Herba Epimedii′s biotransformation in vitro simulated gastrointestinal digestion and faecal fermentation systems

OBJECTIVE Epimedium is rich in a variety of beneficial active ingredients,and has been widely used in the ethnopharmacological practices,however,its biotransformation in gastrointestinal digestions remain unclear.This study aimed to investigate the dynamic changes of components and biological activity of Epimedium in the in vitro simulated digestion and subsequent human faecal fermentation.METHODS The models of in vitro simulated saliva,gastric and intestinal digestion,as well as colonic fermentation were constructed to simulate the digestion process of Epimedium.The dynamic changes of components of Epimedium during the simulated digestions in vitro and subsequent human faecal fermentation were investigated by UPLC-MS,HPLC-DAD combined with principal component analysis(PCA)and multi-ingredient quantitative analysis.RESULTS A variety of metabolites with high contents were produced after 0.5 h of intestinal digestion and colonic fermentation 0.5 h.Application of PCA to HPLC data showed the obvious separation of colonic fermentation 0.5 h stage samples from other colonic fermentation stages samples(24,48 and 72 h).Additionally,non-digestion and saliva digestion stage samples clustered together,and there was obvious separation between intestinal digestion samples and gastric digestion samples.The contents of epimedium C,icariin and baohuside I all increased significantly after intestinal digestion[58.70±7.08,47.15±5.68 and(12.78±0.55)mg·g^(-1)]compared with gastric digestion[29.00±5.65,17.40±4.55 and(2.77±0.19)mg·g^(-1)].There were significant differences between sample after 0.5 h of colonic fermentation[64.22±9.32,51.26±6.33 and(16.68±3.19)mg·g^(-1)]and other time points(24,48 and 72 h)in components and the contents of active ingredient,and the content of these components all decreased with the fermentation time.The ability of scavenging ABTS free radicals[IC50=(0.29±0.02)g·L^(-1)]increased significantly compared with gastric digestion[(1.57±0.02)g·L^(-1)],and after 0.5 h of colonic fermentation,the ability also increased significantly.CONCLUSION Gastrointestinal digestion had a significant impact on the contents of active components in Epimedium,and the metabolism of these components mainly occurred in the colon.The intestinal digestion and colonic fermentation significantly improved the anti-ABTS activity of epimedium.

Biotransformation of Gastrodin by Cell Suspension Cultures of Catharanthus roseus

应用长春花 (Catharanthusroseus (L .)G .Don)悬浮细胞培养体系对天麻素进行了生物转化反应研究。经过8d培养形成一个转化产物 ,应用光谱方法鉴定转化产物的结构为对羟基苯甲醇 ,为天麻素水解后形成的甙元。

Biotransformation of Gastrodin by Mucor spinosus

Microbial transformation of gastrodin by Mucor spinosus strain 3.3450, resulted in a product with a transformation rate close to 100 per cent. This product was identified as p-hydroxy benzyl alcohol on the basis of its 1H, 13C NMR and EI-MS spectral data. It could be inferred that the enzyme responsible for the biotransforma-tion reaction was a kind of extracellular and constitutive enzyme since the transformation reaction of the substrate could be carried out in cell free extracts of the fermentation broth of the Mucor spinosus.

RELATIONSHIP BETWEEN GROWTH AND MORPHOLOGY OF METARRHIZIUM ANISOPLIAE IN BIOTRANSFORMATION OF STEROID

The fermentation with mold metarrhizium anisopliae has been carried out to determine the relationship between mycelial morphology and growth.Like the biomass concentration,the mycelial morphology seems to be coupled to growth phases:the fractal dimension,a feature calculated from the morphological images,increases during the late lag phase and the early exponential phase,and culminates at a value near 2.4 at half of the exponential phase.This can be attributed to the marked change of rough and smooth pellet proportions during growth and the change can be well expressed by the fractal analysis of mycelial morphology.The mycelial morphology is also strongly related to the biotransformation activity:a mycelial sample was withdrawn from the culture to use for fractal analysis before the steroid substrate (16α,17α epoxy 4 pregnene 3,20 dione) was added;the higher fractal dimension corresponds to the higher conversion rate of steroid biotransformation.And the maximum of fractal dimension appeared prior to the maximum of biomass concentration.It is therefore possible to utilize the fractal dimension curve to supervise the fermentation process more timely and availably than to use the conventional biomass curve to do.

Study on Anti-oxidation and Inhibitory Effect on Nonenzymatic Glycation Reaction of Fermentation Extract from Biotransformation of Ginkgo biloba L. (EGB) by Hericium erinaceus

[ Objective] In order to study the anti-oxidation and inhibitory effect on nonenzymatic glycation reaction of EGB fermentation extraction biotransformed by Hericium erinaceus. [ Method ] The free radical scavenging ability and reducing capacity of DPPH as well as inhibitory rate of nonenzymatic glycation reaction were measured targets for comparing changes of anti-oxidation and inhibitory effect on nonenzymatic glycation reaction of fermentation lyophilizer and fermentation extraction before and after EGB fermention adsorbed by AB-8 macroporous resin. The EGB fermention was biotransformed by Hericium erinaceus. [ Result ] After adsorbed by AB-8 macroporous resin, the bioactive matters were concentrated and separated. The free radical scavenging rate, reducing capacity and inhibitory rate of nonenzymatic glycation reaction were increased significantly after adsorbed by AB-8 macroporous resin. [ Conclusion] AB-8 macroporous resin could be used for preliminary concentration of EGB fermentation which was biotransformed by Hericium erinaceus.

Biotransformation of Artemisinin by Hairy Root Cultures of Rheum palmatum L.

The biotransformation of artemisinin by hairy root cultures ofRheum palmatum L. was investigated for the first time. The main product, deoxyartemisinin, was isolated and characterized on the basis of its spectral data.

Technologies of Seperating and Exactly 11-α-Hydroxy-Canerenone from Biotransformation Broth

[Objective] The aim was to investigate the effect of four extraction meth- ods on extraction ratio of 11-α-Hydroxy-Canerenone from biotransformation broth. [Method] Previous reported method, soaking, elution method and homogenate meth- ods were used. [Result] Results showed that more than 90% of the biotransformed 11-α-Hydroxy-Canerenone existed in the broth supernatant and mycelia surface. Elu- tion and soaking methods were effective for 11-α-Hydroxy-Canerenone extraction. Extraction yields of 96.0% and 98.8% were obtained by eluting with 400 ml of ethy- lacetate and soaking with ethylacetate for 90 min, respectively. [Conelusion] The method is simple and efficient, showing industrial application potential.

Biotransformation of aesculin by human gut bacteria and identification of its metabolites in rat urine

AIM： To observe the biotransformation process of a Chinese compound, aesculin, by human gut bacteria, and to identify its metabolites in rat urine.METHODS： Representative human gut bacteria were collected from 20 healthy volunteers, and then utilized in vitro to biotransform aesculin under anaerobic conditions. At 0, 2, 4, 8, 12, 16, 24, 48 and 72 h postincubation, 10 mL of culture medium was collected. Metabolites of aesculin were extracted 3 × from rat urine with methanol and analyzed by HPLC. For in vivo metabolite analysis, aesculetin （100 mg/kg） was administered to rats via stomach gavage, rat urine was collected from 6 to 48 h post-administration, and metabolite analysis was performed by LC/ESI-MS and MS/MS in the positive and negative modes.RESULTS： Human gut bacteria could completely convert aesculin into aesculetin in vitro. The biotransformation process occurred from 8 to 24 h post-incubation, with its highest activity was seen from 8 to 12 h. The in vitro process was much slower than the in vivo process. In contrast to the in vitro model, six aesculetin metabolites were identified in rat urine, including 6-hydroxy-7-glucocoumarin（M1）, 6-hydroxy-7-sulf-coumarin （M2）, 6, 7-digluco-coumarin （M3）, 6-glc-7-gluco-coumarin （M4）, 6-O-methyl-7-gluco-coumarin （MS） and 6-O-methyl-7- sulf-coumarin （M6）. Of which, M2 and M6 were novel metabolites.CONCLUSION： Aesculin can be transferred into aesculetin by human gut bacteria and is further modified by the host in vivo. The diverse metabolites of aesculin may explain its pleiotropic pharmaceutical effects.

BIOTRANSFORMATION OF 2α，5α，10β，14β－TETRA－ACETOXY－4（20），11－TAXADIENE BY THE FUNGIJS CUNNINGHAMELLA ECHINULATA

?5α,10β,14β-Tetra-acctoxy-4(20).11-taxadicne (1) could be regioselectively transfonnedinto 6α 10β-dihydroxy-2α,5α,14β-triacetoxy-4(20). 11- taxadiene (2) by Cunninghamella echinnlata infair yield. 6βM.10β-Dihydroxy-2α,5α,14β-triacet 11-taxadienc (3). 10β-hydroxy-2α,5α,14β-triacetoxy-4(20).11-taxadiene (4), 10 β-hyboxy-4β,20-epoxy -2α,5α,14β-triacetoxy- tax - 11- ene (5)asminor prpducts were also isolated

Recent Progress on Biocatalysis and Biotransformations in Ionic Liquids

Ionic liquids have negligibly low vapor pressure, high stability and polarity. They are regarded as green solvents. Enzymes, especially lipases, as well as whole-cell of microbe, are catalytically active in ionic liquids or aqueous-ionic liquid biphasic systems. Up to date, there have been many reports on enzyme-exhibited features and enzyme-mediated reactions in ionic liquids. In many cases, remarkable results with respect to yield, catalytic activity, stability and (enantio-, regio-) selectivity were obtained in ionic liquids in comparison with those observed in conventional media. Accordingly, ionic liquids provide new possibilities for the application of new type of solvent in biocatalytic reactions.

Biotransformation of 14-Deacetoxy-13-oxo sinenxan A by Ginkgo Cell Cultures

Deacetoxy-13-oxo sinenxan A (1) was converted to 9a-hydroxy-13-oxo-2a, 5a, 10b-triacetoxy-4(20),11-taxadiene (2) and 10b-hydroxy-13-oxo-2a,5a,9a-triacetoxy- 4(20), 11- taxadiene (3) by Ginkgo cell suspension cultures in 45% and 15% yields, respectively.

New polyhydroxylated metabolite derived from biotransformation of diosgenin by the white-rot fungus Coriolus versicolor

Microbial transformation of diosgenin(1) was carried out with the white-rot fungus,Coriolus versicolor.A new polyhydroxyl metabolite,(25R)-spirost-5-ene-3β,7β,21-triol(2),was obtained as a result of hydroxylation.Its structure was elucidated on the basis of 1D and 2D NMR as well as HR-ESI-MS spectroscopic analysis.

Time-course monitoring of in vitro biotransformation reaction via solid-phase microextraction-ambient mass spectrometry approaches

The solid-phase microextraction technique quantifies analytes without considerably affecting the sample composition.Herein,a proof-of-concept study was conducted to demonstrate the use of coated probe electrospray ionization(coated-PESI)and coated blade spray(CBS)as ambient mass spectrometry approaches for monitoring drug biotransformation.The ability of these methods was investigated for monitoring the dephosphorylation of a prodrug,combretastatin A4 phosphate(CA4P),into its active form,combretastatin A4(CA4),in a cell culture medium supplemented with fetal bovine serum.The CBS spot analysis was modified to achieve the same extraction efficiency as protein precipitation and obtained results in 7 min.Because coated-PESI performs extraction without consuming any samples,it is the preferred technique in the case of a limited sample volume.Although coated-PESI only extracts small quantities of analytes,it uses the desorption solvent volume of 5-10 pL,resulting in high sensitivity,thus allowing the detection of compounds after only 1 min of extraction.The biotransformation of CA4P into CA4 via phosphatases occurs within the simple matrix,and the proposed sample preparation techniques are suitable for monitoring the biotransformation.

Fatty acid hydratase for value-added biotransformation:A review

The synthesis of hydroxy fatty acids(HFAs) from renewable oil feedstock by addition of water onto C_C bonds has attracted great attention in recent years. Given that selective asymmetric hydration of non-activated C_C bonds has been proven difficult to achieve with chemical catalysts, enzymatic catalysis by fatty acid hydratases(FAHs) presents an attractive alternative approach to produce value-added HFAs with high regio-, enantioand stereospecificity, as well as excellent atom economy. Even though FAHs have just been investigated as a potential biocatalyst for a decade, remarkable information about FAHs in different aspects is available;however, a comprehensive review has not been archived. Herein, we summarize the research progresses on biochemical characterization, structural and mechanistic determination, enzyme engineering, as well as biotechnological application of FAHs. The current challenges and opportunities for an efficient utilization of FAHs in organic synthesis and industrial applications are critically discussed.

THE OVERLOOKED ECOSYSTEM DRIVING FORCE IN NON-EUTROPHICATED FRESHWATER SYSTEMS:DISSOLVED HUMIC SUBSTANCES-A SHORT REVIEW AND OUTLOOK

This review starts with the description of the quantitative significance of dissolved organic material in general and dissolved humic substances (HS) in particular in various ecosystems. Despite their high quantities, the knowledge about the role of HS is still very low and full of old, but still recycled paradigms. HS are thought to be inert or at least refractory and too large to be taken up by aquatic organisms. Instead, I present evidence that dissolved HS that mainly derives from the terrestrial environment, are taken up and directly and/or indirectly interfere with freshwater organisms and, thus, structure biocenoses. Relatively well known is in the meantime the fuelling function of allochthonous HS, which, upon irradiation, release fatty acids, which serve as substrates for microbial growth. This is an indirect effect of HS. Microbes, in turn, are food for mixotrophic algae and (heterotrophic) zooplankton. Thus, non-eutrophicated freshwaters are net-heterotrophic, meaning that respiration exceeds primary production. Furthermore, model calculations exemplify that only a very small portion of the terrestrial production is sufficient to cause net-heterotrophy in these freshwater bodies. But, recent papers show also that due to different stoichiometries the maximal plankton biomass production with algae or mixotrophs is higher than with bacteria. Very recently, several direct effects of HS have been elucidated. Among them are:induction of chaperons (stress shock proteins), induction and modulation of biotransformation enzymes, modulation (mainly inhibition) of photosynthetic oxygen release of aquatic plants, production of an internal oxidative stress, modulation of the offspring numbers in the nematode Caenorhabditis elegans[WTBZ], feminization of fish and amphibs, interference within the thyroid system, and action as chemical attractant to C. elegans. We are still in the phase of identifying the various physiological, biochemical, and molecular-biological effects. Hence, the ecological and ecophysiological significance of these HS-mediated effects still remain somewhat obscure. Nevertheless, HS appear generally to have an impact on the individual as well as on the community and even ecosystem level comparable to that of, for instance, nutrients.

Biotransformation of Flavor Compositions During Fermentation of Litchi (Litchi Chinensis Sonn.) Fruits into Wine

The aim was to examine the biotransformation of chemical compounds during the fermentation of litchi wine.S.bccyanus BV818 was inoculated to litchi juice(Heiye)to initiate the fermentation.Acetic acid decreased dramatically,succinic acid and DL-malic acid increased sharply.Saturated free fatty acid increased,especially the concentration of the free fatty acid with long carbon chain(more than 10 carbons)increased significantly.The unique flavor compounds of fresh litchi including linalool,α-terpineol,β-citronellol and other terpenoids remained in the litchi wine were transformed to other aroma constituents,by which the primary litchi flavor was retained.The wine had a fruity flavor and delicate bouquet and had harmonious sourness and sweetness.The litchi'Heiye'was suitable for being fermented into litchi wine.

Biotransformation of Carmoisine and Reactive Black 5 Dyes Using <i>Saccharomyces cerevisiae</i>

Saccharomyces cerevisiae (baker’s yeast) is the most important industrial microorganisms. This yeast is commonly used as a leavening agent in baking bread and bakery products, where it produces carbon dioxide from converting of the fermentable sugars present in the dough. Nowadays, industrial and chemical activities led to produce new compounds with new kinds of contamination in the environment. Discharge of untreated or partially treated industrial sewage has created the contamination problems of rivers and lakes such as drugs, oil, heavy metals, paints, pesticides and various chemical compounds in them. Hence, it is necessary to control and reduce the levels of these compounds in wastewater and bring them to permissible values. This study aims to study the bioconversion potential of commonly available Saccharomyces cerevisiae for the two textile dyes of Carmoisine and Reactive Black 5. Reaction mixtures for biotransformation of dyes included 50 mg/l Carmoisine or 25 mg/l Reactive Black 5 and 1% dried harvested cells of S. cerevisiae (bread’s yeast) were tested. Harvested dry and wet yeast were studied for this purpose. The results show that harvested cells of Saccharomyces cerevisiae are able to bioconvert Carmoisine and Reactive Black 5. Reactive Black 5, Carmoisine are degraded by biotransformation 85% and 53% within 24 hours in water at the room temperature.

Unbalanced biotransformation metabolism and oxidative stress status: implications for deficient fatty acid oxidation

The concept of accumulating xenobiotics within the human body as a health risk is well known. However, these compounds can also be endo-genous, as in the case of inborn errors of me-tabolism, and lead to some of the same symp-toms as seen in xenobiotic intoxication. Bio-transformation of both exogenous and endo-genous toxic compounds is an important function of the liver, and the critical balance between these systems is of fundamental importance for cellular health. We propose a novel model, to describe the critical balance between Phase I and Phase II biotransformation and how a disturbance in this balance will increase the oxidative stress status, with resulting pathological consequences. We further used deficient fatty acid oxidation to verify the proposed model, as deficient fatty acid oxidation is associated with the accumulation of characteristic metabolites. These accumulating metabolites undergo both Phase I and Phase II biotransformation reactions, with resulting depletion of biotransformation substrates and co-factors. Depletion of these important biomolecules is capable of disturbing the balance between Phase I and Phase II reactions, and disturbance of this balance will increase oxidative stress status. The value of the proposed model is illustrated by its application to a clinical case investigated in our laboratory. In this case the possibility of deficient fatty acid oxidation only became evident once the critical balance between Phase I and Phase II biotransformation was restored with oral replenishment of biotransformation substrates. In addition to bio-chemical improvement, there was also significant clinical improvement. The significance of this model lies within the treatment possibilities, as the assessment of biotransformation metabolism and oxidative stress status can lead to the development of nutritional treatment strategies to correct imbalances. This in turn may reduce the chances of, or delay the onset of certain disease states.

Biotransformation of 6-deoxypseudoanisatin by Synechocystis sp. PCC6803

Objective:To explore the ability of Synechocystis sp.PCC6803 in transforming 6-deoxypseudoanisatin.Methods:The experiment was performed by incubating 6-deoxypseudoanisatin with the freshwater cyanobacterium Synechocystis sp.PCC6803 under continuous white light at 30C for 5 days.The crude converted product was detected using thin-layer chromatography(TLC)and further analyzed using high-performance liquid chromatography(HPLC)as well as HPLC with electron spray ionization mass spectrometry(HPLC-ESI-MS).Results:TLC results showed that 6-deoxypseudoanisatin was converted into a less polar product.HPLC and MS data indicated that the retention time of the converted product increased in comparison with the standard of 6-deoxypseudoanisatin.Conclusion:Thus,the study appears to demonstrate that Synechocystis sp.PCC6803 can transform 6-deoxypseudoanisatin.The polarity of the converted product is less than that of 6-deoxypseudoanisatin.

Study on Biotransformation of Natural Anti-tumor Drug Podophyllotoxin

[Objective] This study aimed to select stains for transforming podophyllotoxin, to isolate the transformation product and to identify the structure. [ Method~ Structure of podophyllotoxin was modified with microbial batch transformation method. Podophyllotoxin and its transformation products were detected by using TLC and HPLC methods to select strains with ability to transform podophyllotoxin. Scale-up fermentation of one podophyllotoxin-transforming strain was carried out and the fermented material was extracted with EtoAc, isolated with conventional silica gel column chromatography, structure of the transformation product was identified by using classic spectroscopy technology. [ Result ] Based on researches of pedophyllotoxin transformation, seven strains with ability to transform po- dophyllotoxin were selected, including BS. Str-1, BS. Str-2, BS. Str-3, BS. Str-7, BS. Str-18, BS. Str-21 and BS. Str-29. By scale up fermentation, the only trans- formation product of BS. Str-21 strain was extracted and isolated, which was identified as picropodophyllotoxin. [ Conclusion] This study demonstrates that podo- phyllotoxin derivatives can be obtained by biotransformation methods, which lays the foundation for acquisition of natural drugs with high anti-tumor activity.