Human skin pigmentation is a result of constitutive and facultative pigmentation. Facultative pigmentation is frequently stimulated by UV radiation, pharmacologic drugs, and hormones whereby leads to the development of abnormal skin hyperpigmentation. To date, many state-of-art depigmenting compounds have been studied using in vitro model to treat hyperpigmentation problems for cosmetic dermatological applications; little attention has been made to compare the effectiveness of these depigmenting compounds and their mode of actions. In this present article, new and recent depigmenting compounds, their melanogenic pathway targets, and modes of action are reviewed. This article compares the effectiveness of these new depigmenting compounds to modulate several melanogenesis-regulatory enzymes and proteins such as tyrosinase (TYR), TYR-related protein-1 (TRP1), TYR-related protein-2 (TRP2), microphthalmia-associated transcription factor (MITF), extracellular signal-regulated kinase (ERK) and N-terminal kinases (JNK) and mitogen-activated protein kinase p38 (p38 MAPK). Other evidences from in vitro assays such as inhibition on melanosomal transfer, proteasomes, nitric oxide, and inflammation-induced melanogenesis are also highlighted. This article also reviews analytical techniques in different assays performed using in vitro model as well as their advantages and limitations. This article also provides an insight on recent finding and re-examination of some protocols as well as their effectiveness and reliability in the evaluation of depigmenting compounds. Evidence and support from related patents are also incorporated in this present article to give an overview on current patented technology, latest trends, and intellectual values of some depigmenting compounds and protocols, which are rarely highlighted in the literatures.
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.
The depigmenting effect of kojic acid esters synthesized by the esterification of kojic acid using Rhizomucor miehei immobilized lipase was investigated in B16F1 melanoma cells. The depigmenting effect of kojic acid and kojic acid esters was evaluated by the inhibitory effect of melanin formation and tyrosinase activity on alpha-stimulating hormone- (α-MSH-) induced melanin synthesis in B16F1 melanoma cells. The cellular tyrosinase inhibitory effect of kojic acid monooleate, kojic acid monolaurate, and kojic acid monopalmitate was found similar to kojic acid at nontoxic doses ranging from 1.95 to 62.5 μg/mL. However, kojic acid monopalmitate gave slightly higher inhibition to melanin formation compared to other inhibitors at doses ranging from 15.63 to 62.5 μg/mL. Kojic acid and kojic acid esters also show antioxidant activity that will enhance the depigmenting effect. The cytotoxicity of kojic acid esters in B16F1 melanoma cells was significantly lower than kojic acid at high doses, ranging from 125 and 500 μg/mL. Since kojic acid esters have lower cytotoxic effect than kojic acid, it is suggested that kojic acid esters can be used as alternatives for a safe skin whitening agent and potential depigmenting agents to treat hyperpigmentation.
The use of biodegradable material such as simple carbohydrates and recyclable material such as thermo-sensitive polymers is in need to develop a sustainable aqueous two-phase system (ATPS) for the purification of biomolecules. Accurate determination of sucrose concentration is important in liquid-liquid equilibrium (LLE) study of carbohydrate-based ATPS. The well-established phenol-sulfuric acid method has been widely employed in the measurement of carbohydrate concentration. However, the presence of thermo-sensitive polymers, which has a lower critical solution temperature (LCST) below room temperature, in carbohydrate samples could hamper the precision of spectrophotometric analysis due to the formation of two phases or cloudiness in the sample. Thus, the following modifications were made in an attempt to eliminate the interference occurred during conventional phenol-sulfuric acid assay.•The modified assay for sucrose quantification was performed at an ice-cold temperature throughout the reaction in order to avoid the interference from thermo-sensitive polymers.•This method required a sample volume of 3 μL and hence the volume of other reagents employed was also considerably reduced.•The absorbance was measured at 520 nm which allowed a longer linearity range (0.05-7.5%, w/v).
The potential use of n-dodecane and n-hexadecane as oxygen vectors for enhancing hyaluronic acid (HA) biosynthesis by Streptococcus zooepidemicus ATCC 39920 was investigated using a 2-L stirred-tank bioreactor equipped with helical ribbon or Rushton turbine impellers. The volumetric fraction of the oxygen vector influenced the gas-liquid volumetric oxygen transfer coefficient (K(L)a) positively. Batch HA fermentation with 1% (v/v) n-dodecane or 0.5% (v/v) n-hexadecane addition was carried out at different impeller tip speeds. Even though cell growth was lower in the fermentation with oxygen vector addition, the HA productivity and molecular weight were higher when compared to the fermentation without oxygen vector at low impeller tip speed. The highest HA concentration (4.25 gHA/l) and molecular weight (1.54 × 10(7) Da) were obtained when 0.5% (v/v) n-hexadecane and 0.785 m/s impeller tip speed of helical ribbon were used.
The use of pullulanase (EC 3.2.1.41) has recently been the subject of increased applications in starch-based industries especially those aimed for glucose production. Pullulanase, an important debranching enzyme, has been widely utilised to hydrolyse the α-1,6 glucosidic linkages in starch, amylopectin, pullulan, and related oligosaccharides, which enables a complete and efficient conversion of the branched polysaccharides into small fermentable sugars during saccharification process. The industrial manufacturing of glucose involves two successive enzymatic steps: liquefaction, carried out after gelatinisation by the action of α-amylase; saccharification, which results in further transformation of maltodextrins into glucose. During saccharification process, pullulanase has been used to increase the final glucose concentration with reduced amount of glucoamylase. Therefore, the reversion reaction that involves resynthesis of saccharides from glucose molecules is prevented. To date, five groups of pullulanase enzymes have been reported, that is, (i) pullulanase type I, (ii) amylopullulanase, (iii) neopullulanase, (iv) isopullulanase, and (v) pullulan hydrolase type III. The current paper extensively reviews each category of pullulanase, properties of pullulanase, merits of applying pullulanase during starch bioprocessing, current genetic engineering works related to pullulanase genes, and possible industrial applications of pullulanase.
The effect of different carbon and nitrogen sources on the production of mannan-degrading enzymes, focussing on β-mannanase, by Aspergillus niger was investigated using shake flask culture. The β-mannanase activity obtained during growth of A. niger on guar gum (GG, 1495 nkat mL(-1)) was much higher than those observed on other carbon substrates, locust bean gum (1148 nkat mL(-1)), α-cellulose (10.7 nkat mL(-1)), glucose (8.8 nkat mL(-1)) and carboxymethylcellulose (4.6 nkat mL(-1)). For fermentation using GG as a carbon source, bacteriological peptone gave the highest β-mannanase activity (1744 nkat mL(-1)) followed by peptone from meat (1168 nkat mL(-1)), yeast extract (817 nkat mL(-1)), ammonium sulphate (241 nkat mL(-1)), ammonium nitrate (113 nkat mL(-1)) and ammonium chloride (99 nkat mL(-1)) when used as a nitrogen source. The composition of bacteriological peptone and initial pH of the medium were further optimized using response surface methodology (RSM). Medium consisted of 21.3 g L(-1) GG and 57 g L(-1) peptone with initial culture pH of 5.5 was optimum for β-mannanase production (2063 nkat mL(-1)) by A. niger. The β-mannanase production obtained in this study using A. niger was significantly higher than those reported in the literature.
This study was undertaken to optimize yeast extract, glucose, and vitamin concentrations; and also culture pH for maximizing the growth of a probiotic bacterium, Lactobacillus rhamnosus, and to assess the effects of these factors by using response surface methodology. A central composite design was used as an experimental design for the allocation of treatment combinations. A polynomial regression model with cubic and quartic terms was used for analysis of the experimental data. It was found that the effects involving yeast extract, glucose, vitamins and pH on the growth of L. rhamnosus were significant, and the strongest effect was given by the yeast extract concentration. Estimated optimum conditions of the factors for the growth of L. rhamnosus are as follows: pH=6.9; vitamin solution=1.28% (v/v); glucose=5.01% (w/v) and yeast extract=6.0% (w/v).
In this study, the methods for extraction and purification of miraculin from Synsepalum dulcificum were investigated. For extraction, the effect of different extraction buffers (phosphate buffer saline, Tris-HCl and NaCl) on the extraction efficiency of total protein was evaluated. Immobilized metal ion affinity chromatography (IMAC) with nickel-NTA was used for the purification of the extracted protein, where the influence of binding buffer pH, crude extract pH and imidazole concentration in elution buffer upon the purification performance was explored. The total amount of protein extracted from miracle fruit was found to be 4 times higher using 0.5M NaCl as compared to Tris-HCl and phosphate buffer saline. On the other hand, the use of Tris-HCl as binding buffer gave higher purification performance than sodium phosphate and citrate-phosphate buffers in IMAC system. The optimum purification condition of miraculin using IMAC was achieved with crude extract at pH 7, Tris-HCl binding buffer at pH 7 and the use of 300 mM imidazole as elution buffer, which gave the overall yield of 80.3% and purity of 97.5%. IMAC with nickel-NTA was successfully used as a single step process for the purification of miraculin from crude extract of S. dulcificum.
Research was undertaken to investigate the treatment of fishery washing water using Bacillus sphaericus, and to recover the spores for subsequent use as bioinsecticide to control the population of mosquitoes. This treatment method could reduce pollution due to organic matter by decreasing the value of Chemical Oxygen Demand (COD) and Biological Oxygen Demand (BOD) by about 85% and 92%, respectively. The maximum concentration of spores (83.3 x 10(7) spores ml(-1)) using normal concentration of filtered fishery washing water was only about 27% lower than that obtained in fermentation using 0.25% (w/v) yeast extract. The larvicidal activity of the spores produced in fermentation using fishery washing water to Culex quinquefaciatus, as measured by LD50 after 48 h, was almost the same as the larvicidal activity of spores obtained from fermentation using yeast extract.
The effects of medium strategy, number of impellers, aeration mode, and mode of operation on Morinda elliptica cell suspension cultures in a stirred-tank bioreactor are described. A lower number of impellers and continuous aeration contributed toward high cell growth rate, whereas a higher number of impellers reduced cell growth rate, although not anthraquinone yield. The semicontinuous mode could indirectly imitate the larger scale version of production medium strategy and improved anthraquinone production even with 0. 012% (v/v) antifoam addition. Production medium promoted both growth (maximum dry cell weight of 24.6 g/L) and anthraquinone formation (maximum content of 19.5 mg/g of dry cell weight), without any necessity for antifoam addition. Cultures in production medium or with higher growth rate and anthraquinone production were less acidic than cultures in growth medium or with lower growth rate and anthraquinone production. Using the best operating variables, growth of M. elliptica cells (24.6 g/L) and anthraquinone yield (0.25 g/L) were 45% and 140%, respectively, lower than those using a shake flask culture after 12 days of cultivation.
Silver nanoparticles (AgNPs) have been found to have extensive biomedical and biological applications. They can be synthesised using chemical and biological methods, and coated by polymer to enhance their stability. Hence, the changes in the physico-chemical characteristics of AgNPs must be scrutinised due to their importance for biological activity. The UV-Visible absorption spectra of polyethylene glycol (PEG) -coated AgNPs displayed a distinctive narrow peak compared to uncoated AgNPs. In addition, High-Resolution Transmission Electron Microscopy analysis revealed that the shapes of all AgNPs, were predominantly spherical, triangular, and rod-shaped. Fourier-Transform Infrared Spectroscopy analysis further confirmed the role of PEG molecules in the reduction and stabilisation of the AgNPs. Moreover, dynamic light scattering analysis also revealed that the polydispersity index values of PEG-coated AgNPs were lower than the uncoated AgNPs, implying a more uniform size distribution. Furthermore, the uncoated and PEG-coated biologically synthesised AgNPs demonstrated antagonisms activities towards tested pathogenic bacteria, whereas no antagonism activity was detected for the chemically synthesised AgNPs. Overall, generalisation on the interrelations of synthesis methods, PEG coating, characteristics, and antimicrobial activity of AgNPs were established in this study.
In a conventional protein downstream processing (DSP) scheme, chromatography is the single most expensive step. Despite being highly effective, it often has a low process throughput due to its semibatch nature, sometimes with nonreproducible results and relatively complex process development. Hence, more work is required to develop alternative purification methods that are more cost-effective, but exhibiting nearly comparable performance. In recent years, surfactant precipitation has been heralded as a promising new method for primary protein recovery that meets these criteria and is a simple and cost-effective method that purifies and concentrates. The method requires the direct addition of a surfactant to a complex solution (e.g. a fermentation broth) containing the protein of interest, where the final surfactant concentration is maintained below its critical micelle concentration (CMC) in order to allow for electrostatic and hydrophobic interactions between the surfactant and the target protein. An insoluble (hydrophobic) protein-surfactant complex is formed and backextraction of the target protein from the precipitate into a new aqueous phase is then carried out using either solvent extraction, or addition of a counter-ionic surfactant. Importantly, as highlighted by past researchers, the recovered proteins maintain their activity and structural integrity, as determined by circular dichroism (CD). In this review, various aspects of surfactant precipitation with respect to its general methodology and process mechanism, system parameters influencing performance, protein recovery, process selectivity and process advantages will be highlighted. Moreover, comparisons will be made to reverse micellar extraction, and the current drawbacks/challenges of surfactant precipitation will also be discussed. Finally, promising directions of future work with this separation technique will be highlighted.
Bacteriocin is a proteinaceous biomolecule produced by bacteria (both Gram-positive and Gram-negative) that exhibits antimicrobial activity against closely related species, and food-borne pathogens. It has recently gained importance and attracted the attention of several researchers looking to produce it from various substrates and bacterial strains. This ushers in a new era of food preservation where the use of bacteriocin in food products will be an alternative to chemical preservatives, and heat treatment which are understood to cause unwanted side effects, and reduce sensory and nutritional quality. However, this new market depends on the success of novel downstream separation schemes from various types of crude feedstocks which are both effective and economic. This review focuses on the downstream separation of bacteriocin from various sources using both conventional and novel techniques. Finally, recommendations for future interesting areas of research that need to be pursued are highlighted.
Bacteriocin is an important peptide which can be used as an anti-microbial agent in food. However, simpler and more cost-effective purification methods need to be developed compared to chromatography to enhance its commercial viability. Surfactant precipitation was employed for the first time to purify bacteriocin-like inhibitory substance (BLIS) from a fermentation broth of Pediococcus acidilactici Kp10, and the amount precipitated was investigated as a function of anionic surfactant (AOT) concentration, and pH. Protein recovery from the precipitate was accomplished using solvent extraction, and solvent type, NaCl concentration, and ionic strength of the final solution were optimised. Optimal conditions were; 1.05mM of AOT at pH 4 for precipitation, and acetone extraction (with 1mM NaCl), which resulted in an 86.3% yield, and 53.8 purification factor. This study highlighted the fact that surfactant precipitation can be used as a primary recovery method for BLIS from a complex fermentation broth.
Direct conversion of gelatinized sago starch into kojic acid by Aspergillus flavus strain having amylolytic enzymes was carried out at two different scales of submerged batch fermentation in a 250-mL shake flask and in a 50-L stirred-tank fermentor. For comparison, fermentations were also carried out using glucose and glucose hydrolyzate from enzymic hydrolysis of sago starch as carbon sources. During kojic acid fermentation of starch, starch was first hydrolyzed to glucose by the action of alpha-amylase and glucoamylase during active growth phase. The glucose remaining during the production phase (non-growing phase) was then converted to kojic acid. Kojic acid production (23.5 g/L) using 100 g/L sago starch in a shake flask was comparable to fermentation of glucose (31.5 g/L) and glucose hydrolyzate (27.9 g/L) but in the 50-L fermentor was greatly reduced due to non-optimal aeration conditions. Kojic acid production using glucose was higher in the 50-L fermentor than in the shake flask.
Lactic acid bacteria are industrially important microorganisms recognized for their fermentative ability mostly in their probiotic benefits as well as lactic acid production for various applications. Nevertheless, lactic acid fermentation often suffers end-product inhibition which decreases the cell growth rate. The inhibition of lactic acid is due to the solubility of the undissociated lactic acid within the cytoplasmic membrane and insolubility of dissociated lactate, which causes acidification of cytoplasm and failure of proton motive forces. This phenomenon influences the transmembrane pH gradient and decreases the amount of energy available for cell growth. In general, the restriction imposed by lactic acid on its fermentation can be avoided by extractive fermentation techniques, which can also be exploited for product recovery.
Lactic acid bacteria are industrially important microorganisms recognized for fermentative ability mostly in their probiotic benefits as well as lactic acid production for various applications. Fermentation conditions such as concentration of initial glucose in the culture, concentration of lactic acid accumulated in the culture, types of pH control strategy, types of aeration mode and different agitation speed had influenced the cultivation performance of batch fermentation of Pediococcus acidilactici. The maximum viable cell concentration obtained in constant fed-batch fermentation at a feeding rate of 0.015 L/h was 6.1 times higher with 1.6 times reduction in lactic acid accumulation compared to batch fermentation. Anion exchange resin, IRA 67 was found to have the highest selectivity towards lactic acid compared to other components studied. Fed-batch fermentation of P. acidilactici coupled with lactic acid removal system using IRA 67 resin showed 55.5 and 9.1 times of improvement in maximum viable cell concentration compared to fermentation without resin for batch and fed-batch mode respectively. The improvement of the P. acidilactici growth in the constant fed-batch fermentation indicated the use of minimal and simple process control equipment is an effective approach for reducing by-product inhibition. Further improvement in the cultivation performance of P. acidilactici in fed-bath fermentation with in situ addition of anion-exchange resin significantly helped to enhance the growth of P. acidilactici by reducing the inhibitory effect of lactic acid and thus increasing probiotic production.
The study is to identify the extraction of intracellular inulinase (exo- and endoinulinase) and invertase as well as optimization medium composition for maximum productions of intra- and extracellular enzymes from Aspergillus niger ATCC 20611. From two different methods for extraction of intracellular enzymes, ultrasonic method was found more effective. Response surface methodology (RSM) with a five-variable and three-level central composite design (CCD) was employed to optimize the medium composition. The effect of five main reaction parameters including sucrose, yeast extract, NaNO₃, Zn⁺², and Triton X-100 on the production of enzymes was analyzed. A modified quadratic model was fitted to the data with a coefficient of determination (R²) more than 0.90 for all responses. The intra-extracellular inulinase and invertase productions increased in the range from 16 to 8.4 times in the optimized medium (10% (w/v) sucrose, 2.5% (w/v) yeast extract, 2% (w/v) NaNO₃, 1.5 mM (v/v) Zn⁺², and 1% (v/v) Triton X-100) by RSM and from around 1.2 to 1.3 times greater than in the medium optimized by one-factor-at-a-time, respectively. The results of bioprocesses optimization can be useful in the scale-up fermentation and food industry.