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1.
Ana Figueiras Jorge M. G. Sarraguça Alberto A. C. C. Pais Rui A. Carvalho J. Francisco Veiga 《AAPS PharmSciTech》2010,11(1):233-240
In this study, we investigate how the effect of l-arginine (ARG) and cyclodextrins upon omeprazole (OME) stability and solubility. The effect of the presence of ARG on the
apparent stability constants (K1:1) of the inclusion complexes formed between OME and each cyclodextrin, β-cyclodextrin (βCD), and methyl-β-cyclodextrin (MβCD)
is studied by phase solubility diagrams and nuclear magnetic resonance (NMR) spectroscopy. The interaction of OME with those
cyclodextrins, in the presence of ARG, is characterized using NMR spectroscopy and molecular dynamics simulations. ARG significantly
increases the drug solubility and complex stability, in comparison to inclusion complexes formed in its absence. The effect
is more pronounced for the OME:βCD complex. ARG also contributes to a larger stability of OME when free in aqueous solution.
The combination of ARG with cyclodextrins can represent an important tool to develop stable drug formulations. 相似文献
2.
Francisella tularensis ssp. tularensis is a category A select agent and the causal organism for the zoonotic disease tularemia. The vast majority of F. tularensis isolates are β-lactamase-positive. β-lactamase production is widely believed to be responsible for the inefficacy of β-lactams in the treatment of tularemia. In this study, we report the cloning and characterization of the two chromosomally encoded F. tularensis ssp. holarctica live-vaccine strain (LVS) β-lactamases. The two LVS β-lactamases were homologous to F. tularensis Schu S4 open reading frames FTT0681c and FTT0611c and have been named bla1
LVS
and bla2
LVS
, respectively. Recombinant expression in Escherichia coli suggested that bla1
LVS
did not encode a functional β-lactamase, whereas bla2
LVS
encoded a functional β-lactamase that hydrolyzed penicillins but was inactive against third-generation cephalosporins, including cefprozil. As both LVS and Schu S4 were susceptible to cefprozil, we developed three new shuttle vectors based on selection for the production of the Blashv-2 extended-spectrum β-lactamase with cefprozil. The resulting shuttle vectors were suitable for recombinant gene expression and complementation studies in LVS and Schu S4. 相似文献
3.
Kawai R Igarashi K Yoshida M Kitaoka M Samejima M 《Applied microbiology and biotechnology》2006,71(6):898-906
When Phanerochaete chrysosporium was grown with laminarin (a β-1,3/1,6-glucan) as the sole carbon source, a β-1,3-glucanase with a molecular mass of 36 kDa was produced as a major extracellular protein. The cDNA encoding this enzyme was cloned, and the deduced amino acid sequence revealed that this enzyme belongs to glycoside hydrolase family 16; it was named Lam16A. Recombinant Lam16A, expressed in the methylotrophic yeast Pichia pastoris, randomly hydrolyzes linear β-1,3-glucan, branched β-1,3/1,6-glucan, and β-1,3-1,4-glucan, suggesting that the enzyme is a typical endo-1,3(4)-β-glucanase (EC 3.2.1.6) with broad substrate specificity for β-1,3-glucans. When laminarin and lichenan were used as substrates, Lam16A produced 6-O-glucosyl-laminaritriose (β-d-Glcp-(1–>6)-β-d-Glcp-(1–>3)-β-d-Glcp-(1–>3)-d-Glc) and 4-O-glucosyl-laminaribiose (β-d-Glcp-(1–>4)-β-d-Glcp-(1–>3)-d-Glc), respectively, as one of the major products. These results suggested that the enzyme strictly recognizes β-d-Glcp-(1–>3)-d-Glcp at subsites −2 and −1, whereas it permits 6-O-glucosyl substitution at subsite +1 and a β-1,4-glucosidic linkage at the catalytic site. Consequently, Lam16A generates non-branched oligosaccharide from branched β-1,3/1,6-glucan and, thus, may contribute to the effective degradation of such molecules in combination with other extracellular β-1,3-glucanases. 相似文献
4.
Songmee Bae Jaehoon Lee Eunah Kim Jaehwa Lee Jaeyon Yu Yeonho Kang 《Journal of microbiology (Seoul, Korea)》2010,48(1):84-88
Haemophilus influenzae is a frequent causative bacterial pathogen of respiratory tract infections. Resistance to β-lactam antibiotics has been a
significant clinical problem in treatment for H. influenzae respiratory infections. This study describes the serotype, antibiotic resistance and distribution of TEM-1 or ROB-1 β-lactamase
in H. influenzae isolates from local private hospitals from 2002 to 2004. Among the 100 H. influenzae respiratory isolates, only 7% were identified as serotypes a, b, e, and f, with the remaining 93% being nontypeable. Resistance
to ampicillin, cefaclor, and tetracycline was 57%, 46%, and 16%, respectively. All strains were susceptible to azithromycin
and ciprofloxacin, whereas amoxicillin/clavulanate, cefotaxime, and imipenem exhibited reduced susceptibilities of 99%, 99%,
and 91%, respectively. All 57 ampicillinresistant strains (minimum inhibitory concentration, MIC≥4 μg/ml) were β-lactamase-positive
and possessed the TEM-1 type β-lactamase. One β-lactamase-positive amoxicillin/clavulanate-resistant isolate that was resistant
to ampicillin (MIC>128 μg/ml) had the TEM-1 type β-lactamase and not susceptible to cefaclor and cefotaxime. Analysis of penicillin
binding protein 3 revealed six residues (Asp-350, Met-377, Ala-502, Asn-526, Val-547, and Asn-569) that were substituted by
Asn, Ile, Val, Lys, Ile, and Ser, respectively. 相似文献
5.
Ferenc Ötvös Dmitry S. Gembitsky Richard F. Murphy Sándor Lovas 《International journal of peptide research and therapeutics》2007,13(1-2):329-336
Steric requirements of binding [Nle10]NKA(4–10) to NK-2 receptor were studied by introducing conformationally constrained amino acid analogs into its sequence.
Two series of [Nle10]NKA(4–10) analogs were synthesized to investigate (i) the significance of a putative β-turn in the receptor-ligand interaction by insertion of either (S)- or (R)-Gly8{ANC-2}Leu9 γ-lactams to mimic a β-turn constraint, and (ii) the effect of hindered rotation in the Φ, χ1 and χ2 dihedral angle space of the crucially important Phe6 which was replaced systematically with d-Phe, d- and l-Tyr, as well as with their conformationally constrained analogs, Tic, HOTic and β-MePhe. Competition binding experiments
with [3H]NKA were performed using cloned human NK-2 receptors expressed in CHO cells. The analog possessing only an (R)-Gly8{ANC-2}Leu9 constraint, had the same binding affinity as that of the parent peptide. The rank order of potency of the other analogs showed
a cumulative effect of different structural modifications in decreasing the binding affinity, i.e., when changing the configuration of the lactam ring to S, replacing Phe6 with constrained analogs, Tic or β-MePhe, changing the configuration of the amino acid at position six to d, and introducing a hydroxyl group on the aromatic ring.
Ferenc ?tv?s and Dmitry S. Gembitsky - Made an equal contribution.
Abbreviations used for amino acids and peptides follow the recommendations of the IUPAC-IUB Commission of Biochemical Nomenclature,
Eur. J. Biochem. (1984) 138, 9–37 相似文献
6.
Amyloid β peptides appear to play a role in physiological processes; however, they are also involved in the pathogenesis of
Alzheimer disease. Their actions under normal conditions are probably mediated by soluble monomeric l-isoforms at low concentrations, perhaps via highly specific interactions. On the contrary, toxic effects of aggregated natural
l-isoforms/synthetic d-isoforms on membranes are very similar, but synthetic reverse/random l-isoforms without pronounced aggregation properties are not toxic. Our previous work reported interactions of non-aggregated/aggregated
l-isoforms of amyloid β peptides 1–40/1–42 with racemic 24-hydroxycholesterol. In this study, stereospecificity in the interactions
of natural 24(S)hydroxycholesterol (cerebrosterol) or synthetic 24(R)hydroxycholesterol with soluble fragment 1–40 was evaluated
by means of an in vitro test based on increased vulnerability of the hemicholinium-3 sensitive high-affinity choline uptake
system in rat hippocampal cholesterol-depleted synaptosomes to the actions of amyloid β; computational simulations were also
performed. Our results suggest that: (1) 24(S)hydroxycholesterol interacts with l-peptide 1–40 but not with the reverse l-peptide 40–1, (2) 24(R)hydroxycholesterol does not interact with l-peptide 1–40 or reverse 40–1, and (3) both enantiomers can probably interact with d-peptide 1–40. Therefore, the binding of 24(S)hydroxycholesterol is not fully stereospecific and the interaction could not
reflect a physiological mechanism. Data from the computational simulation indicate that the hydrophobic core of the amyloid
β molecule interacts with the hydrophobic part of 24(S)hydroxycholesterol, but no hydrogen bonds with high stability were
found. Using this procedure, globular amyloid β could retain 24(S)hydroxycholesterol and thus contribute to its pathological
accumulation in the brains of patients with Alzheimer disease. 相似文献
7.
[(4-methoxy-4(3-β-d-galactose-4-chlorophenyl)]spiro[1,2-dioxetane-3-1,3-tricyclo[7.3.1.02,7]tridec-2,7-ene] (“sβ-Gal 102”) and sodium [4-methoxy-4(3-β-d-glucuronic acid-4-chlorophenyl)]spiro[1,2-dioxetane-3-1,3-tricyclo[7.3.1.02,7]tridec-2,7-ene] (“sβ-Glucor 102”) are carbohydrate-containing 1,2-dioxetane compounds that produce chemiluminescence upon
enzymatic hydrolysis by β-d-galactosidase, and β-d-glucuronidase, respectively. In this study, we have characterized and validated a sensitive detection principle for viable
Escherichia coli based on enzymatic cleavage of sβ-Gal 102 and sβ-Glucor 102 (“ColiLight II”). The proposed chemiluminescent assay was optimized
with respect to analytical requirements including incubation time, temperature, pH, enzyme induction, and cell permeabilization.
The sensitivity and specificity rates of the assay were tested on ten different bacterial genera. The assay was found to be
representative based on low coefficients of variations for both accuracy and precision. The analysis time was less than 1 h
and the analytical detection limit was 102 to 103
E. coli cells. In combination with membrane filtration and a brief resuscitation step of 4 h, the proposed assay was capable of detecting
low concentrations of stressed E. coli in potable water (<30 CFU 100 ml−1). The proposed chemiluminescent enzyme assay may be used for assessing the metabolic activity of E. coli in oligotrophic environments and for early warning detection of low concentrations of E. coli in water for human consumption. 相似文献
8.
Chen GT Yang M Song Y Lu ZQ Zhang JQ Huang HL Wu LJ Guo DA 《Applied microbiology and biotechnology》2008,77(6):1345-1350
Preparative-scale fermentation of ginsenoside Rb1 (1) with Acremonium strictum AS 3.2058 gave three new compounds, 12β-hydroxydammar-3-one-20 (S)-O-β-d-glucopyranoside (7), 12β, 25-dihydroxydammar-(E)-20(22)-ene-3-O-β-d-glucopyranosyl-(1→2)-β-d-glucopyranoside (8), and 12β, 20 (R), 25-trihydroxydammar-3-O-β-d-glucopyranosyl-(1→2)-β-d-glucopyranoside (9), along with five known compounds, ginsenoside Rd (2), gypenoside XVII (3), ginsenoside Rg3 (4), ginsenoside F2 (5), and compound K (6). The structural elucidation of these metabolites was based primarily on one- and two-dimensional nuclear magnetic resonance
and high-resolution electron spray ionization mass spectra analyses. Among these compounds, 2–6 are also the metabolites of ginsenoside Rb1 in mammals. This result demonstrated that microbial culture parallels mammalian metabolism; therefore, A. strictum might be a useful tool for generating mammalian metabolites of related analogs of ginsenosides for complete structural identification
and for further use in pharmaceutical research in this series of compounds. In addition, the biotransformation kinetics was
also investigated. 相似文献
9.
Hemicellulose bioconversion 总被引:24,自引:0,他引:24
Saha BC 《Journal of industrial microbiology & biotechnology》2003,30(5):279-291
Various agricultural residues, such as corn fiber, corn stover, wheat straw, rice straw, and sugarcane bagasse, contain about
20–40% hemicellulose, the second most abundant polysaccharide in nature. The conversion of hemicellulose to fuels and chemicals
is problematic. In this paper, various pretreatment options as well as enzymatic saccharification of lignocellulosic biomass
to fermentable sugars is reviewed. Our research dealing with the pretreatment and enzymatic saccharification of corn fiber
and development of novel and improved enzymes such as endo-xylanase, β-xylosidase, and α-l-arabinofuranosidase for hemicellulose bioconversion is described. The barriers, progress, and prospects of developing an
environmentally benign bioprocess for large-scale conversion of hemicellulose to fuel ethanol, xylitol, 2,3-butanediol, and
other value-added fermentation products are highlighted. 相似文献
10.
Three different types of β-d-galactosidase (EC 3.2.1.23) could be distinguished in rabbit tissues using electrophoretic procedures. (1) Acid β-d-galactosidase with a low mobility and maximal activity atpH 3–5 was found in the particulate fraction of various tissue homogenates. This enzyme hydrolyzed 4-methylumbelliferyl-d-galactoside, but no activity against other glycoside substrates could be demonstrated. The enzyme was inhibited by galactono-(1
→ 4)-lactone. (2) Lactose-hydrolyzing β-d-galactosidase with an intermediate mobility was found only in juvenile small intestine. Most of the activity was found in
the particulate fraction of the cell. The enzyme hydrolyzed several other synthetic glycoside substrates besides lactose.
It was most active atpH 5–6 and strongly inhibited by glucono-(1 → 5)-lactone but not much affected by galactono-(1 → 4)-lactone. (3) Neutral β-d-galactosidase with a fast mobility and maximal activity atpH 6–8 was found in the soluble fraction of homogenates from liver, kidney, and small intestine. This enzyme also showed a
broad substrate specificity; it possessed activity against aryl-β-d-glucoside, -fucoside, and -galactoside substrates but not against lactose. The enzyme was strongly inhibited by glucono-(1
→ 5)-lactone and (less) by galactone-(1 → 4)-lactone. Neutral β-d-galactosidase and neutral β-d-glucosidase (EC 3.2.1.21) are probably identical enzymes in the rabbit. Individual variation, in both electrophoretic mobility
and activity, was found for neutral β-d-galactosidase. Genetic analysis of the electrophoretic variants revealed that two alleles at an autosomal locus are responsible
for this variation.
This investigation was supported in part by Public Health Service Grant RR-00251 from the Division of Research Resources and
by funds of the University of Utrecht. 相似文献
11.
S Chakraborti R K Sani U C Banerjee R C Sobti 《Journal of industrial microbiology & biotechnology》2000,24(1):58-63
An extracellular β-galactosidase which catalyzed the production of galacto-oligosaccharide from lactose was harvested from
the late stationary-phase of Bacillus sp MTCC 3088. The enzyme was purified 36.2-fold by ZnCl2 precipitation, ion exchange, hydrophobic interaction and gel filtration chromatography with an overall recovery of 12.7%.
The molecular mass of the purified enzyme was estimated to be about 484 kDa by gel filtration on a Sephadex G-200 packed column
and the molecular masses of the subunits were estimated to be 115, 86.5, 72.5, 45.7 and 41.2 kDa by sodium dodecyl sulfate-polyacrylamide
gel electrophoresis. The isoelectric point of the native enzyme, determined by polyacrylamide gel electrofocusing, was 6.2.
The optimum pH and temperature were 8 and 60°C, respectively. The Michaelis–Menten constants determined with respect to o-NO2-phenyl-β-D-galactopyranoside and lactose were 6.34 and 6.18 mM, respectively. The enzyme activity was strongly inhibited (68%) by galactose,
the end product of lactose hydrolysis reaction. The β-galactosidase was specific for β-D anomeric linkages. Enzyme activity was significantly inhibited by metal ions (Hg2+, Cu2+ and Ag+) in the 1–2.5 mM range. Mg2+ was a good activator. Catalytic activity was not affected by the chelating agent EDTA. Journal of Industrial Microbiology & Biotechnology (2000) 24, 58–63.
Received 09 February 1999/ Accepted in revised form 24 September 1999 相似文献
12.
Paula Alayón-Luaces Eduardo A. Pagano Luis A. Mroginski Gabriel O. Sozzi 《Plant Cell, Tissue and Organ Culture》2008,95(3):257-263
α-l-Arabinofuranosidase, α- and β-d-xylosidase, and β-d-glucosidase activity was detected in the soluble fraction (S-F) extracted with water and in the NaCl-released fraction (NaCl-F)
extracted with a high-salt concentration buffer from apple callus cultures. The activity was found to be differentially modulated
by the addition of various plant growth regulators (PGRs) to calluses that had lost their requirement for specific PGRs (“habituation”
phenomenon). α-l-Arabinofuranosidase activity was 93%, 130%, 126% and 186% higher in the NaCl-F from IAA-, IBA-, ABA- and GA3-treated callus than in that extracted from untreated callus while S-F α-l-arabinofuranosidase activity was only 71%, 24%, 55% and 66% higher, respectively. α-d-Xylosidase displayed low activity levels in both S-F and NaCl-F but 2iP-treated callus showed higher α-d-xylosidase activity in both fractions than the control. 2,4-D increased α-d-xylosidase activity by 110% in the NaCl-F but decreased it by 40% in the S-F. β-d-Xylosidase activity increased by 99% in S-F from 2iP-treated callus but slightly decreased in the NaCl-F. In GA3-treated callus, NaCl-F β-d-xylosidase activity increased by 188%. S-F and NaCl-F from Picloram-treated callus showed undetectable or only slightly noticeable
α-l-arabinofuranosidase, α-d-xylosidase and β-d-xylosidase activity. Interestingly, β-d-glucosidase activity rose 28-fold in the S-F extracted from Picloram-treated callus. β-d-glucosidase was the only enzyme assayed that greatly increased its NaCl-F activity after 10 subcultures, and the addition
of any PGR to the callus culture –except for Picloram and ABA– decreased its activity, suggesting that this enzyme may be
associated with certain stress conditions, such as PGR starvation or Picloram addition. This is the first report on glycoside
hydrolases from fruit callus as modulated by different PGRs. 相似文献
13.
Sateeshkumar Sathigari Gurkishan Chadha Y-H. Phillip Lee Nydeia Wright Daniel L. Parsons Vijay K. Rangari Oladiran Fasina R. Jayachandra Babu 《AAPS PharmSciTech》2009,10(1):81-87
Efavirenz (EFV) is an oral antihuman immunodeficiency virus type 1 drug with extremely poor aqueous solubility. Thus, its
gastrointestinal absorption is limited by the dissolution rate of the drug. The objective of this study was to characterize
the inclusion complexes of EFV with β-cyclodextrin (β-CD), hydroxypropyl β-CD (HPβCD), and randomly methylated β-CD (RMβCD)
to improve the solubility and dissolution of EFV. The inclusion complexation of EFV with cyclodextrins in the liquid state
was characterized by phase solubility studies. The solid-state characterization of various EFV and CD systems was performed
by X-ray diffraction, differential scanning calorimetry, and scanning electron microscopy analyses. Dissolution studies were
carried out in distilled water using US Pharmacopeia dissolution rate testing equipment. Phase solubility studies provided
an AL-type solubility diagram for β-CD and AP-type solubility diagram for HPβCD and RMβCD. The phase solubility data enabled calculating stability constants (K
s) for EFV-βCD, EFV-HPβCD, and EFV-RMβCD systems which were 288, 469, and 1,073 M−1, respectively. The physical and kneaded mixtures of EFV with CDs generally provided higher dissolution of EFV as expected.
The dissolution of EFV was substantially higher with HPβCD and RMβCD inclusion complexes prepared by the freeze drying method.
Thus, complexation with HPβCD and RMβCD could possibly improve the dissolution rate-limited absorption of EFV. 相似文献
14.
Nielsen KA Hrmova M Nielsen JN Forslund K Ebert S Olsen CE Fincher GB Møller BL 《Planta》2006,223(5):1010-1023
Barley (Hordeum vulgare L.) produces a leucine-derived cyanogenic β-d-glucoside, epiheterodendrin that accumulates specifically in leaf epidermis. Barley leaves are not cyanogenic, i.e. they
do not possess the ability to release hydrogen cyanide, because they lack a cyanide releasing β-d-glucosidase. Cyanogenesis was reconstituted in barley leaf epidermal cells through single cell expression of a cDNA encoding
dhurrinase-2, a cyanogenic β-d-glucosidase from sorghum. This resulted in a 35–60% reduction in colonization rate by an obligate parasite Blumeria graminis f. sp. hordei, the causal agent of barley powdery mildew. A database search for barley homologues of dhurrinase-2 identified
a (1,4)-β-d-glucan exohydrolase isozyme βII that is located in the starchy endosperm of barley grain. The purified barley (1,4)-β-d-glucan exohydrolase isozyme βII was found to hydrolyze the cyanogenic β-d-glucosides, epiheterodendrin and dhurrin. Molecular modelling of its active site based on the crystal structure of linamarase
from white clover, demonstrated that the disposition of the catalytic active amino acid residues was structurally conserved.
Epiheterodendrin stimulated appressoria and appressorial hook formation of B. graminis in vitro, suggesting that loss of cyanogenesis in barley leaves has enabled the fungus to utilize the presence of epiheterodendrin
to facilitate host recognition and to establish infection. 相似文献
15.
Rau U Kuenz A Wray V Nimtz M Wrenger J Cicek H 《Applied microbiology and biotechnology》2009,81(5):827-837
Trametes versicolor ATCC 200801 secretes 4.1 g L−1 of exopolysaccharide (EPS) when synthetic minimal medium and low-shear bioreactor cultivation technique are used. Structural
and compositional analyses by thin layer chromatography, gas chromatography–mass spectrometry, electrospray ionization tandem
mass spectrometry, and nuclear magnetic resonance spectroscopy yielded predominantly glucose and small amounts of galactose,
mannose, arabinose, and xylose. The main EPS is composed of β-1,3/β-1,6-linked d-glucose molecules which is identical with Schizophyllan but does not possess a triple helical arrangement as secondary structure.
Two molar mass fractions were detected by size exclusion chromatography yielding weight-average molecular weights of 4,100
and 2.6 kDa. Protein content varies between 2–3.6% (w/w). The exopolysaccharide is different in the nature of the glycosidic linkage, composition of monosaccharides, protein content,
and weight-average molecular weight compared to the well-known polysaccharopeptide (PSP) and polysaccharopeptide Krestin (PSK). 相似文献
16.
Surface plasmon resonance imaging for real-time, label-free analysis of protein interactions with carbohydrate microarrays 总被引:1,自引:0,他引:1
Karamanska R Clarke J Blixt O Macrae JI Zhang JQ Crocker PR Laurent N Wright A Flitsch SL Russell DA Field RA 《Glycoconjugate journal》2008,25(1):69-74
Plant lectin recognition of glycans was evaluated by SPR imaging using a model array of N-biotinylated aminoethyl glycosides of β-d-glucose (negative control), α-d-mannose (conA-responsive), β-d-galactose (RCA120-responsive) and N-acetyl-β-d-glucosamine (WGA-responsive) printed onto neutravidin-coated gold chips. Selective recognition of the cognate ligand was observed
when RCA120 was passed over the array surface. Limited or no binding was observed for the non-cognate ligands. SPR imaging of an array
of 40 sialylated and unsialylated glycans established the binding preference of hSiglec7 for α2-8-linked disialic acid structures
over α2-6-sialyl-LacNAcs, which in turn were recognized and bound with greater affinity than α2-3-sialyl-LacNAcs. Affinity
binding data could be obtained with as little as 10–20 μg of lectin per experiment. The SPR imaging technique was also able
to establish selective binding to the preferred glycan ligand when analyzing crude culture supernatant containing 10–20 μg
of recombinant hSiglec7-Fc. Our results show that SPR imaging provides results that are in agreement with those obtained from
fluorescence based carbohydrate arrays but with the added advantage of label-free analysis. 相似文献
17.
An DS Cui CH Sung BH Yang HC Kim SC Lee ST Im WT Kim SG 《Applied microbiology and biotechnology》2012,94(3):673-682
The gene encoding an α-l-arabinofuranosidase that could biotransform ginsenoside Rc {3-O-[β-d-glucopyranosyl-(1–2)-β-d-glucopyranosyl]-20-O-[α-l-arabinofuranosyl-(1–6)-β-d-glucopyranosyl]-20(S)-protopanaxadiol} to ginsenoside Rd {3-O-[β-d-glucopyranosyl-(1–2)-β-d-glucopyranosyl]-20-O-β-d-glucopyranosyl-20(S)-protopanaxadiol} was cloned from a soil bacterium, Rhodanobacter ginsenosidimutans strain Gsoil 3054T, and the recombinant enzyme was characterized. The enzyme (AbfA) hydrolyzed the arabinofuranosyl moiety from ginsenoside
Rc and was classified as a family 51 glycoside hydrolase based on amino acid sequence analysis. Recombinant AbfA expressed
in Escherichia coli hydrolyzed non-reducing arabinofuranoside moieties with apparent K
m values of 0.53 ± 0.07 and 0.30 ± 0.07 mM and V
max values of 27.1 ± 1.7 and 49.6 ± 4.1 μmol min−1 mg−1 of protein for p-nitrophenyl-α-l-arabinofuranoside and ginsenoside Rc, respectively. The enzyme exhibited preferential substrate specificity of the exo-type
mode of action towards polyarabinosides or oligoarabinosides. AbfA demonstrated substrate-specific activity for the bioconversion
of ginsenosides, as it hydrolyzed only arabinofuranoside moieties from ginsenoside Rc and its derivatives, and not other sugar
groups. These results are the first report of a glycoside hydrolase family 51 α-l-arabinofuranosidase that can transform ginsenoside Rc to Rd. 相似文献
18.
The study was designed to investigate the effect of cyclodextrins (CDs) on the solubility, dissolution rate, and bioavailability
of cilostazol by forming inclusion complexes. Natural CDs like β-CD, γ-CD, and the hydrophilic β-CD derivatives, DM-β-CD and
HP-β-CD, were used to prepare inclusion complexes with cilostazol. Phase solubility study was carried out and the stability
constants were calculated assuming a 1:1 stoichiometry. Solid cilostazol complexes were prepared by coprecipitation and kneading
methods and compared with physical mixtures of cilostazol and cyclodextrins. Prepared inclusion complexes were characterized
by Fourier transform infrared spectroscopy, differential scanning calorimetry (DSC), and X-ray diffraction (XRD) studies.
In vitro dissolution study was performed using phosphate buffer pH 6.4, distilled water, and HCl buffer pH 1.2 as dissolution medium.
The optimized inclusion complex was studied for its bioavailability in rabbit and the results were compared with those of
pure cilostazol and Pletoz-50. Phase solubility study showed dramatic improvement in the solubility of drug by formation of
complexes, which was further increased by pH adjustment. The dissolution rate of cilostazol was markedly augmented by the
complexation with DM-β-CD. DSC and XRD curves showed sharp endothermic peaks indicating the reduction in the microcrystallinity
of cilostazol. Selected inclusion complex was also stable at ambient temperature up to 6 months. The in vivo study revealed that DM-β-CD increased the bioavailability of cilostazol with low variability in the absorption. Among all
cilostazol–cyclodextrins complexes, cilostazol–DM-β-CD inclusion complex (1:3) prepared by coprecipitation method showed 1.53-fold
and 4.11-fold increase in absorption along with 2.1-fold and 2.97-fold increase in dissolution rate in comparison with Pletoz-50
and pure cilostazol, respectively. 相似文献
19.
Søren Bastholm Lasse Wahlstrøm Louise Appel Bjergbæk Peter Roslev 《World journal of microbiology & biotechnology》2008,24(10):2323-2330
Traditional cultivation-dependent tests for coliform bacteria in food and drinking water take 18–24 h to complete. Bioluminescence-based
enzyme assays can potentially reduce analysis time for indicator bacteria such as coliforms. In the present study, we developed
a simple presence/absence (P/A) bioluminescence procedure for rapid detection of coliform bacteria in groundwater-based drinking
water. The bioluminescence procedure targeting β-d-galactosidase activity in coliform bacteria was based on hydrolysis of 6-O-β-galactopyranosyl-luciferin. Bacteria immobilized on membrane filters were enriched for 6–8 h in selective media containing
isopropyl-β-d-thiogalactopyranoside (IPTG) to induce β-d-galactosidase activity in coliform bacteria. The equivalent of approximately 300 E. coli cells was required for bioluminescence detection of β-d-galactosidase activity. In comparison, PCR based detection of E. coli in drinking water required approximately 30 target cells. Analysis of contaminated drinking water samples showed comparable
results for coliform bacteria using traditional multiple-tube fermentation, Colilert-18, and the bioluminescence procedure.
Aeromonas hydrophila or indigenous groundwater bacteria did not interfere with the procedure. The bioluminescence procedure can be combined with
commercial substrates such as Fluorocult or Colilert-18, and will allow the detection of one coliform in 100 ml drinking water
within one working day. The results suggest the bioluminescence assays targeting β-d-galactosidase activity may be used for or for early warning screening of drinking water and/or rapid identification of contaminated
drinking water wells. 相似文献
20.
The purpose of the study was to investigate the effect of hydroxypropyl beta cyclodextrin (HPβCD) on aqueous solubility, stability,
and in vitro corneal permeation of acyl ester prodrugs of ganciclovir (GCV). Aqueous solubility and stability of acyl ester
prodrugs of Ganciclovir (GCV) were evaluated in pH 7.4 isotonic phosphate buffer solution (IPBS) in the presence and absence
of HPβCD. Butyryl cholinesterase-mediated enzymatic hydrolysis of the GCV prodrugs was studied using various percentage w/v
HPβCD. In vitro corneal permeation of GCV and its prodrugs (with and without 5% HPβCD) across isolated rabbit cornea was studied
using side-by-side diffusion cells. HPβCD-prodrug complexation was of the AL type with values for complexation constants ranging between 12 and 108 M−1. Considerable improvement in chemical and enzymatic stability of the GCV prodrugs was observed in the presence of HPβCD.
The stabilizing effect of HPβCD was found to depend on the degree of complexation and the degradation rate of prodrug within
the complex. Five percent w/v HPβCD was found to enhance the corneal permeation of only the most lipophilic prodrug GCV dibutyrate
(2.5-fold compared with 0% HPβCD). All other prodrugs showed little or no difference in transport in the presence of 5% w/v
HPβCD. Agitation in the donor chamber largely influenced the transport kinetics of GCV dibutyrate across cornea. Results indicate
the presence of an unstirred aqueous diffusion layer at the corneal surface that restricts the transport of the highly lipophilic
GCV dibutyrate prodrug. HPβCD improves corneal permeation by solubilizing the hydrophobic prodrug and delivering it across
the mucin layer at the corneal surface. 相似文献