Enhancement of Biomass and Calcium Carbonate Biomineralization of Chlorella vulgaris through Plackett-Burman Screening and Box-Behnken Optimization Approach.

The biosynthesis of calcium carbonate (CaCO3) minerals through a metabolic process known as microbially induced calcium carbonate precipitation (MICP) between diverse microorganisms, and organic/inorganic compounds within their immediate microenvironment, gives rise to a cementitious biomaterial that may emerge as a promissory alternative to conventional cement. Among photosynthetic microalgae, Chlorella vulgaris has been identified as one of the species capable of undergoing such activity in nature. In this study, response surface technique was employed to ascertain the optimum condition for the enhancement of biomass and CaCO3 precipitation of C. vulgaris when cultured in Blue-Green (BG)-11 aquaculture medium. Preliminary screening via Plackett-Burman Design showed that sodium nitrate (NaNO3), sodium acetate, and urea have a significant effect on both target responses (p < 0.05). Further refinement was conducted using Box-Behnken Design based on these three factors. The highest production of 1.517 g/L C. vulgaris biomass and 1.143 g/L of CaCO3 precipitates was achieved with a final recipe comprising of 8.74 mM of NaNO3, 61.40 mM of sodium acetate and 0.143 g/L of urea, respectively. Moreover, polymorphism analyses on the collected minerals through morphological examination via scanning electron microscopy and crystallographic elucidation by X-ray diffraction indicated to predominantly calcite crystalline structure.

Influence of amino acids and vitamins on the growth of gdhA derivative Pasteurella multocida B:2 for use as an animal vaccine.

Pasteurella multocida serotype B:2 is the causative agent of haemorrhagic septicaemia, a fatal disease in cattle and buffaloes. For use as a vaccine in the treatment of HS disease, an efficient cultivation of attenuated gdhA derivative P. multocida B:2 (mutant) for mass production of viable cells is required. In this study, the role of amino acids and vitamins on the growth of this particular bacterium was investigated. Initially, three basal media (Brain-heart infusion, Terrific broth, and defined medium YDB) were assessed in terms of growth performance of P. multocida B:2. YDB medium was selected and redesigned to take into account the effects of amino acids (glutamic acid, cysteine, glycine, methionine, lysine, tyrosine, and histidine) and vitamins (vitamin B1, nicotinic acid, riboflavin, pyridoxine, pantothenic acid, and biotin). High viable cell number was largely affected by the availability of micronutrient components and macronutrients. Histidine was essential for the growth whereby a traceable amount (20 mM) was found to greatly enhance the growth of gdhA derivative P. multocida B:2 mutant (6.6 × 109 cfu/mL) by about 19 times as compared to control culture (3.5 × 108 cfu/mL). In addition, amongst the vitamins added, riboflavin exhibited the highest impact on the viability of gdhA derivative P. multocida B:2 mutant (5.3 × 109 cfu/mL). Though the combined histidine and riboflavin in the culture eventually did not promote the stacking impact on cell growth and cell viability, nonetheless, they were still essential and important in either growth medium or production medium.

Development of Palm Fatty Acid Distillate-Containing Medium for Biosurfactant Production by Pseudomonas sp. LM19.

High production costs of biosurfactants are mainly caused by the usage of the expensive substrate and long fermentation period which undermines their potential in bioremediation processes, food, and cosmetic industries even though they, owing to the biodegradability, lower toxicity, and raise specificity traits. One way to circumvent this is to improvise the formulation of biosurfactant-production medium by using cheaper substrate. A culture medium utilizing palm fatty acid distillate (PFAD), a palm oil refinery by-product, was first developed through one-factor-at-a-time (OFAT) technique and further refined by means of the statistical design method of factorial and response surface modeling to enhance the biosurfactant production from Pseudomonas sp. LM19. The results shows that, the optimized culture medium containing: 1.148% (v/v) PFAD; 4.054 g/L KH2PO4; 1.30 g/L yeast extract; 0.023 g/L sodium-EDTA; 1.057 g/L MgSO4·7H2O; 0.75 g/L K2HPO4; 0.20 g/L CaCl2·2H2O; 0.080 g/L FeCl3·6H2O gave the maximum biosurfactant productivity. This study demonstrated that the cell concentration and biosurfactant productivity could reach up to 8.5 × 109 CFU/mL and 0.346 g/L/day, respectively after seven days of growth, which were comparable to the values predicted by an RSM regression model, i.e., 8.4 × 109 CFU/mL and 0.347 g/L/day, respectively. Eleven rhamnolipid congeners were detected, in which dirhamnolipid accounted for 58% and monorhamnolipid was 42%. All in all, manipulation of palm oil by-products proved to be a feasible substrate for increasing the biosurfactant production about 3.55-fold as shown in this study.

Culture Medium Development for Microbial-Derived Surfactants Production-An Overview.

Surfactants are compounds that can reduce the surface tension between two different phases or the interfacial tension of the liquid between water and oil, possessing both hydrophilic and hydrophobic moieties. Biosurfactants have traits that have proven to be advantageous over synthetic surfactants, but these compounds do not compete economically with synthetic surfactants. Different alternatives increase the yield of biosurfactants; development of an economical production process and the usage of cheaper substrates during process have been employed. One of the solutions relies on the suitable formulation of a production medium by including alternative raw materials sourced from agro-wastes, hydrocarbons, or by-products of a process might help in boosting the biosurfactant production. Since the nutritional factors required will be different among microorganisms, the establishment of a suitable formulation for biosurfactant production will be challenging. The present review describes various nutrients and elements considered in the formulation of a production medium with an approach focusing on the macronutrient (carbon, nitrogen source, and C/N ratio), minerals, vitamins, metabolic regulators, and salinity levels which may aid in the study of biosurfactant production in the future.

Kinetics and Optimization of Lipophilic Kojic Acid Derivative Synthesis in Polar Aprotic Solvent Using Lipozyme RMIM and Its Rheological Study.

The synthesis of kojic acid derivative (KAD) from kojic and palmitic acid (C16:0) in the presence of immobilized lipase from Rhizomucor miehei (commercially known as Lipozyme RMIM), was studied using a shake flask system. Kojic acid is a polyfunctional heterocycles that acts as a source of nucleophile in this reaction allowing the formation of a lipophilic KAD. In this study, the source of biocatalyst, Lipozyme RMIM, was derived from the lipase of Rhizomucor miehei immobilized on weak anion exchange macro-porous Duolite ES 562 by the adsorption technique. The effects of solvents, enzyme loading, reaction temperature, and substrate molar ratio on the reaction rate were investigated. In one-factor-at-a-time (OFAT) experiments, a high reaction rate (30.6 × 10-3 M·min-1) of KAD synthesis was recorded using acetone, enzyme loading of 1.25% (w/v), reaction time of 12 h, temperature of 50 °C and substrate molar ratio of 5:1. Thereafter, a yield of KAD synthesis was optimized via the response surface methodology (RSM) whereby the optimized molar ratio (fatty acid: kojic acid), enzyme loading, reaction temperature and reaction time were 6.74, 1.97% (w/v), 45.9 °C, and 20 h respectively, giving a high yield of KAD (64.47%). This condition was reevaluated in a 0.5 L stirred tank reactor (STR) where the agitation effects of two impellers; Rushton turbine (RT) and pitch-blade turbine (PBT), were investigated. In the STR, a very high yield of KAD synthesis (84.12%) was achieved using RT at 250 rpm, which was higher than the shake flask, thus indicating better mixing quality in STR. In a rheological study, a pseudoplastic behavior of KAD mixture was proposed for potential application in lotion formulation.

Comparative analyses of response surface methodology and artificial neural network on medium optimization for Tetraselmis sp. FTC209 grown under mixotrophic condition.

Mixotrophic metabolism was evaluated as an option to augment the growth and lipid production of marine microalga Tetraselmis sp. FTC 209. In this study, a five-level three-factor central composite design (CCD) was implemented in order to enrich the W-30 algal growth medium. Response surface methodology (RSM) was employed to model the effect of three medium variables, that is, glucose (organic C source), NaNO3 (primary N source), and yeast extract (supplementary N, amino acids, and vitamins) on biomass concentration, X(max), and lipid yield, P(max)/X(max). RSM capability was also weighed against an artificial neural network (ANN) approach for predicting a composition that would result in maximum lipid productivity, Pr(lipid). A quadratic regression from RSM and a Levenberg-Marquardt trained ANN network composed of 10 hidden neurons eventually produced comparable results, albeit ANN formulation was observed to yield higher values of response outputs. Finalized glucose (24.05 g/L), NaNO3 (4.70 g/L), and yeast extract (0.93 g/L) concentration, affected an increase of X(max) to 12.38 g/L and lipid a accumulation of 195.77 mg/g dcw. This contributed to a lipid productivity of 173.11 mg/L per day in the course of two-week cultivation.

Improvement of ɣ-Aminobutyric Acid Production and Cell Viability of Lactiplantibacillus plantarum B7 via Whole-Cell Immobilisation in Repeated Batch Fermentation System.

Whole-cell immobilisation technology involving ℽ-aminobutyric acid GABA biosynthesis using lactic acid bacteria (LAB) has been extensively studied owing to its numerous benefits over free-living bacteria, including enhanced productivity, improved cell viability, ability to prevent cell lysis and protect cells against bacteriophages and other stressful conditions. Therefore, a novel LAB biocatalyst was developed using various fruit and fruit waste, immobilising a potential probiotic strain, Lactiplantibacillus plantarum B7, via an adsorption method to improve GABA and cell viability. Apple and watermelon rind have been known to be the ideal natural supports for L. plantarum B7 owing to higher GABA and lactic acid production and improved cell viability among the other natural supports tested and selected to be used in repeated batch fermentation (RBF) to improve GABA production and cell viability. In general, immobilisation of L. plantarum B7 on natural support has better GABA and lactic acid production with improved cell viability via RBF compared to free cells. Watermelon rind-supported cells and apple-supported cells could produce nine and eight successful GABA cycles, respectively, within RBF, whereas free cells could only produce up to four cycles. When using watermelon rind-supported cells and apple-supported cells in RBF, the GABA titer may be raised by up to 6.7 (218.480 ± 0.280 g/L) and 6 (195.439 ± 0.042 g/L) times, respectively, in comparison to GABA synthesis by free cells in single batch fermentation (32.65 ± 0.029 g/L). Additionally, natural support immobilised L. plantarum B7 could retain half of its cell viability even after the 12th cycle of RBF, while no cell was observed in control.

Strategies for improvement of gamma-aminobutyric acid (GABA) biosynthesis via lactic acid bacteria (LAB) fermentation.

Gamma-aminobutyric acid (GABA) is a non-protein amino acid widely distributed in nature and extensively explored for its numerous physiological functions and effects on metabolic disorders. Lactic acid bacteria (LAB) are one of the most important GABA producers, vigorously pursued due to their high GABA content and generally regarded as safe (GRAS) status that allows for direct formulation in various GABA-enriched food products. To meet the strict requirements of the food and nutraceutical industries, the biosynthesis of GABA is typically preferred over the chemical synthesis route. The production of GABA varies among various strains of LAB and is affected by different fermentation conditions. Hence, optimizing the fermentation conditions to enhance the activity of the key enzyme glutamic acid decarboxylase is essential to maximize GABA production. This paper reviews the beneficial effects of GABA on human health and its applications in fermented food products. A particular emphasis is given to the biosynthetic approach for producing GABA by various LAB species via the microbial fermentation route. Efficient strategies for enhancing GABA production through optimization of the fermentation conditions, mode of fermentation, two-step fermentation, co-culturing approach, immobilization technique and genetic engineering are discussed in detail.

Production of single cell oil by Yarrowia lipolytica JCM 2320 using detoxified desiccated coconut residue hydrolysate.

Nowadays, the replacement of petro-diesel with biodiesel has raised the concern among the community for the utilization of improper feedstocks and the cost involved. However, these issues can be solved by producing single cell oil (SCO) from lignocellulosic biomass hydrolysates by oleaginous microorganisms. This study introduced Yarrowia lipolytica JCM 2320 with a desiccated coconut residue (DCR) hydrolysate (obtained from the 2% dilute sulphuric acid pretreatment) as a carbon source in generating SCO. However, common inhibitors formed during acid pretreatment of biomass such as five-hydroxymethylfurfural (HMF), furfural, acetic acid and levulinic acid resulting from the sugar degradations may have detrimental effects towards the fermentation process. To visualize the effect of inhibitors on Y. lipolytica, an inhibitory study was conducted by adding 0.5-5.0 g/L of potential inhibitors to the YPD (yeast, peptone and D-glucose) medium. It was found that the presence of furfural at 0.5 g/L would increase the lag phase, which beyond that was detrimental to Y. lipolytica. Furthermore, increasing the five-hydroxymethylfurfural (HMF) concentration would increase the lag phase of Y. lipolytica, whereas, for acetic acid and levulinic acid, it showed a negligible effect. Detoxification was hence conducted to remove the potential inhibitors from the DCR hydrolysate prior its utilization in the fermentation. To examine the possibility of using adsorption resins for the detoxification of DCR hydrolysate, five different resins were tested (Amberlite® XAD-4, Amberlite® XAD-7, Amberlite® IR 120, Amberlite® IRA 96 and Amberlite® IRA 402) with five different concentrations of 1%, 3%, 5%, 10% and 15% (w/v), respectively. At resin concentration of 10%, Amberlite® XAD-4 recorded the highest SCO yield, 2.90 ± 0.02 g/L, whereas the control and the conventional overliming detoxification method, recorded only 1.29 ± 0.01 g/L and 1.27 ± 0.02 g/L SCO accumulation, respectively. Moreover, the fatty acid profile of the oil produced was rich in oleic acid (33.60%), linoleic acid (9.90%), and palmitic acid (14.90%), which indicates the potential as a good biodiesel raw material.

The State of Starch/Hydroxyapatite Composite Scaffold in Bone Tissue Engineering with Consideration for Dielectric Measurement as an Alternative Characterization Technique.

Hydroxyapatite (HA) has been widely used as a scaffold in tissue engineering. HA possesses high mechanical stress and exhibits particularly excellent biocompatibility owing to its similarity to natural bone. Nonetheless, this ceramic scaffold has limited applications due to its apparent brittleness. Therefore, this had presented some difficulties when shaping implants out of HA and for sustaining a high mechanical load. Fortunately, these drawbacks can be improved by combining HA with other biomaterials. Starch was heavily considered for biomedical device applications in favor of its low cost, wide availability, and biocompatibility properties that complement HA. This review provides an insight into starch/HA composites used in the fabrication of bone tissue scaffolds and numerous factors that influence the scaffold properties. Moreover, an alternative characterization of scaffolds via dielectric and free space measurement as a potential contactless and nondestructive measurement method is also highlighted.

Valorization of biodiesel side stream waste glycerol for rhamnolipids production by Pseudomonas aeruginosa RS6.

Biodiesel side stream waste glycerol was identified as a cheap carbon source for rhamnolipids (RLs) production which at the same time could improve the management of waste. The present study aimed to produce RLs by using Pseudomonas aeruginosa RS6 utilizing waste glycerol as a substrate and to evaluate their physico-chemicals properties. Fermentation conditions such as temperature, initial medium pH, waste glycerol concentration, nitrogen sources and concentrations resulted in different compositions of the mono- and di-RLs produced. The maximum RLs production of 2.73 g/L was obtained when P. aeruginosa RS6 was grown in a basal salt medium supplemented with 1% waste glycerol and 0.2 M sodium nitrate at 35 °C and pH 6.5. At optimal fermentation conditions, the emulsification index (E24) values of cooking oil, diesel oil, benzene, olive oil, petroleum, and kerosene were all above E24=50%. The surface tension reduction obtained from 72.13 mN/m to 29.4-30.4 mN/m was better than the surface activity of some chemical-based surfactants. The RLs produced possessed antimicrobial activities against Gram-negative and Gram-positive bacteria with values ranging from 37% to 77% of growth inhibition when 1 mg/mL of RLs was used. Concentrations of RLs below 1500 µg/mL did not induce phytotoxicity effects on the tested seeds (Vigna radiata) compared to the chemical-based- surfactant, SDS. Furthermore, RLs tested on zebrafish (Danio rerio) embryos only exhibited low acute toxicity with an LC50 value of 72.97 µg/mL at 48 h of exposure suggesting a green and eco-biochemical worthy of future applications to replace chemical-based surfactants.

A Review on Haematococcus pluvialis Bioprocess Optimization of Green and Red Stage Culture Conditions for the Production of Natural Astaxanthin.

As the most recognizable natural secondary carotenoid astaxanthin producer, the green microalga Haematococcus pluvialis cultivation is performed via a two-stage process. The first is dedicated to biomass accumulation under growth-favoring conditions (green stage), and the second stage is for astaxanthin evolution under various stress conditions (red stage). This mini-review discusses the further improvement made on astaxanthin production by providing an overview of recent works on H. pluvialis, including the valuable ideas for bioprocess optimization on cell growth, and the current stress-exerting strategies for astaxanthin pigment production. The effects of nutrient constituents, especially nitrogen and carbon sources, and illumination intensity are emphasized during the green stage. On the other hand, the significance of the nitrogen depletion strategy and other exogenous factors comprising salinity, illumination, and temperature are considered for the astaxanthin inducement during the red stage. In short, any factor that interferes with the cellular processes that limit the growth or photosynthesis in the green stage could trigger the encystment process and astaxanthin formation during the red stage. This review provides an insight regarding the parameters involved in bioprocess optimization for high-value astaxanthin biosynthesis from H. pluvialis.

Heterotrophic cultivation of microalgae for production of biodiesel.

High cell density cultivation of microalgae via heterotrophic growth mechanism could effectively address the issues of low productivity and operational constraints presently affecting the solar driven biodiesel production. This paper reviews the progress made so far in the development of commercial-scale heterotrophic microalgae cultivation processes. The review also discusses on patentable concepts and innovations disclosed in the past four years with regards to new approaches to microalgal cultivation technique, improvisation on the process flow designs to economically produced biodiesel and genetic manipulation to confer desirable traits leading to much valued high lipid-bearing microalgae strains.