Photoswitchable carbon-dot liposomes mediate catalytic cascade reactions for amplified dynamic treatment of tumor cells.

Most biochemical reactions that occur in living organisms are catalyzed by a series of enzymes and proceed in a tightly controlled manner. The development of artificial enzyme cascades that resemble multienzyme complexes in nature is of current interest due to their potential in various applications. In this study, a nanozyme based on photoswitchable carbon-dot liposomes (CDsomes) was developed for use in programmable catalytic cascade reactions. These CDsomes prepared from triolein are amphiphilic and self-assemble into liposome-like structures in an aqueous environment. CDsomes feature excitation-dependent photoluminescence and, notably, can undergo reversible switching between a fluorescent on-state and nonfluorescent off-state under different wavelengths of light irradiation. This switching ability enables the CDsomes to exert photocatalytic oxidase- and peroxidase-like activities in their on- (bright) and off- (dark) states, respectively, resulting in the conversion of oxygen molecules into hydrogen peroxide (H2O2), followed by the generation of active hydroxyl radicals (OH). The two steps of oxygen activation can be precisely controlled in a sequential manner by photoirradiation at different wavelengths. Catalytic reversibility also enables the CDsomes to produce sufficient reactive oxygen species (ROS) to effectively kill tumor cells. Our results reveal that CDsomes is a promising photo-cycling nanozyme for precise tumor phototherapy through regulated programmable cascade reactions.

Anti-oxidative and anti-hyperglycemic properties of Agastache foeniculum essential oil and oily fraction in hyperglycemia-stimulated and lipopolysaccharide-stimulated macrophage cells: In vitro and in silico studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Hyperglycemia (HG) and lipopolysaccharide (LPS) often promote superoxide accumulation, which may increase oxidative stress. Reducing superoxide production in hyperglycemia and the inflammatory condition is an emerging way to reduce protein and lipid oxidation and diabetes complication. AIM OF STUDY: To examine the effect of Agastache foeniculum essential oil (AFEO) and oil fraction (AFoil) on HG- and LPS-stimulated oxidative stress, the pathogenicity of AFEO and AFoil on oxidative stress was assessed. METHODS: The stimulatory effects of AFEO and AFoil on the activity and expression of NADH oxide (NOX), catalase (CAT), superoxide dismutase (SOD), and the expression of nuclear respiratory factor 2 (NRF2) and nuclear factor-kappa B (NF-kB) in the stimulated macrophage cell line, J774.A1, was studied. The interaction patterns of AFEO and AFoil components with NOX, SOD, CAT, NRF2, and NF-kB proteins were also deduced using molecular docking. RESULTS: Estragole was the main ingredient in AFEO (97%). Linolenic acid (32.10%), estragole (16.22%), palmitic acid (12.62%), linoleic acid (12.04%), and oleic acid (8.73%) were the major chemical components of the AFoil. NOX activation was stimulated in macrophage cells by HG and LPS. At 20 µg/mL, AFEO and AFoil decreased NOX activity while increased SOD and CAT activities in stimulated macrophages. AFoil with estragole and omega-3 fatty acids was better than AFEO with estragole in anti-hyperglycemic and anti-oxidative activity. According to molecular docking research, estragole, linoleic acid, and linolenic acid bind to different hydrophobic pockets of NOX, SOD, CAT, NFR2, and NF-kB using hydrogen bonds, van der Waals bonds, pi-alkyl, and pi-anion interactions, with different binding energies. CONCLUSION: AFEO and AFoil showed antioxidant and anti-diabetic activity. The mechanisms in lowering oxidative stress markers depended on down-regulating superoxide-producing enzymes and up-regulating superoxide-removing enzymes at gene and protein levels. The AFoil emulsion can be used to reduce the detrimental impacts of hyperglycemia and oxidative stress.

Discovery and investigation of natural Diels-Alderases.

It has been proposed that biosyntheses of many natural products involve pericyclic reactions, including Diels-Alder (DA) reaction. However, only a small set of enzymes have been proposed to catalyze pericyclic reactions. Most surprisingly, there has been no formal identification of natural enzymes that can be defined to catalyze DA reactions (DAases), despite the wide application of the reaction in chemical syntheses of complex organic compounds. However, recent studies began to accumulate a growing body of evidence that supports the notion that enzymes that formally catalyze DA reactions, in fact exist. In this review, I will begin by describing a short history behind the discovery and characterization of macrophomate synthase, one of the earliest enzymes that was proposed to catalyze an intermolecular DA reaction during the biosynthesis of a substituted benzoic acid in a phytopathogenic fungus Macrophoma commelinae. Then, I will discuss representative enzymes that have been chemically authenticated to catalyze DA reactions, with emphasis on more recent discoveries of DAases involved mainly in fungal secondary metabolite biosynthesis except for one example from a marine streptomycete. The current success in identification of a series of DAases and enzymes that catalyze other pericyclic reactions owes to the combined efforts from both the experimental and theoretical approaches in discovering natural products. Such efforts typically involve identifying the chemical features derived from cycloaddition reactions, isolating the biosynthetic genes that encode enzymes that generate such chemical features and deciphering the reaction mechanisms for the enzyme-catalyzed pericyclic reactions.

Structural insights into Escherichia coli phosphopantothenoylcysteine synthetase by native ion mobility-mass spectrometry.

CoaBC, part of the vital coenzyme A biosynthetic pathway in bacteria, has recently been validated as a promising antimicrobial target. In this work, we employed native ion mobility-mass spectrometry to gain structural insights into the phosphopantothenoylcysteine synthetase domain of E. coli CoaBC. Moreover, native mass spectrometry was validated as a screening tool to identify novel inhibitors of this enzyme, highlighting the utility and versatility of this technique both for structural biology and for drug discovery.

Enzymatic Production and Characterization of Pectic Oligosaccharides Derived from Citrus and Apple Pectins: A GC-MS Study Using Random Forests and Association Rule Learning.

Pectic oligosaccharides (POS) from citrus and apple pectin hydrolysis using ViscozymeL and Glucanex200G have been obtained. According to the results, maximum POS formation was achieved from citrus pectin after 30 min of hydrolysis with ViscozymeL, with a yield of 652 mg g-1 and average molecular mass ( Mw) of 0.8-2.5 kDa, while with Glucanex200G, the yield was 518 mg g-1 and Mw was 0.8-7.1 kDa. Digalacturonic and trigalacturonic acids were identified among other low Mw compounds as di- and tri-POS. In addition, differences in GC-MS spectra of all oligosaccharides found in the hydrolysates were studied by employing random forests and other algorithms to identify structural differences between the obtained POS, and high prediction rates were shown for new samples. Chemical structures were proposed for some influential m/ z ions, and 12 association rules that explain differences according to pectin and enzyme origin were built. This information could be used to establish structure-function relationships of POS.

Characteristic anti-inflammatory and antioxidative effects of enzymatic- and acidic- hydrolysed mycelium polysaccharides by Oudemansiella radicata on LPS-induced lung injury.

The present work investigated the antioxidative, anti-inflammatory and pulmonary protective effects of enzymatic- and acid- hydrolysed mycelia polysaccharides (En-MPS and Ac-MPS) from Oudemansiella radicata on LPS-induced acute lung injury (ALI) mice. The results demonstrated that both En-MPS and Ac-MPS showed potential pulmonary protective effects by decreasing serum levels of hs-CRP and C3, increasing pulmonary enzyme values of SOD, GSH-Px, CAT and the level of T-AOC; reducing the activity of MPO; and down-regulating the contents of MDA and LPO. In addition, the levels of TNF-ɑ, IL-1ß, and IL-6 in BALF of mice treated with En-MPS at a dosage of 400 mg/kg/d were significantly lower than those in the ALI mice. The in vitro antioxidant effects also showed that the En-MPS was more effective than Ac-MPS. Furthermore, the physical properties of polysaccharides were also investigated by GC, HPGPC, FT-IR and NMR. These results indicated that both En-MPS and Ac-MPS possessed potent antioxidant and anti-inflammatory activities, which could be used as an ingestible drug in preventing lung injury.

Steered molecular dynamics simulations reveal the role of Ca2+ in regulating mechanostability of cellulose-binding proteins.

The conversion of cellulosic biomass into biofuels requires degradation of the biomass into fermentable sugars. The most efficient natural cellulase system for carrying out this conversion is an extracellular multi-enzymatic complex named the cellulosome. In addition to temperature and pH stability, mechanical stability is important for functioning of cellulosome domains, and experimental techniques such as Single Molecule Force Spectroscopy (SMFS) have been used to measure the mechanical strength of several cellulosomal proteins. Molecular dynamics computer simulations provide complementary atomic-resolution quantitative maps of domain mechanical stability for identification of experimental leads for protein stabilization. In this study, we used multi-scale steered molecular dynamics computer simulations, benchmarked against new SMFS measurements, to measure the intermolecular contacts that confer high mechanical stability to a family 3 Carbohydrate Binding Module protein (CBM3) derived from the archetypal Clostridium thermocellum cellulosome. Our data predicts that electrostatic interactions in the calcium binding pocket modulate the mechanostability of the cellulose-binding module, which provides an additional design rule for the rational re-engineering of designer cellulosomes for biotechnology. Our data offers new molecular insights into the origins of mechanostability in cellulose binding domains and gives leads for synthesis of more robust cellulose-binding protein modules. On the other hand, simulations predict that insertion of a flexible strand can promote alternative unfolding pathways and dramatically reduce the mechanostability of the carbohydrate binding module, which gives routes to rational design of tailormade fingerprint complexes for force spectroscopy experiments.

Biomedical Applications of Lumazine Synthase.

Lumazine synthase (LS) is a family of enzyme involved in the penultimate step in the biosynthesis of riboflavin. Its enzymatic mechanism has been well defined, and many LS structures have been solved using X-ray crystallography or cryoelectron microscopy. LS is composed of homooligomers, which vary in size and subunit number, including pentamers, decamers, and icosahedral sixty-mers, depending on its species of origin. Research on LS has expanded beyond the initial focus on its enzymatic function to properties related to its oligomeric structure and exceptional conformational stability. These attributes of LS systems have now been repurposed for use in various biomedical fields. This review primarily focuses on the applications of LS as a flexible vaccine presentation system. Presentation of antigens on the surface of LS results in a high local concentration of antigens displayed in an ordered array. Such repetitive structures enable the cross-linking of B-cell receptors and result in strong immune responses through an avidity effect. Potential issues with the use of this system and corresponding solutions are also discussed with the objective of improved utilization of the LS system in vaccine development.

Viscozyme L pretreatment on palm kernels improved the aroma of palm kernel oil after kernel roasting.

With an interest to enhance the aroma of palm kernel oil (PKO), Viscozyme L, an enzyme complex containing a wide range of carbohydrases, was applied to alter the carbohydrates in palm kernels (PK) to modulate the formation of volatiles upon kernel roasting. After Viscozyme treatment, the content of simple sugars and free amino acids in PK increased by 4.4-fold and 4.5-fold, respectively. After kernel roasting and oil extraction, significantly more 2,5-dimethylfuran, 2-[(methylthio)methyl]-furan, 1-(2-furanyl)-ethanone, 1-(2-furyl)-2-propanone, 5-methyl-2-furancarboxaldehyde and 2-acetyl-5-methylfuran but less 2-furanmethanol and 2-furanmethanol acetate were found in treated PKO; the correlation between their formation and simple sugar profile was estimated by using partial least square regression (PLS1). Obvious differences in pyrroles and Strecker aldehydes were also found between the control and treated PKOs. Principal component analysis (PCA) clearly discriminated the treated PKOs from that of control PKOs on the basis of all volatile compounds. Such changes in volatiles translated into distinct sensory attributes, whereby treated PKO was more caramelic and burnt after aqueous extraction and more nutty, roasty, caramelic and smoky after solvent extraction.

Structural insights into natural compounds as inhibitors of Fasciola gigantica thioredoxin glutathione reductase.

Fascioliasis is caused by the helminth parasites of genus Fasciola. Thioredoxin glutathione reductase (TGR) is an important enzyme in parasitic helminths and plays an indispensable role in its redox biology. In the present study, we conducted a structure-based virtual screening of natural compounds against the Fasciola gigantica TGR (FgTGR). The compounds were docked against FgTGR in four sequential docking modes. The screened ligands were further assessed for Lipinski and ADMET prediction so as to evaluate drug proficiency and likeness property. After refinement, three potential inhibitors were identified that were subjected to 50 ns molecular dynamics simulation and free energy binding analyses to evaluate the dynamics of protein-ligand interaction and the stability of the complexes. Key residues involved in the interaction of the selected ligands were also determined. The results suggested that three top hits had a negative binding energy greater than GSSG (-91.479 KJ · mol-1 ), having -152.657, -141.219, and -92.931 kJ · mol-1 for compounds with IDs ZINC85878789, ZINC85879991, and ZINC36369921, respectively. Further analysis showed that the compound ZINC85878789 and ZINC85879991 displayed substantial pharmacological and structural properties to be a drug candidate. Thus, the present study might prove useful for the future design of new derivatives with higher potency and specificity.

Effects of Snailase Treatment on Wettability of Raw Cotton Yarns in Pre-wetting Process of Foam Sizing.

The main objective of the present study was to meticulously investigate an inclusive set of physicochemical properties of pre-wetted cotton yarns and optimize the snailase treatment for raw cotton yarns. Based on single factor design, effects of snailase treatment on the removal percentage of pectins and cotton waxes, wettability, and weight loss were studied in different snailase concentrations and the optimal concentration range was obtained. The Box-Behnken design was employed to determine the optimal condition of snailase treatment for achieving the maximum wettability of cotton yarns. The maximum wettability was obtained when treated at 16.90 g/L of snailase concentration at 56.85 °C for 27.77 s. The theoretical and observed values were in reasonably good agreement, and the deviation was less than 1%. Results revealed that snailase had a positive effect on pectin and wax removal, had an obvious impact on weight loss, and significantly enhanced the wettability of raw cotton yarns.

Functional characterization of the cutI gene for the transcription of carbon monoxide dehydrogenase genes in Mycobacterium sp. strain JC1 DSM 3803.

Carbon monoxide dehydrogenase (CO-DH) in Mycobacterium sp. strain JC1 is a key enzyme for the carboxydotrophic growth, when carbon monoxide (CO) is supplied as a sole source of carbon and energy. This enzyme is also known to act as nitric oxide dehydrogenase (NO-DH) for the detoxification of NO. Several accessory genes such as cutD, cutE, cutF, cutG, cutH, and cutI, are clustered together with two copies of the CO-DH structural genes (cutB1C1A1 and cutB2C2A2) in Mycobacterium sp. strain JC1 and are well conserved in carboxydotrophic mycobacteria. Transcription of the CO-DH structural and accessory genes was demonstrated to be increased significantly by acidified sodium nitrate as a source of NO. A cutI deletion (ΔcutI) mutant of Mycobacterium sp. strain JC1 was generated to identity the function of CutI. Lithoautotrophic growth of the ΔcutI mutant was severely affected in mineral medium supplemented with CO, while the mutant grew normally with glucose. Western blotting, CO-DH activity staining, and CO-DH-specific enzyme assay revealed a significant decrease in the cellular level of CO-DH in the ΔcutI mutant. Northern blot analysis and promoter assay showed that expression of the cutB1 and cutB2 genes was significantly reduced at the transcriptional level in the ΔcutI mutant, compared to that of the wildtype strain. The ΔcutI mutant was much more susceptible to NO than was the wild type.

Biodegradation of pyrene by pseudomonas sp. JPN2 and its initial degrading mechanism study by combining the catabolic nahAc gene and structure-based analyses.

In this study, a pyrene-degrading bacterial strain Pseudomonas sp. JPN2 was isolated from crude oil in Dagang Oilfield, China. The degrading percent of the strain JPN2 to pyrene was increased with the extension of culture time and achieved a maximum of 82.88% after 25 d culture. Meanwhile, four metabolites 4,5-dihydroxy-4,5-dihydropyrene, 4-phenanthrol, 1-hydroxy-2-naphthoic acid and phthalate were detected in the culture solution by GC-MS analysis. In addition, DNA fragments of nahAc gene, encoding α subunit of naphthalene dioxygenase, were amplified by PCR program and sequenced. As a result, it was presumed that the initial cleavage of the aromatic rings on pyrene was occurred at C4 and C5 positions and formed the intermediate 4,5-dihydroxy-4,5-dihydropyrene. This issue had been verified by the interaction analysis between pyrene and the active site of naphthalene dioxygenase in the strain JPN2 by molecular docking. Meanwhile, the differences of the amino acid residues in the active sites of template and target enzymes may be a factor leading to the different biological activity between the strain JPN2 and the other bacteria from the genus Pseudomonas. Additionally, the microcalorimetry analysis displayed that the strain JPN2 had high tolerance for pyrene, and the effect could be negligible under the experimental concentration (100 mg L-1). Consequently, the strain JPN2 was considered as an excellent candidate for the further bioremediation study of pyrene and the other aromatic contaminants.

QSAR-Driven Discovery of Novel Chemical Scaffolds Active against Schistosoma mansoni.

Schistosomiasis is a neglected tropical disease that affects millions of people worldwide. Thioredoxin glutathione reductase of Schistosoma mansoni (SmTGR) is a validated drug target that plays a crucial role in the redox homeostasis of the parasite. We report the discovery of new chemical scaffolds against S. mansoni using a combi-QSAR approach followed by virtual screening of a commercial database and confirmation of top ranking compounds by in vitro experimental evaluation with automated imaging of schistosomula and adult worms. We constructed 2D and 3D quantitative structure-activity relationship (QSAR) models using a series of oxadiazoles-2-oxides reported in the literature as SmTGR inhibitors and combined the best models in a consensus QSAR model. This model was used for a virtual screening of Hit2Lead set of ChemBridge database and allowed the identification of ten new potential SmTGR inhibitors. Further experimental testing on both shistosomula and adult worms showed that 4-nitro-3,5-bis(1-nitro-1H-pyrazol-4-yl)-1H-pyrazole (LabMol-17) and 3-nitro-4-{[(4-nitro-1,2,5-oxadiazol-3-yl)oxy]methyl}-1,2,5-oxadiazole (LabMol-19), two compounds representing new chemical scaffolds, have high activity in both systems. These compounds will be the subjects for additional testing and, if necessary, modification to serve as new schistosomicidal agents.

Quartz crystal microbalance with dissipation and microscale thermophoresis as tools for investigation of protein complex formation between thymidylate synthesis cycle enzymes.

Thymidylate synthase (TS) and dihydrofolate reductase (DHFR) play essential role in DNA synthesis, repair and cell division by catalyzing two subsequent reactions in thymidylate biosynthesis cycle. The lack of either enzyme leads to thymineless death of the cell, therefore inhibition of the enzyme activity is a common and successful tool in cancer chemotherapy and treatment of other diseases. However, the detailed mechanism of thymidylate synthesis cycle, especially the interactions between cycle enzymes and its role remain unknown. In this paper we are the first to show that human TS and DHFR enzymes form a strong complex which might be essential for DNA synthesis. Using two unique biosensor techniques, both highly sensitive to biomolecular interactions, namely quartz crystal microbalance with dissipation monitoring (QCM-D) and microscale thermophoresis (MST) we have been able to determine DHFR-TS binding kinetic parameters such as the Kd value being below 10 µM (both methods), k(on) = 0.46 × 10(4) M(-1) s(-1) and k(off) = 0.024 s(-1) (QCM-D). We also calculated Gibbs free energy as in the order of -30 kJ/mol and DHFR/TS molar ratio pointing to binding of 6 DHFR monomers per 1 TS dimer (both methods). Moreover, our data from MST analysis have pointed to positive binding cooperativity in TS-DHFR complex formation. The results obtained with both methods are comparable and complementary.

Role of protons in the pump cycle of KdpFABC investigated by time-resolved kinetic experiments.

The time-resolved kinetics of the KdpFABC complex solubilized in Aminoxide WS-35 was investigated by ATP concentration jump experiments. ATP was photoreleased from its inactive precursor, caged ATP, and charge movements in the membrane domain of the KdpFABC were detected by the electrochromic dye RH421. At low ATP concentrations, the ATP binding step became rate-limiting with an apparent, pH-independent ATP binding affinity of ~70 µM. At saturating ATP concentrations, the rate-limiting step is the conformational transition (E1-P → P-E2) with a rate constant of ~1.7 s(-1) at 20 °C that was independent of K(+) concentration. This observation together with the detected fluorescence decrease indicates that K(+) (or another positive ion) is bound in the membrane domain after enzyme phosphorylation and the conformational transition to the P-E2 state. pH dependence experiments revealed different roles of H(+) in the transport mechanism. Two different functions of protons for the ion pump must be distinguished. On one hand, there are electrogenically bound "functional" protons, which are not transported but prerequisite for the performance of the ATP-driven half-cycle. On the other hand, protons bind to the transport sites, acting as weak congeners of K(+). There possibly are noncompetitively bound protons, affecting the enzyme activity and/or coupling between KdpA and KdpB subunits. Finally, the recently proposed Post-Albers model for the KdpFABC complex was supplemented with stoichiometry factors of 2 for K(+) and 3 for H(+), and additional inhibitory side reactions controlled by H(+) were introduced, which are relevant at pH <6.5 and/or in the absence of K(+).

In vitro and in vivo antioxidant activities of polysaccharide purified from aloe vera (Aloe barbadensis) gel.

The in vitro and in vivo antioxidant potentials of a polysaccharide isolated from aloe vera gel were investigated. Enzymatic extracts were prepared from aloe vera gel by using ten digestive enzymes including five carbohydrases and five proteases. Among them, the highest yield was obtained with the Viscozyme extract and the same extract showed the best radical scavenging activity. An active polysaccharide was purified from the Viscozyme extract using ethanol-added separation and anion exchange chromatography. Purified aloe vera polysaccharide (APS) strongly scavenged radicals including DPPH, hydroxyl and alkyl radicals. In addition, APS showed a protective effect against AAPH-induced oxidative stress and cell death in Vero cells as well as in the in vivo zebrafish model. In this study, it is proved that both the in vitro and in vivo antioxidant potentials of APS could be further utilized in relevant industrial applications.

(13)C and (63,65)Cu ENDOR studies of CO dehydrogenase from Oligotropha carboxidovorans. Experimental evidence in support of a copper-carbonyl intermediate.

We report here an ENDOR study of an S = 1/2 intermediate state trapped during reduction of the binuclear Mo/Cu enzyme CO dehydrogenase by CO. ENDOR spectra of this state confirm that the (63,65)Cu nuclei exhibits strong and almost entirely isotropic coupling to the unpaired electron, show that this coupling atypically has a positive sign, aiso = +148 MHz, and indicate an apparently undetectably small quadrupolar coupling. When the intermediate is generated using (13)CO, coupling to the (13)C is observed, with aiso = +17.3 MHz. A comparison with the couplings seen in related, structurally assigned Mo(V) species from xanthine oxidase, in conjunction with complementary computational studies, leads us to conclude that the intermediate contains a partially reduced Mo(V)/Cu(I) center with CO bound at the copper. Our results provide strong experimental support for a reaction mechanism that proceeds from a comparable complex of CO with fully oxidized Mo(VI)/Cu(I) enzyme.

Employing bifunctional enzymes for enhanced extraction of bioactives from plants: flavonoids as an example.

A cost-effective and environmentally friendly approach was developed to improve the extraction of active ingredients from plants, in which a bifunctional enzyme was employed for not only facilitating cell wall degradation but also increasing the bioactivity of target compounds in the extract. In the aqueous extraction of flavonoids from Glycyrrhizae radix, Trichoderma viride cellulase, a commercial cell-wall-degrading enzyme, was found to efficiently deglycosylate liquiritin and isoliquiritin, which are of high content but low bioactivity, into their aglycones that have much higher physiological activities for dietary and medicinal uses. Under optimized conditions, the extraction yield of liquiritigenin and isoliquiritigenin aglycones reached 4.23 and 0.39 mg/g of dry weight (dw) with 6.51- and 3.55-fold increases, respectively. The same approach was expanded to the extraction of flavonoids from Scutellariae radix using Penicillium decumbens naringinase, where enhanced production of more bioactive bacalein and wogonin was achieved via enzymatic deglycosylation of bacalin and wogonoside.

Enzyme-assisted extraction of bioactive compounds from ginger (Zingiber officinale Roscoe).

Ginger (Zingiber officinale R.) is a popular spice used in various foods and beverages. 6-Gingerol is the major bioactive constituent responsible for the antiinflammatory, antitumour and antioxidant activities of ginger. The effect of application of α-amylase, viscozyme, cellulase, protease and pectinase enzymes to ginger on the oleoresin yield and 6-gingerol content has been investigated. Pre-treatment of ginger with α-amylase or viscozyme followed by extraction with acetone afforded higher yield of oleoresin (20%±0.5) and gingerol (12.2%±0.4) compared to control (15%±0.6 oleoresin, 6.4%±0.4 gingerol). Extraction of ginger pre-treated with enzymes followed by extraction with ethanol provided higher yield of gingerol (6.2-6.3%) than the control (5.5%) with comparable yields of the oleoresin (31-32%). Also, ethanol extract of cellulase pre-treated ginger had the maximum polyphenol content (37.5 mg/g). Apart from 6-gingerol, 6-paradol along with 6- and 8-methyl shogaols were the other important bio-active constituents in the oleoresin from cellulase-treated ginger.