first hydrogen production peak. Elevated reducing sugars were observed in the second stage, with Prevotella_9 and Bacteroides becoming the dominant hydrogen-producing bacteria in the third stage, leading to a second hydrogen production peak. Overall, cumulative hydrogen production was enhanced by 50.9 % compared to the control. The synergistic pretreatment with an active electric field and cellulase provides a novel approach for efficiently utilizing wheat straw.

To explore a sustainable sophorolipid production, several hydrolysates from agricultural byproducts, such as wheat feed, rapeseed meal, coconut waste and palm waste were used as nitrogen sources. The four hydrolysates overperformed the controls after 168 h of fermentation using Starmerella bombicola ATCC 22214. Wheat feed and coconut waste hydrolysates were the most promising feedstocks presenting a linear relationship between yeast growth and diacetylated lactonic C18:1 production at total nitrogen concentrations below 1.5 g/L (R2 = 0.90 and 0.83, respectively). At 0.31 g/L total nitrogen, wheat feed hydrolysate achieved the highest production, yielding 72.20 ± 1.53 g/L of sophorolipid crude extract and 60.05 ± 0.56 g/L of diacetylated lactonic C18:1 at shake flask scale with productivities of 0.43 and 0.36 g/L/h, respectively. Results were confirmed in a 2-L bioreactor increasing 15 % diacetylated lactonic C18:1 production. Moreover, wheat feed hydrolysate supplemented only with a hydrophobic carbon source was able to produce mainly diacetylated lactonic C18:1 congener (88.5 % wt.), suggesting that the composition of the hydrolysate significantly influences the congeners profile. Overall, this study provides valuable insights into agricultural byproduct hydrolysates as potential nitrogen feedstocks for sophorolipid production and their further application on industrial biotechnology.

Sophorolipids (SLs) are promising glycolipid biosurfactants as they are easily produced and functional. SLs from microorganisms are comprised of mixtures of multiple derivatives that have different structures and properties, including well-known acidic and lactonic SL (ASLs and LSLs, respectively). In this study, we established a method for analyzing all SL derivatives in the products of Starmerella bombicola, a typical SL-producing yeast. Detailed component analyses of S. bombicola products were carried out using reversed-phase high-performance liquid chromatography and mass spectrometry. Methanol was used as the eluent as it is a good solvent for all SL derivatives. With this approach, it was possible to not only quantify the ratio of the main components of ASL, LSL, and SL glycerides but also confirm trace components such as SL mono-glyceride and bola-form SL (sophorose at both ends); notably, this is the first time these components have been isolated and identified successfully in naturally occurring SLs. In addition, our results revealed a novel SL derivative in which a fatty acid is bonded in series to the ASL, which had not been reported previously. Using the present analysis method, it was possible to easily track compositional changes in the SL components during culture. Our results showed that LSL and ASL are produced initially and that SL glycerides accumulate from the middle stage during the fermentation process. KEY POINTS: • An easy and detailed component analysis method for sophorolipids (SLs) is introduced. • Multiple SL derivatives were identified different from known SLs. • A novel hydrophobic acidic SL was isolated and characterized.

Surfactants are compounds with high surface activity and emulsifying property. These compounds find application in food, medical, pharmaceutical, and petroleum industries, as well as in agriculture, bioremediation, cleaning, cosmetics, and personal care product formulations. Due to their widespread use and environmental persistence, ensuring biodegradability and sustainability is necessary so as not to harm the environment. Biosurfactants, i.e., surfactants of plant or microbial origin produced from lignocellulosic feedstock, perform better than their petrochemically derived counterparts on the scale of net-carbon-negativity. Although many biosurfactants are commercially available, their high cost of production justifies their application only in expensive pharmaceuticals and cosmetics. Besides, the annual number of new biosurfactant compounds reported is less, compared to that of chemical surfactants. Multiple operational issues persist in the biosurfactant value chain. In this review, we have categorized some of these issues based on their relative position in the value chain - hurdles occurring during planning, upstream processes, production stage, and downstream processes - alongside plausible solutions. Moreover, we have presented the available paths forward for this industry in terms of process development and integrated pretreatment, combining conventional tried-and-tested strategies, such as reactor designing and statistical optimization with cutting-edge technologies including metabolic modeling and artificial intelligence. The development of techno-economically feasible biosurfactant production processes would be instrumental in the complete substitution of petrochemical surfactants, rather than mere supplementation.

The use of alternative feedstocks such as industrial or food waste is being explored for the sustainable production of sophorolipids (SLs). Microbial biosurfactants are mainly produced via submerged fermentation (SmF); however, solid-state fermentation (SSF) seems to be a promising alternative for using solid waste or byproducts that could not be exploited by SmF. Applying the advantages that SSF offers and with the aim of revalorizing industrial organic waste, the impact of carbon and nitrogen sources on the relationship between yeast growth and SL production was analyzed. The laboratory-scale system used winterization oil cake as the solid waste for a hydrophobic carbon source. Pure hydrophilic carbon (glucose) and nitrogen (urea) sources were used in a Box-Behnken statistical design of experiments at different ratios by applying the response surface methodology. Optimal conditions to maximize the production and productivity of diacetylated lactonic C18:1 were a glucose:nitrogen ratio of 181.7:1.43 (w w-1 based on the initial dry matter) at a fermentation time of 100 h, reaching 0.54 total gram of diacetylated lactonic C18:1 with a yield of 0.047 g per gram of initial dry mass. Moreover, time course fermentation under optimized conditions increased the SL crude extract and diacetylated lactonic C8:1 production by 22% and 30%, respectively, when compared to reference conditions. After optimization, industrial wastes were used to substitute pure substrates. Different industrial sludges, OFMSW hydrolysate, and sweet candy industry wastewater provided nitrogen, hydrophilic carbon, and micronutrients, respectively, allowing their use as alternative feedstocks. Sweet candy industry wastewater and cosmetic sludge are potential hydrophilic carbon and nitrogen sources, respectively, for sophorolipid production, achieving yields of approximately 70% when compared to the control group.

Exploring the natural, safe, and effective antimicrobial is one of the preferable ways to control foodborne bacteria. In this work, novel oil-in-water nanoemulsions were formulated with sophorolipids and eugenol without any co-surfactant using a self-assembling strategy. These nanoemulsions showed high stability with sizes less than 200 nm when exposure to low concentrations of salt ions, various pH values (5.0, 7.0, 10.0), storage temperature and time. The synergistic antibacterial effects against both Gram-negative Escherichia coli and Gram-positive Bacillus cereus were determined with a minimum inhibitory concentration (MIC) value of 0.5 mg/mL and 0.125 mg/mL, respectively. Further microscopy (SEM, TEM, LCSM) examination and ATP/Na+-K+-ATPase assay results showed that the morphological changes, intensive cell membrane permeability, leakage of ATP, and decreased Na+-K+-ATPase contributed to the antibacterial effects. Moreover, the bonding mechanism between nanoemulsions and cell membranes were further evaluated by FTIR and ITC using a DPPC vesicle model, which demonstrated that the nanoemulsions adsorbed on the surface of bilayer, interacted with the hydrophobic chains of DPPC membrane mainly through the hydrophobic interaction, and altered the structural integrity of the lipid bilayer. These results not only provide a facile green strategy for fabricating stable nanoemulsions, but also highlight a new perspective for stabilizing essential oils for their widely application in food industry.

Sophorolipids (SLs) are surface active compounds that have excellent surface-lowering properties. SLs were produced by Starmerella bombicola (CGMCC1576) yeast with sunflower seed oil, fried waste oil, cooked tung oil and raw tung oil used as hydrophobic carbon sources. The results showed that the strain could use sunflower seed oil and fried waste oil as hydrophobic carbon sources to produce SLs, and the yields were 44.52 and 39.09 gl-1. It could not be used as cooked tung oil and raw tung oil. The analysis by high-performance liquid chromatography/high resolution mass spectrometry (HPLC-MS/MS) showed that the main composition and structure of SLs produced by fermentation using fried waste oil were similar to that of sunflower seed oil as hydrophobic carbon source. The yield of SLs was the highest when the fried waste oil was used as hydrophobic carbon source, glucose (8%), waste oil (6%) and yeast (0.3%). When fried waste oil was used as a hydrophobic carbon source in a parallel 4-strand fermentation tank (FT), the combination with the largest yield and the most cost saving was that 3% of fried waste oil was added into the initial medium, and another 3% was again added after 72 h of fermentation. The total yield of SLs was 121.28 gl-1, and the yield of lactone SLs was 48.07 gl-1.

Introduction: New bioresources for catalytic application and fine chemical synthesis are the need of the hour. In an effort to find out new biocatalyst for oxidation-reduction reaction, leading to the synthesis of chiral intermediates, novel yeast were isolated from unique niche and employed for the synthesis of value added compounds. Methods: To determine the genetic relatedness of the isolated strain, HSB-15T, sequence analysis of the internal transcribed spacer (ITS) and D1/D2 domains of the 26S rRNA gene sequence was carried out. The distinctive features of the strain HSB-15T were also identified by phenotypic characterization. The isolated strain HSB-15T was employed for the reduction of selected naphthyl ketones to their corresponding alcohols and a biosurfactant was isolated from its culture broth. Results: The analysis of the ITS and D1/D2 domains of the 26S rRNA gene revealed that strain HSB-15T is closely related to the type strain of Starmerella vitae (CBS 15147T) with 96.3% and 97.7% sequence similarity, respectively. However, concatenated sequences of the ITS gene and D1/D2 domain showed 94.6% sequence similarity. Phenotypic characterization indicated significant differences between strain HSB-15T and its closely related species and consequently, it was identified as a novel species, leading to the proposal of the name Starmerella cerana sp. nov. The strain was able to reduce selected naphthyl ketones to their corresponding alcohols with remarkable efficiency, within a 12-hours. The strain HSB-15T also produced a surfactant in its culture broth, identified as sophorolipid upon analysis. Discussion: The study explored the potential of the novel strain, HSB-15T, as a whole-cell biocatalyst for the reduction of naphthyl ketones to their corresponding alcohols and also reports its capability to produce sophorolipid, a biosurfactant, in its culture broth. This dual functionality of HSB-15T both as biocatalyst and biosurfactant producer enhances its applicability in biotechnology and environmental science.

The removal of NH4+, NO3-, and NH3- from wastewater can be difficult and expensive. Through physical, chemical, and biological processes, metals and nutrients can be extracted from wastewater. Very few scientific investigations have employed surfactants with high biodegradability, low toxicity, and suitability for ion removal from wastewater at different pH and salinity levels. This research employed a highly biodegradable biosurfactant generated from yeast (sophorolipid) through micellar-enhanced ultrafiltration (MEUF). MEUF improves nutrient removal efficiency and reduces costs by using less pressure than reverse osmosis (RO) and nanofiltration (NF). The biosurfactant can be recovered after the removal of nutrient- and ion-containing micelles from the filtration membrane. During the experiment, numerous variables, including temperature, pH, biosurfactant concentration, pollutant ions, etc., were evaluated. The highest amount of PO43- was eliminated at a pH of 6.0, which was reported at 94.9%. Maximum NO3- removal occurred at 45.0 °C (96.9%), while maximum NH4+ removal occurred at 25.0 mg/L (94.5%). Increasing TMP to 200 kPa produced the maximum membrane flow of 226 L/h/m2. The concentrations of the contaminating ion and sophorolipid were insignificant in the permeate, demonstrating the high potential of this approach.