Modeling of Rhamnolipid Biosurfactant Production: Estimation of Kinetic Parameters by Genetic Algorithm

Studies about kinetics and modeling of production parameters for biosurfactants are essential to the development of efficient processes from an economic point of view. In this sense, this work evaluated the performance of four nonstructured models to explain the experimental data for biomass growth, substrate consumption, and rhamnolipid production using glycerol as carbon source and a Pseudomonas aeruginosa strain. The kinetic parameters of each model were estimated using a global search method known as genetic algorithm and numerical discretization of differential equations by the Runge–Kutta 4th order method. The main result of this study showed that the Monod model best represented the experimental data, with μ_max values of 0.06 h⁻¹, K_S of 50.8 g L⁻¹, Y_X/S of 0.43 g g⁻¹, and Y_P/X equal to 0.017 g g⁻¹.     

高效产表面活性剂菌株（Lz2～1）的筛选及其特性研究

从24份含油土壤和水等样品中,经富集培养、摇瓶培养和排油活性测定等方法筛选出一株能以原油为碳源产生表面活性剂的菌株Lz2—1;该菌株可以将水的表面张力由72mN／m降到28mN／m,且发酵液具有较好的乳化活性;经生理生化16SrDNA及生理生化实验确定该菌株为铜绿假单胞菌;提取其代谢产物经薄层层析及红外色谱分析显示,主要的表面活性剂类物质为鼠李糖脂类,其临界胶束质量浓度为0．63047g／L。结果表明,表面活性物质是Lz2—1菌株在微生物采油过程中发挥作用的主要因素。     

Biolytic Effect of Rhamnolipid Biosurfactant and Dodecyl Sulfate Against Phagotrophic Alga <Emphasis Type="Italic">Ochromonas danica</Emphasis>

Surfactants can affect biological activities and pose serious threats to the aquatic ecosystem if discharged without proper treatment. Rhamnolipid is a biosurfactant with promising agricultural, industrial and biomedical applications. It is important to assess the risks posed by rhamnolipid before it is adopted for large scale use. In this study, toxicity of rhamnolipid was estimated with the effects of motility loss, membrane permeability, and cell lysis using a phagotrophic alga Ochromonas danica. O. danica is a sensitive algal species without a protective cell wall and it represents a group of phagotrophic flagellates ecologically important in the aquatic ecosystem. A common synthetic surfactant, sodium dodecyl sulfate (SDS), was used for comparison. Results suggested that rhamnolipid is significantly less biolytic than SDS and the effects required longer exposure. Lysis of O. danica cells by rhamnolipid was serious only at concentrations over its critical micelle concentration. Motility loss was, however, significantly more sensitive; about 75% loss after 20 min exposure to 50 mg/L rhamnolipid.     

Recycling of Bakery Waste as an Alternative Carbon Source for Rhamnolipid Biosurfactant Production

Production of a rhamnolipid biosurfactant (RBS) using discarded mixed bakery waste (BW) employing bacterial strain Pseudomonas aeruginosa strain PG1 (identified by 16 s rDNA sequencing) was investigated for bioconversion of the food waste. Dry and powder form BW was supplemented with mineral salt media (MSM) as a sole carbon source for production of RBS. RBS production was measured based on the drop collapse assay and surface tension (ST) reduction of the culture media. Production of RBS in the culture media was enhanced by optimizing the carbon source (BW) concentration and the proper nitrogen source along with the pH of the MSM. Under optimized culture conditions, 11.56 g L⁻¹ day⁻¹ crude biosurfactant (BS) was achieved. The RBS had the ability to reduce the ST of the optimized MSM from 72.0 to 25.8 mN m⁻¹ during culture, where the critical micelle concentration (CMC) of the biosurfactant was found to be 100 mg L⁻¹. Liquid Chromatography Mass Spectroscopy (LC-MS), Fourier Transform Infrared spectroscopy (FTIR), and scanning electron microscopy (SEM)–energy dispersive X-ray spectroscopy (EDS) analyses of the purified BS confirmed that it is of rhamnolipid in nature and it is made up of both monorhamnolipid and dirhamnolipid congeners. Furthermore, the RBS did not express any cytotoxic effect on the cell line of mouse L292 fibroblastic cell indicating the biosafety nature of the high-value biomolecule.     

Sunflower Acid Oil-Based Production of Rhamnolipid Using Pseudomonas aeruginosa and Its Application in Liquid Detergents

Utilization of industrial waste as substrates for the rhamnolipid synthesis by Pseudomonas aeruginosa is a worthy alternative for conventionally used vegetable oils and fatty acids to reduce the production cost of rhamnolipid. Sunflower acid oil (SAO), a by-product of the oil industry, contains 70% 18:0 fatty acid, with oleic acid as a major component. In this scope, production and analysis of rhamnolipid was successfully demonstrated using SAO as a new substrate. Pseudomonas aeruginosa produced rhamnolipid (a glycolipid biosurfactant) at a maximum concentration of 4.9 g L⁻¹ with 60 g L⁻¹ of SAO in the medium. Structural properties of rhamnolipid biosurfactant are confirmed using thin layer chromatography (TLC), high performance liquid chromatography (HPLC), and fourier transformed infrared spectroscopy (FTIR) analysis. Further surface-active properties of the crude rhamnolipid were evaluated by measuring surface tension and emulsification properties. The synthesized rhamnolipid reduced the surface tension of water to 30.12 mN m⁻¹ and interfacial tension (against heptane) to 0.52 mN m⁻¹. Moreover, rhamnolipid shows the highest emulsification index (above 80%) for vegetable oils. This study confirms the use of SAO as a potential substrate for rhamnolipid production. The synthesized rhamnolipid was incorporated in liquid detergent formulation along with alpha olefin sulfonate (AOS) and sodium lauryl ether sulfate (SLES). The performance properties including foaming and cleaning efficiency of liquid detergent were compared.     

Rhamnolipid–Metal Ions (CrVI and PbII) Complexes: Spectrophotometric,Conductometric, and Surface Tension Measurement Studies

The formation of rhamnolipid complexes with metal ions of chromium (VI) and lead (II) has been studied spectrophotometrically, conductometrically, and by surface tension measurement. The values of the critical micelle concentration (CMC) of rhamnolipid, obtained by spectroscopic, conductometric, and surface tension measurements, were 5.2 × 10⁵ , 5.0 × 10⁵ , and 5.3 × 10⁵ mol.dm⁻³, respectively, which are in close agreement. An increase in CMC on increasing metal ion content in the rhamnolipid solution and a shift in λ_max in the spectra of rhamnolipid indicated the formation of a complex between rhamnolipid and both the metal ions, namely, chromium (VI) and lead (II). The values of stability constants for the {rhamnolipid–chromium (VI)} and {rhamnolipid–lead (II)} complexes have been determined by spectroscopic data and were as 0.58 × 10⁴ and 0.50 × 10⁴ at 308 K, respectively. The thermodynamic parameters for micellization, namely, free energy change (∆G_mic) , entropy change (∆S_mic) , and heat enthalpy change (∆H_mic) , have been determined by conductivity measurements. An increase in the negative value of ∆G_mic and a decrease in the value of ∆S_mic on increasing metal ion content in the surfactant solution indicated lower micellization of rhamnolipid in the presence of metal ions. The electrostatic attractions and entrapment of chromium (VI) in the micelles of the biosurfactants were found to be responsible for {rhamnolipid–lead (II)} and {rhamnolipid–chromium (VI)} complexes, respectively.     

Mechanism Study on the Severe Foaming of Rhamnolipid in Fermentation

Although the biosurfactant rhamnolipid has been previously characterized as having low foam ability, its fermentation is largely impeded by severe foaming. Hence, the investigation of this paradox is critically important for improving the mass production of rhamnolipid. Unexpectedly, the hydrophobic cell, instead of rhamnolipid, has been claimed to explain such severe foaming in rhamnolipid fermentation. This study tried to systematically investigate the severe foaming in fermentation, aiming to propose an effective strategy for foam control. The overflowing foam sustained a super high stability in terms of half‐time for over 30 min. The major product of rhamnolipid largely contributed to the severe foaming in the fermentation process whereas other products like cells elicited much more limited effects. Furthermore, the foam stability of the fermentation broth increased with rhamnolipid concentration and noticeably increased with agitation speed. In the classic Bikerman foam test system without stirring, rhamnolipid showed foam stability as low as Tween 20 which is well known for its poor foam stability. However, in a stirring Bikerman system, rhamnolipid exhibited a foam stability almost as high as sodium dodecyl sulfate (SDS) at 10 g/L and even surpassed SDS at a higher concentration of 20 g/L. Hence, the extraordinarily increased foam stability of rhamnolipid with both agitation and concentration could explain the severe foaming at its late‐stage fermentation when rhamnolipid‐rich solution is mechanically agitated.     

DLVO AND NON-DLVO FORCES IN THIN LIQUID FILMS FROM RHAMNOLIPIDS

Microscopic foam films (r?=?100?μm) stabilized with a single rhamnolipid with a well-known structure (α-L-rhamnopyranosyl-β-hydroxydecanoyl-β-hydroxydecanoate (R1)) are investigated, and the obtained results are compared with results obtained from studies of foam films formed from solutions of rhamnolipid mixtures. The studies are carried out employing the Scheludko-Exerowa microinterferometric method. The dependence of foam film thickness on the electrolyte concentration (C _el ) of the solution is monitored, and formation of common films (CF), common black films (CBF) and Newton black films (NBF) is found. The continuous CBF-to-NBF transition is considered as evidence of the action of repulsive forces that are not described by the classic Derjaguin–Landau–Verwey–Overbeek (DLVO) theory of colloid stability. These non-DLVO repulsive forces lead to an additional positive component of the disjoining pressure. To understand better the surface forces operating in the rhamnolipid foam films, direct measurements of the disjoining pressure/film thickness (Π(h)) isotherms are carried out employing the thin liquid film–pressure balance technique. The comparison of the obtained experimental Π(h) isotherm for CF (C _el ?=?10^?3 mol dm^?3 NaCl) to the DLVO theoretical predictions yields a diffuse electric layer potential of?～?5?mV and surface charge density of?～?50?mC?m^?2 at the film solution–air interfaces. The deviation of the experimental curve from the theoretical one found for films thinner than about 40?nm evidences the action of non-DLVO surface forces. The experimental steplike Π(h) isotherms obtained for the CBF (C _el ?=?0.15?mol?dm^?3 NaCl) are considered to result from an aggregation process, leading to the formation of lamellar structures in the foam film. The obtained results show that the surface forces operative in rhamnolipid foam films are determined not only by the structure and organization of the adsorbed layers but also by the molecular state of the bulk solution.     

Two-stage co-hydrolysis of rice straw by Trichoderma reesei ZM4-F3 and Pseudomonas aeruginosa BSZ-07

Rhamnolipid biosurfactant was added to rice straw hydrolysis system to enhance the production of reducing sugars. Differing from the traditional method, on-site production of rhamnolipid made the rice straw decomposing fungus Trichoderma reesei ZM4-F3 and rhamnolipid producing bacteria Pseudomonas aeruginosa BSZ-07 work together. As the growth periods of these two strains are 96 h and 48 h, respectively, a new two-stage co-hydrolysis bioprocess was achieved. T. reesei ZM4-F3 was cultivated for rice straw hydrolysis in the first 48 h at suitable conditions. For the next 48 h, the results showed that the temperature of 34 °C and pH value of 5.5 were the optimum conditions, and the optimum adding inoculation concentration ratio of P. aeruginosa BSZ-07 to T. reesei ZM4-F3 was 4%. Under these conditions, the co-hydrolysis sample could improve the production of reducing sugars to 2.57 g l⁻¹, 15.20% higher than that of the control. The increased enzyme stability was indicated to be one of the mechanisms of the stimulatory effect of rhamnolipid on rice straw hydrolysis system. Compared with Tween-80 and sodium dodecylsulphate, rhamnolipid biosurfactant exhibited a better stimulatory effect on rice straw hydrolysis. Since the two-stage co-hydrolysis system could leave out the rhamnolipid purification process, thus reducing the biosurfactant production cost effectively, it seems to be a new prospective bioprocess for rice straw hydrolysis.