Upcycling food waste as a low-cost cultivation medium for Chlorella sp. microalgae.

BACKGROUND: Global food loss and waste have raised environmental concerns regarding the generation of greenhouse gases (e.g., carbon dioxide and methane gas), which directly contribute to climate change. To address these concerns, the present research aims to upcycle food waste into an alternative culture medium for the cultivation of microalgae. Various parameters including pretreatment of food waste (i.e., autoclave and non-autoclave), concentration of food waste culture medium (i.e., 10%, 30%, 50%, 70%, 90% and 100%), harvesting efficiency and biochemical compounds of Chlorella sp. microalgae were carried out. RESULTS: Based on the preliminary findings, the highest biomass concentration obtained from 10% food waste culture medium in the autoclave for Chlorella sp., including strains FSP-E, ESP-31 and CY-1, were 2.869 ± 0.022, 2.385 ± 0.018 and 0.985 ± 0.0026 g L-1, respectively. Since Chlorella vulgaris FSP-E exhibited the highest biomass concentration, this microalgal strain was selected to examine the subsequent parameters. Cultivation of C. vulgaris FSP-E in 100FW achieves a biomass concentration of 4.465 ± 0.008 g L-1 with biochemical compounds of 6.94 ± 1.396, 248.24 ± 0.976 and 406.23 ± 0.593 mg g-1 for lipids, carbohydrates and proteins, respectively. CONCLUSION: This study shows that using food waste as an alternative culture medium for C. vulgaris FSP-E can achieve substantial biomass productivity and biochemical content. This research work would contribute to the concept of net zero emission and transitioning toward a circular bioeconomy by upcycling food waste as an alternative culture medium for the cultivation of microalgae. © 2024 Society of Chemical Industry.

Identification of microorganisms from fermented biowaste and the potential for wastewater treatment.

Food loss or waste is a far-reaching problem and has indeed become a worrying issue that is growing at an alarming rate. Fruits and vegetables are lost or wasted at the highest rate among the composition of food waste. Furthermore, the world is progressing toward sustainable development; hence, an efficient approach to valorise fruit and vegetable waste (FVW) is necessary. A simple phenotypic characterisation of microbiota isolated from the fermented FVW was conducted, and its effectiveness toward wastewater treatment was investigated. Presumptive identification suggested that yeast is dominant in this study, accounting for 85% of total isolates. At the genus level, the enriched medium's microbial community consists of Saccharomyces, Bacillus and Candida. Ammonium in the wastewater can enhance certain bacteria to grow, such as lactic acid bacteria, resulting in decreased NH4+ concentration at the end of the treatment to 0.5 mg/L. In addition, the fermented biowaste could reduce PO43- by 90% after the duration of treatment. Overall, FVW is a valuable microbial resource, and the microbial population enables a reduction in organic matter such as NH4+ and PO43-. This study helps explore the function and improve the effectiveness of utilising biowaste by understanding the microorganisms responsible for producing eco-enzyme.

Insect biorefinery: A circular economy concept for biowaste conversion to value-added products.

With rapid growing world population and increasing demand for natural resources, the production of sufficient food, feed for protein and fat sources and sustainable energy presents a food insecurity challenge globally. Insect biorefinery is a concept of using insect as a tool to convert biomass waste into energy and other beneficial products with concomitant remediation of the organic components. The exploitation of insects and its bioproducts have becoming more popular in recent years. This review article presents a summary of the current trend of insect-based industry and the potential organic wastes for insect bioconversion and biorefinery. Numerous biotechnological products obtained from insect biorefinery such as biofertilizer, animal feeds, edible foods, biopolymer, bioenzymes and biodiesel are discussed in the subsequent sections. Insect biorefinery serves as a promising sustainable approach for waste management while producing valuable bioproducts feasible to achieve circular bioeconomy.

Epitomizing biohydrogen production from microbes: Critical challenges vs opportunities.

Hydrogen is a clean and green biofuel choice for the future because it is carbon-free, non-toxic, and has high energy conversion efficiency. In exploiting hydrogen as the main energy, guidelines for implementing the hydrogen economy and roadmaps for the developments of hydrogen technology have been released by several countries. Besides, this review also unveils various hydrogen storage methods and applications of hydrogen in transportation industry. Biohydrogen productions from microbes, namely, fermentative bacteria, photosynthetic bacteria, cyanobacteria, and green microalgae, via biological metabolisms have received significant interests off late due to its sustainability and environmentally friendly potentials. Accordingly, the review is as well outlining the biohydrogen production processes by various microbes. Furthermore, several factors such as light intensity, pH, temperature and addition of supplementary nutrients to enhance the microbial biohydrogen production are highlighted at their respective optimum conditions. Despite the advantages, the amounts of biohydrogen being produced by microbes are still insufficient to be a competitive energy source in the market. In addition, several major obstacles have also directly hampered the commercialization effors of biohydrogen. Thus, this review uncovers the constraints of biohydrogen production from microbes such as microalgae and offers solutions associated with recent strategies to overcome the setbacks via genetic engineering, pretreatments of biomass, and introduction of nanoparticles as well as oxygen scavengers. The opportunities of exploiting microalgae as a suastainable source of biohydrogen production and the plausibility to produce biohydrogen from biowastes are accentuated. Lastly, this review addresses the future perspectives of biological methods to ensure the sustainability and economy viability of biohydrogen production.

Evaluation of Aqueous Biphasic Electrophoresis System Based on Halide-Free Ionic Liquids for Direct Recovery of Keratinase.

Aqueous biphasic electrophoresis system (ABES) incorporates electric fields into the biphasic system to separate the target biomolecules from crude feedstock. Ionic liquid (IL) is regarded as an excellent candidate as the phase-forming components for ABES because of the great electrical conductivity, which can promote the electromigration of biomolecules in ABES, and thereby enhances the separation efficiency of the target biomolecules from crude feedstock. The application of electric fields to the conventional biphasic system expedites the phase settling time of the biphasic system, which eases the subsequent scaling-up steps and reduces the overall processing time of the recovery process. Alkyl sulphate-based IL is a green and economical halide-free surfactant when compared to the other halide-containing IL. The feasibility of halide-free IL-based ABES to recover Kytococcus sedentarius TWHK01 keratinase was studied. Optimum partition coefficient (Ke = 7.53 ± 0.35) and yield (YT = 80.36% ± 0.71) were recorded with IL-ABES comprised of 15.0% (w/w) [EMIM][ESO4], 20.0% (w/w) sodium carbonate and 15% (w/w) crude feedstock. Selectivity (S) of 5.75 ± 0.27 was obtained with the IL-ABES operated at operation time of 5 min with 10 V voltage supplied. Halide-free IL is proven to be a potential phase-forming component of IL-ABES for large-scale recovery of keratinase.

Recent development in the production strategies of microbial carotenoids.

Carotenoids are a diverse group of lipid-soluble pigments that exhibit potent biological activities such as antioxidant, anti-inflammatory, and provitamin A activities. The potent health benefits of carotenoids result in the surge in the market demands for carotenoids, especially natural carotenoids from sustainable sources. Microbial carotenoids have attracted considerable interests for many industrial applications because of the low costs and ease of scaling-up with shorter production time. There is a growing interest in the search of new and sustainable microbial sources and cost-efficient production strategies following the high economical values and vast commercial applications of carotenoids. This article presents a review on the industrial production strategies of microbial carotenoids from microalgae, fungi, and bacteria sources. The industrial significance of the mass production of microbial carotenoids is also discussed. The structure, classification, and biosynthesis pathway of the carotenoids are also presented in this review.

Recent development of unconventional aqueous biphasic system: characteristics, mechanisms and applications.

Aqueous biphasic system (ABS) is widely used in the recovery, extraction, purification and separation of proteins, enzymes, nucleic acids and antibodies. The ABS with high water content and low interfacial tension offers a biocompatible environment for the recovery of labile biomolecules. Process integration can be achieved using ABS by incorporating multiple-steps of purification, concentration and purification of biomolecules in a single-step operation which often results in high product recovery yield and purity. Conventional ABS is usually formed by aqueous solutions of two polymers or a polymer and a salt above a critical concentration. The high viscosity of polymer-based ABS causes slow phase separation and hinders the mass transfer of biomolecules, whereas polymer/salt ABS is characterized by high ionic strength resulting in the loss of bioactivity of recovered biomolecules. These limitations have encouraged the development of novel ABS which is more cost-effective for various biotechnological applications. This review discusses the characteristics and mechanisms of several types of emerging unconventional ABS using phase-forming components such as hyperbranched polymers, special salts, surfactants, magnetic fields, the addition of nanoparticles and incorporation of various solvent. Moreover, several novel applications of ABS for different separation purposes such as microfluidic-based ABS, ABS bioreactors, application of ABS as an analytical tool, and ABS micropatterning are discussed in this review. In the last section of this review, a comprehensive summary of process integration using ABS for extractive fermentations, bioconversion, crystallization and precipitation is also supplemented for the comprehensive review of various types and applications of ABS in recent years.

Thermoseparating aqueous two-phase systems: Recent trends and mechanisms.

Having the benefits of being environmentally friendly, providing a mild environment for bioseparation, and scalability, aqueous two-phase systems (ATPSs) have increasingly caught the attention of industry and researchers for their application in the isolation and recovery of bioproducts. The limitations of conventional ATPSs give rise to the development of temperature-induced ATPSs that have distinctive thermoseparating properties and easy recyclability. This review starts with a brief introduction to thermoseparating ATPSs, including its history, unique characteristics and advantages, and lastly, key factors that influence partitioning. The underlying mechanism of temperature-induced ATPSs is covered together with a summary of recent applications. Thermoseparating ATPSs have been proven as a solution to the demand for economically favorable and environmentally friendly industrial-scale bioextraction and purification techniques.

Upcycling fruit waste into microalgae biotechnology: Perspective views and way forward.

Fruit and vegetable wastes are linked to the depletion of natural resources and can pose serious health and environmental risks (e.g. eutrophication, water and soil pollution, and GHG emissions) if improperly managed. Current waste management practices often fail to recover high-value compounds from fruit wastes. Among emerging valorization methods, the utilization of fruit wastes as a feedstock for microalgal biorefineries is a promising approach for achieving net zero waste and sustainable development goals. This is due to the ability of microalgae to efficiently sequester carbon dioxide through photosynthesis, utilize nutrients in wastewater, grow in facilities located on non-arable land, and produce several commercially valuable compounds with applications in food, biofuels, bioplastics, cosmetics, nutraceuticals, pharmaceutics, and various other industries. However, the application of microalgal biotechnology towards upcycling fruit wastes has yet to be implemented on the industrial scale due to several economic, technical, operational, and regulatory challenges. Here, we identify sources of fruit waste along the food supply chain, evaluate current and emerging fruit waste management practices, describe value-added compounds in fruit wastes, and review current methods of microalgal cultivation using fruit wastes as a fermentation medium. We also propose some novel strategies for the practical implementation of industrial microalgal biorefineries for upcycling fruit waste in the future.

In vitro evidence of chain transfer to tetraethylene glycols in enzymatic polymerization of polyhydroxyalkanoate.

A polyhydroxyalkanoate (PHA) was enzymatically synthesized in vitro, and the end structure of PHA associated with a chain transfer (CT) reaction was investigated. In the CT reaction, PHA chain transfers from PHA synthase (PhaC) to a CT agent, resulting in covalent bonding of CT agent to the PHA chain at its carboxyl end. In vitro CT reaction has never been demonstrated because of relatively low yields of in vitro synthesized poly[(R)-3-hydroxybutyrate)] (P(3HB)), which makes it difficult to characterize the end structures of the polymers by nuclear magnetic resonance (NMR). To overcome these difficulties, a novel in vitro synthesis method that produced relatively larger amounts of P(3HB) was developed by employing PhaCDa from Delftia acidovorans and two enantioselective enoyl-coenzyme A (CoA) hydratases which were R-hydratase (PhaJAc) from Aeromonas caviae and S-hydratase (FadB1x) from Pseudomonas putida KT2440 with ß-butyrolactone and CoA as starting materials. Using this method, P(3HB) synthesis was performed with tetraethylene glycols (TEGs) as a discriminable CT agent, and the resultant P(3HB) was characterized by (1)H-NMR. NMR analysis revealed that the carboxylic end of P(3HB) was covalently linked to TEGs, providing the first direct evidence of in vitro CT reaction.

Functional ginger extracts from supercritical fluid carbon dioxide extraction via in vitro and in vivo assays: antioxidation, antimicroorganism, and mice xenografts models.

Supercritical fluid carbon dioxide extraction technology was developed to gain the active components from a Taiwan native plant, Zingiber officinale (ginger). We studied the biological effects of ginger extracts via multiple assays and demonstrated the biofunctions in each platform. Investigations of ginger extracts indicated antioxidative properties in dose-dependant manners on radical scavenging activities, reducing powers and metal chelating powers. We found that ginger extracts processed moderate scavenging values, middle metal chelating levels, and slight ferric reducing powers. The antibacterial susceptibility of ginger extracts on Staphylococcus aureus, Streptococcus sobrinus, S. mutans, and Escherichia coli was determined with the broth microdilution method technique. The ginger extracts had operative antimicroorganism potentials against both Gram-positive and Gram-negative bacteria. We further discovered the strong inhibitions of ginger extracts on lethal carcinogenic melanoma through in vivo xenograft model. To sum up, the data confirmed the possible applications as medical cosmetology agents, pharmaceutical antibiotics, and food supplements.

Expression of His-tagged NADPH-dependent acetoacetyl-CoA reductase in recombinant Escherichia coli BL-21(DE3).

Poly-3-hydroxybutyrate (P(3HB)), a member of the polyhydroxyalkanoate (PHA) family, is a biodegradable polyester with diverse industrial applications. NADPH-dependent acetoacetyl-CoA reductase (phaB) is the enzyme which plays an essential role in P(3HB) synthesis by catalyzing the conversion of the intermediates. The expression of phaB enzyme using the recombinant Escherichia coli BL-21(DE3) and the purification of the synthesized enzyme were studied. The pET-B3 plasmid harbouring the phaB gene derived from Ralstonia eutropha H16, was driven by the lac promoter in E. coli BL-21(DE3). The enzyme was expressed with different induction time, temperatures and cell age. Results showed that the cell age of 4 h, induction time of 12 h at 37°C were identified as the optimal conditions for the enzyme reductase expression. A specific activity of 0.151 U mg-1 protein and total protein concentration of 0.518 mg mg-1 of dry cell weight (DCW) were attained. Affinity chromatography was performed to purify the His-tagged phaB enzyme, in which enhanced the specific activity (14.44 U mg-1) and purification fold (38-fold), despite relative low yield (44.6%) of the enzyme was obtained. The purified phaB showed an optimal enzyme activity at 30°C and pH 8.0. The findings provide an alternative for the synthesis of the reductase enzyme which can be used in the industrial-scale production of the biodegradable polymers.

Tropical Seaweeds as a Sustainable Resource Towards Circular Bioeconomy: Insights and Way Forward.

Seaweeds are photosynthetic marine macroalgae known for their rapid biomass growth and their significant contributions to global food and feed production. Seaweeds play a crucial role in mitigating various environmental issues, including greenhouse gases, ocean acidification, hypoxia, and eutrophication. Tropical seaweeds are typically found in tropical and subtropical coastal zones with warmer water temperatures and abundant sunlight. These tropical seaweeds are rich sources of proteins, vitamins, minerals, fibers, polysaccharides, and bioactive compounds, contributing to their health-promoting properties and their diverse applications across a range of industries. The productivity, cultivability, nutritional quality, and edibility of tropical seaweeds have been well-documented. This review article begins with an introduction to the growth conditions of selected tropical seaweeds. Subsequently, the multifunctional properties of tropical seaweeds including antioxidant and anti-inflammatory, anti-coagulant, anti-carcinogenic and anti-proliferative, anti-viral, therapeutic and preventive properties were comprehensively evaluated. The potential application of tropical seaweeds as functional foods and feeds, as well as their contributions to sustainable cosmetics, bioenergy, and biofertilizer production were also highlighted. This review serves as a valuable resource for researchers involved in seaweed farming as it provides current knowledge and insights into the cultivation and utilization of seaweeds.

Enhancement of protein production using synthetic brewery wastewater by Haematococcus pluvialis.

Microalgae is a sustainable protein source that has been widely applied in animal feeds, functional foods, pharmaceutical, and cosmeceutical industries. Waste products could be a potential cost-saving and nutrient-rich substrate in the cultivation of microalgae for protein production. This study aims to investigate the cultivation condition of Haematococcus pluvialis for protein synthesis using synthetic brewery wastewater (BW). H. pluvialis was cultivated in the Bold's Basal Medium (BBM) mixed with synthetic BW at different concentrations. Various cultivation conditions including brewer's spent grain hydrolysate (BSGH) concentrations, pH, and light sources were studied. The molecular weight, amino acids profile and antioxidant activity of synthesized protein were determined. Fed-batch cultivation using different percentages of fresh medium replacement for enhancing protein production was investigated. The 20% fed-batch cultivation reached 27 × 105 ± 0.42 cells/mL, and 4-fold of the protein content of 64.93 ± 5.30% of dry weight was recorded on day-13. Seven essential amino acids (lysine, threonine, histidine, phenylalanine, isoleucine, leucine, methionine) were produced in fed-batch cultivation. Red LED obtained the highest DPPH radical scavenging activity of 27.47 ± 0.98%. The findings suggested that BW is a promising substrate in the cultivation of H. pluvialis to commercially produce protein for numerous industrial applications.

Efficacy of alcohol/sugar aqueous biphasic system on partition of bovine serum albumin.

The efficacy of alcohol/sugar aqueous biphasic (ABS) system on protein extraction was investigated. A model protein, bovine serum albumin (BSA), was adopted to evaluate the effects of types and concentration of phase-forming components, protein concentration, and system pH on the protein partition efficiency. The 1-propanol/maltose ABS exhibited an overall better partition efficiency of BSA to the alcohol-rich top phase. A maximum partition coefficient (K) of 20.01 ± 0.05 and recovery yield (Y) of 95.42% ± 0.01% of BSA were achieved with 35% (w/w) 1-propanol/22% (w/w) maltose ABS at pH 5.0 for 10% (w/w) BSA load. The K and Y of BSA in 1-propanol/maltose ABS was slightly improved with the addition of 3% (w/w) of ionic liquid, 1-butyl-3-methylimidazolium bromide ([Bmim]Br) as the adjuvant that could provide protein stabilizing effect. The Fourier Transform Infrared Spectrum (FTIR) analysis revealed that the protein structure remained unaltered upon the separation process.

Enhanced recovery of astaxanthin from recombinant Kluyveromyces marxianus with ultrasonication-assisted alcohol/salt aqueous biphasic system.

Microbial astaxanthin with strong antioxidant activity is greatly demanded for diverse applications. Extractive disruption in aqueous biphasic system (ABS) integrates the cells disruption and biomolecules recovery processes in one-step operation, allowing the direct recovery of intracellular biomolecules with biphasic system upon released from cells. In this study, astaxanthin was recovered from recombinant Kluyveromyces marxianus yeast cells via extractive disruption using alcohol/salt ABS. Recombinant K. marxianus yeast is engineered to produce high concentration of free form astaxanthin. Highest partition coefficient (K = 90.02 ± 2.25) and yield (Y = 96.80% ± 0.05) of astaxanthin were obtained with ABS composed of 20% (w/w) 1-propanol and 20% (w/w) sodium citrate of pH 5, 0.5% (w/w) yeast cells loading and additional of 1% (w/w) 1-butyl-3-methylimidazolium tetrafluoroborate (Bmim)BF4 to improve the migration of astaxanthin to alcohol-rich top phase. The incorporation of 2.5 h of ultrasonication to the biphasic system further enhanced the astaxanthin recovery in ABS. The direct recovery of astaxanthin from recombinant K. marxianus cells was demonstrated with the ultrasonication-assisted alcohol/salt ABS which integrates the extraction and concentration of astaxanthin in a single-step operation.

Characterization of alcohol/salt aqueous two-phase system for optimal separation of gallic acids.

Gallic acid (GA) is a hydrophilic polyphenol which is noteworthy for strong antioxidant capacity. The drawbacks of conventional extraction approaches such as time-consuming and high processing cost are often viewed as a hurdle to extract GA from plant sources in industrial scale. Aqueous two-phase system (ATPS) is a separation approach which can be employed as an alternative to the conventional approaches. The partition behaviour of GA in an alcohol/salt ATPS was investigated in this study to aid the development of industrial scale ATPS to extract GA from natural sources. The separation of GA was characterized by determining the types of alcohol and salt, phase composition, sample load, pH of the system and addition of adjuvants applied in the alcohol/salt ATPS construction. The hydrophilic GA was targeted to the salt-rich phase of the alcohol/salt ATPS with a partition coefficient (KGA) of 25.00 ± 0.00. The optimum condition of ATPS for the maximum partition of GA was achieved in ATPS comprised of 24% (w/w) 1-propanol and 22% (w/w) phosphate salt at pH 8 with 5% (w/w) of 1 mg/mL sample loading and 2% (w/w) NaCl addition. The findings suggest that ATPS can be applied for separation of GA from various natural sources.

Primary purification of intracellular Halomonas salina ectoine using ionic liquids-based aqueous biphasic system.

Ectoine is a zwitterionic amino acid derivative that can be naturally sourced from halophilic microorganisms. The increasing demands of ectoine in various industries have urged the researches on the cost-effective approaches on production of ectoine. Ionic liquids-based aqueous biphasic system (ILABS) was applied to recover Halomonas salina ectoine from cells hydrolysate. The 1-butyl-3-methylimidazolium tetrafluoroborate (Bmim)BF4 was used in the ILABS and the recovery efficiency of ILABS to recover ectoine from H. salina cells lysate was evaluated by determining the effects of phase composition; pHs; crude loading and additional neutral salt (NaCl). The hydrophilic ectoine was targeted to partition to the hydrophilic salt-rich phase. A total yield (YB) of 96.32% ± 1.08 of ectoine was obtained with ILABS of phase composition of 20% (w/w) (Bmim)BF4 and 30% (w/w) sulfate salts; system pH of 5.5 when the 20% (w/w) of crude feedstock was applied to the ILABS. There was no significant enhancement on the ectoine recovery efficiency using the ILABS when NaCl was added, therefore the ILABS composition without the additional neutral salt was recommended for the primary purification of ectoine. Partition coefficient (KE) of 30.80 ± 0.42, purity (PE) of 95.82% and enrichment factor (Ef) of 1.92 were recorded with the optimum (Bmim)BF4/sulfate ILABS. These findings have provided an insight on the feasibility of recovery of intracellular biomolecules using the green solvent-based aqueous system in one single-step operation.

Exploring the fermentation characteristics of a newly isolated marine bacteria strain, Gordonia terrae TWRH01 for carotenoids production.

Carotenoids serve as one of the most important group of naturally-occurring lipid-soluble pigments which exhibit great biological activities such as antioxidant, anti-inflammatory and provitamin A activities. Owing to their advantageous health effects, carotenoids are widely applied in various industries. Microbial carotenoids synthesis therefore has attracted increasing attention in recent years. In the present study, a marine microorganism originally isolated from seawater in northern Taiwan was determined to be a strain of Gordonia terrae based on its 16S rRNA gene sequence. The strain G. terrae TWRH01 has the ability to synthesize and accumulate the intracellular pigments was identified by gas chromatography-mass spectrometry (GC-MS). The biochemical production characteristics of this strain were studied by employing different fermentation strategies. Findings suggested that G. terrae TWRH01 can actively grow and efficiently synthesize carotenoids in medium adjusted to pH 7 containing 16 g L-1 sucrose as the carbon source, 16 g L-1 yeast extract as the nitrogen source, 0.6 M NaCl concentration, and supplemented with 0.45% (v/v) 1 M CaCl2. Results revealed that the optimization of fermentation yielded 15.29 g L-1 dry biomass and 10.58 µmol L-1 relative ß-carotene concentration. According to GC-MS analysis, the orange-red colored pigments produced were identified as carotenoid derivatives, mainly echinenone and adonixanthin 3'-ß-d-glucoside. Therefore, the new bacterial strain showed a highly potential bioresource for the commercial production of natural carotenoids.

Partition efficiency of cytochrome c with alcohol/salt aqueous biphasic flotation system.

Aqueous biphasic flotation (ABF) integrates aqueous biphasic system (ABS) and solvent sublation for recovery of target biomolecules. The feasibility of the alcohol/salt ABF for exclusive partition of cytochrome c to one specific phase of the system was investigated. Aliphatic alcohols of different carbon chain length (ethanol, 1-propanol and 2-propanol) and salts (sulfate, phosphate and citrate) were used for the phase formation. The effects of phase composition, concentration of sample loading, pH, flotation time and flow rate of the system on the partition efficiency of cytochrome c were determined. Cytochrome c was exclusively partitioned to the alcohol-rich top phase of the ABF of 18% (w/w) ethanol and 26% (w/w) ammonium sulfate with pH 6 and 20% (w/w) of sample loading. Highest partition coefficient (K) of 6.85 ± 0.21 and yield (YT) of 99.40% ± 0.02 were obtained with optimum flotation rate of 10 mL/min and flow rate of 10 min.