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PubMed1999

[Chromatographic properties of tetradecylamine bonded stationary phase for reversed-phase liquid chromatography].

A novel bonded stationary phase, tetradecylamine bonded stationary phase (TABP), for reversed-phase HPLC was prepared by bonding 1-tetradecylamine to YWG-80 silica gel through 3-glycidoxypropyltrimethoxysilane. Hydrophobicity, selectivity and silanophilic activity of TABP were evaluated by using aromatic compounds as analytes and methanol-water as binary mobile phase. The orgainc components including acidic, basic and neutral aromatic analytes could be separated satisfactorily with excellent selectivity and peak shape. The relative retention alpha for ethylbenzene-toluene was found to be 1.66 and the asymmetry factors of basic aniline, p-toluidine and N,N-dimethylaniline were found to be 1.26, 1.21 and 1.10, respectively, with the V(methanol):V(water) = 55:45 mobile phase. Aniline was eluted before phenol due to the suppress of the ion exchange activity of residual silanols by the internal masking interaction.

Amines↗
PubMed2002

[Formation of the third phase and spectroscopic research of the loading organic phases in the gold extraction].

The formation of the third phase during the gold extraction from cyanide solution by quaternary ammonium, tetradecyldimethylbenzylammonium chloride (TDMBAC) was studied by 198Au tracer method. The microstructure of the organic phases was investigated by FTIR, 31P-NMR. With the increasing of gold concentration, aggregations were formed in the gold-loaded organic phase. With the further increasing of gold concentration, the organic phase turned into two layers. Gold, water, and tri-n-butyl phosphate (TBP) were mainly in the down organic phase. In the down layer, distinct O-H stretching vibration peak was found by FTIR; the frequency of P=O stretching vibration shifted to the low frequency; 31P-NMR chemical shift was markedly shifted to the high frequency field. The radius of aggregation was measured by the dynamic laser scattering (DLS) method.

English Abstract↗
PubMed2003

[Organic acids conversion in methanogenic-phase reactor of the two-phase anaerobic process].

Experiments on organic acids conversion in methanogenic-phase of the two-phase anaerobic process were conducted. The results showed that when the acidogenic-phase was in mixed acids fermentation and its load was 41.5 kg/(m3-d), the load of methanogenic-phase was 6.05 kg/(m3.d), the substrate conversion velocity of bacteria inhabited at the same height of UASB reactor as follows: acetic acid > ethanol > butyric acid > propionic acid; ethanol-utilized microbial had the same pH range as methanogenic bacteria; ethanol-type fermentation was the optimal acidogenci-type fermentation for the two-phase anaerobic process; the conversion of acetic acid was rather high, but the others organic acids will be convert to acetic acid make it the rate-limiting step for anaerobic degradation.

Acetic Acid↗
PubMed2005

[Repair of articular cartilage defects with "two-phase" tissue engineered cartilage constructed by autologous marrow mesenchymal stem cells and "two-phase" allogeneic bone matrix gelatin].

OBJECTIVE: To investigate the effect of "two-phase" tissue engineered cartilage constructed by autologous marrow mesenchymal stem cells(MSCs) and allogeneic bone matrix gelatin(BMG) in repairing articular cartilage defects. METHODS: Thirty-two New Zealand white rabbits were involved in the experiment. "Two-phase" allogeneic BMG scaffold (one side of porous cancellous bone and the other side of cortical bone; 3 mm both in diameter and in thickness) was prepared from iliac bone and limb bone of 5 rabbits by sequentially chemical method. The MSCs were separated from 18 New Zealand white rabbits and induced to express chondrocytic phenotype. The chondrocyte precursor cells were seeded onto "two-phase" allogeneic BMG to construct tissue engineering cartilage. Masson's trichrome staining, PAS staining and scanning electronic microscopic observation were carried out at 1, 3 and 5 weeks. The defects of full thickness articular cartilage (3 mm both in diameter and in depth) were made at both sides of femoral medial condyles in 27 rabbits (including 18 of separated MSCs and the remaining 9). The defects were repaired with the tissue engineered cartilage at the right side (group A, n = 18), with BMG at the left side (group B, n = 18), and without any implant at both sides in the remaining 9 rabbits as a control (group C, n = 18). After 1, 3 and 6 months, the 6 specimens of femoral condyles were harvested in 3 groups, respectively. Gross observation, Masson's trichrome and Alcian blue staining, modified Wakitani scoring and in situ hybridization of collagen type I were carried out to assess the repair efficacy of tissue engineered cartilage. RESULTS: The "two-phase" BMG consisted of the dense cortical part and the loose cancellous part. In cancellous part, the pore size ranged 100-800 microm, in which the chondrocyte precursor cells being induced from MSCs proliferated and formed the cell-rich cartilaginous part of tissue engineered cartilage. In cortical part, the pore size ranged 10-40 microm, on which the cells arranged in a layer and formed the hard part of subchondral bone. After 1 month of transplantation, the cartilage and subchondral bone were regenerated in group A; during observation, the regenerated cartilage gradually thinned, but defect was repaired and the structure of the articular surface and subchondral bone was in integrity. In groups B and C, defects were not repaired, the surrounding cartilage of defect was abrased. According to the modified Wakitani scoring, the indexes in group A were significantly higher than those in group B and C (P < 0.01) except the thickness of cartilage at 6 months. The positive cell rate of in situ hybridization for collagen type II in group A was also higher than those in groups B and C (P < 0.01). CONCLUSION: "Two-phase" allogeneic BMG is a prospective scaffold for tissue engineered cartilage, which combines with autologous chondrocyte precursor cells induced from MSCs to construct the tissue engineering cartilage. The tissue engineered cartilage can repair defects of articular cartilage and subchondral bone.

Animals↗
PubMedDate not supplied

Identification of phase-I and phase-II metabolites of fluphenazine in rat bile. Intact glucuronide and sulfate conjugates.

In rat bile following ip administration of fluphenazine (FLU) dihydrochloride (20 mg/kg body weight), phase-I metabolites 7-hydroxyfluphenazine (7-HOFLU) and FLU sulfoxide (FLUSO), together with unmetabolized FLU, were isolated and identified by HPLC and fast atom bombardment spectrometry (FAB/MS) and also by comparison with authentic compounds. Two intact glucuronide conjugates of FLU were isolated and identified as phase-II metabolites: 7-hydroxyfluphenazine ring glucuronide (glucuronic acid linked to the aromatic hydroxyl group of 7-hydroxyfluphenazine), and FLU glucuronide (glucuronide linked to the aliphatic group of the side chain of FLU) by HPLC and FAB/MS in comparison with authentic compounds. Further confirmed by FAB/MS were several sulfate conjugates of FLU that were isolated and identified indirectly as phase-II sulfate metabolites: FLU sulfate, 7-hydroxyfluphenazine sulfate and/or 7-hydroxyfluphenazine ring sulfate, and FLU sulfoxide sulfate by HPLC and FAB/MS; their aglycones were identified after sulfatase hydrolysis as FLU, 7-HOFLU and FLUSO. A further phase-II metabolite, for which no authentic standard was available, was tentatively identified as a monoglucuronide of dihydroxy derivative of FLU. To our knowledge, this report provides the first direct evidence of the presence of intact phase-II metabolites of FLU in rat bile.

Animals↗
PubMed1990

Relationship of bcr breakpoint to chronic phase duration, survival, and blast crisis lineage in chronic myelogenous leukemia patients presenting in early chronic phase.

Strong evidence implicates fusion of control elements and 5' sequences of the bcr gene of chromosome 22 with 3' sequences of the c-abl gene of chromosome 9 in the pathogenesis of Ph-positive and certain cases of Ph-negative chronic myelogenous leukemia (CML). Since this fusion gene gives rise to a chimeric tyrosine protein kinase with transforming potential, and since the bcr exon contribution to this chimeric protein is variable, the question has arisen as to whether bcr breakpoint location and bcr exon contribution could influence the clinical course of CML. Prior studies have yielded conflicting results on this point. Here we have looked, in a manner approximating a prospective analysis, at the relation of bcr breakpoint localization to the duration of chronic phase, total survival, and blast crisis phenotype in 81 patients presenting in the chronic phase of CML. We have found no significant differences in chronic phase duration or total survival among patients with breakpoints in the three major subregions of a breakpoint cluster region within the bcr gene. These findings indicate that chronic phase duration and total survival cannot be predicted from bcr breakpoint for CML patients presenting in chronic phase and suggest that unknown oncogenic events determining the onset of blast crisis are the prime determinants of prognosis. Combined analysis of blast crisis cell lineage in our patients and patients presented in a previous study has revealed an overall ratio of myeloid:lymphoid (M:L) crisis of 3.4:1, but a striking predominance of myeloid crisis in patients with breakpoints in subregion 2 (M:L of 9:1), and a lower than expected M:L ratio (1.6:1) among patients with breakpoints in subregion 3 (P for subregion 2 versus 3 = .012; subregions 0,1,2 versus 3 = .012; subregions 0,1,3 versus 2 = .032). The molecular basis for this divergence from the anticipated M:L ratio in patients with breakpoints in bcr subregions 2 and 3 is unknown.

Adult↗
PubMed1988

[Plasmid profile types of virulent and avirulent strains of phase-I Shigella sonnei and their rough phase-II and R-form variants].

The study of S. sonnei in phase I, irrespective of their virulence, has revealed the existence of at least 3 types of profiles of large plasmids: (I)A having a single plasmid with a molecular weight of about 120 MD; (I)B having, alongside plasmid pSS120, a plasmid with a molecular weight of about 60 MD; (I)C, represented only by vaccine strain 6S, having three plasmids with molecular weights of about 80, 60 and 37 MD. The plasmid profiles of rough S. sonnei in phase II are characterized by the absence of large plasmids with a molecular weight of 120-80 MD, typical of bacteria in phase I, and can be in their turn subdivided, in accordance with the type of the initial culture, into three subvariants (II)A, (II)B and (II)C. The plasmid profiles of rough S. sonnei (R-forms and phase II) completely coincide. The biosynthesis of the specific antigen of S. sonnei in phase I can be determined by smaller derivatives obtained from large plasmid pSS120 by deletion (e.g., by a plasmid with a molecular weight of about 80 MD, such as plasmid pSS80).

Animals↗
PubMed1987

Monoclonal antibodies used in solid-phase and liquid-phase assays, as exemplified by progesterone assay.

Two immunoglobulins secreted by hybridoma cell lines have been systematically investigated to determine if they could be used in solid-phase assays to give results comparable with those obtained by conventional liquid-phase radioimmunoassay (RIA). The antibodies, BQ.1 and BQ.2, bind with high specificity to the steroid hormone progesterone. The affinity constants, Ka, to 125I-labeled progesterone derivatives are 1.1 X 10(11) L/mol and 9.1 X 10(9) L/mol, respectively. Progesterone inhibited the binding of radioiodinated derivatives (amides of tyramine, histamine, and tyrosine methylester with 11 alpha-progesterone hemisuccinate) equally well. For solid-phase assays, we immobilized antibody BQ.1 via Protein A to different polystyrene surfaces (about 30 pg per tube at 50% inhibition of radiolabeled tracer). Under these conditions, the performance of this antibody for the quantification of progesterone was equivalent to that obtained in RIA. For the immobilized antibody BQ.2, only 1/10 of the amount used for optimal results in RIA was required in solid-phase assays. Binding of either antibody to the antigen was undiminished after several freezing-thawing cycles. When immobilized on solid matrices, both antibodies retained up to 95% of their binding properties for one year. Thus high-affinity, high-specificity monoclonal antibodies can be obtained for haptens and, when suitably immobilized, can be used in solid-phase assays with results equal to or better than those obtained with liquid RIA.

Antibodies, Monoclonal↗
PubMed1983

Different antigenic properties of lipopolysaccharides isolated from Coxiella burnetii in phase I and pure phase II.

Lipopolysaccharides (LPSs) isolated from Coxiella burnetii cells in phase I and pure phase II behaved as antigens reacting in serological tests (passive haemolysis, passive haemolysis inhibition, complement-fixation, immunoprecipitation) with sera of rabbits immunized with C. burnetii strains in the corresponding phase. No cross-reactions were observed between LPSs isolated from C. burnetii in phase I and phase II, respectively.

Complement Fixation Tests↗
PubMed1994

Linear solvation energy relationships in reversed-phase liquid chromatography. Examination of RP-8 stationary phases for measuring lipophilicity parameters.

Molecular lipophilicity can be expressed by log P or more conveniently by log kw, determined by the so-called "shake-flask" (SF) technique or by reversed-phase high-performance liquid chromatography (RP-HPLC), respectively. In the present study, the lipophilicity of a large set of solutes was measured by RP-HPLC on a silanol-deactivated octylsilane (OS) reversed phase, namely Inertsil OS (IN). IN lipophilicity parameters were found to be fairly well correlated with log P. A Linear Solvation Energy Relationship (LSER) study revealed that cavity term (modeled by intrinsic molar volume, V1) and hydrogen-bond acceptor capacity (modeled by the solvatochromic parameter beta) of solutes are the most important factors affecting retention on Inertsil OS. The comparison of LSER model for retention on Inertsil with those obtained for two other silanol-deactivated OS stationary phases, namely Deltabond OS and RP Select B, shows that alpha (H-bond donor activity of solutes) can be used as a parameter able to discriminate the behaviour of different stationary phases. QSAR studies indicated that lipophilicity parameters determined on silanol-deactivated OS phases have potential in medicinal chemistry applications.

Chromatography, High Pressure Liquid↗
PubMed1977

Effect of an aqueous phase on the solubility of cholesterol in an oil phase.

In the absence of water, the solubility of cholesterol in triolein at 21 degrees C was 2.8%. When water was added to the system, the solubility of cholesterol in the oil phase decreased to 1.9%, and cholesterol monohydrate precipitated. The precipitation of the sterol evidently resulted from the excess concentration of the surface-active cholesterol at the interface, allowing the rapid interaction of water with cholesterol and the resulting formation of cholesterol monohydrate with its attendant lower energy and less soluble crystalline lattice. The ternary phase diagram for the system cholesterol--triolein--water, modified to include cholesterol monohydrate formation with the consequent decrease in sterol solubility; differs from the previously reported phase diagram. Other cholesterol--oil--aqueous systems related to biologically important lipids were studied. Cholesteryl oleate was more soluble than cholesterol in triolein (23% at 21 degrees C), and this value did not decrease when water was present. Water caused cholesterol to precipitate from cholesteryl linoleate at 37 degrees C. Thus crystalline cholesterol may be present in lipids found in atherosclerotic plaques at a lower concentration of free cholesterol than had been predicted previously. In another experiment, a micellar taurocholate solution precipitated cholesterol from triolein and from corn oil. These effects of aqueous systems suggest the possibility of cholesterol precipitation from dietary fat when it becomes mixed with water in the diet or stomach, or with the micellar phase in the intestine. Plant sterols were precipitated also from oil solutions by an aqueous phase. Water-induced sterol precipitation is a phenomenon that could occur in a variety of biological systems, and may be applicable to sterols in general.

Cholesterol↗
PubMed1995

Comparisons of phase I and phase II in vitro hepatic enzyme activities of human, dog, rhesus monkey, and cynomolgus monkey.

The metabolism of probe substrates of phase I and phase II enzymes in vitro were compared in hepatic subcellular fractions from humans, cynomolgus monkeys, rhesus monkeys, and beagle dogs. These studies were undertaken to compare the suitability of these species as models of metabolism in drug development. Eight cytochrome P450-dependent activities were measured in microsomal incubations: ethoxyresorufin O-deethylase, coumarin 7-hydroxylase, tolbutamide 4-hydroxylase, S-mephenytoin 4'-hydroxylase, bufuralol 1'-hydroxylase, N-nitrosodimethylamine N-demethylase, midazolam 1'-hydroxylase, and erythromycin N-demethylase. Seven phase II activities were determined in the appropriate subcellular fractions:acetaminophen UDP-glucurono-syltransferase, acetaminophen sulfotransferase, 17 alpha-ethinylestradiol UDP-glucuronosyltransferase, 17 alpha-ethinylestradiol sulfotransferase, 6-mercaptopurine methylase, dichloronitrobenzene (DCNB) glutathione S-transferase, and isoniazid N-acetylase. Hepatic subcellular fractions from cynomolgus and rhesus monkeys showed significantly higher activities than those from humans for ethoxyresorufin O-deethylase, bufuralol 1'-hydroxylase, midazolam 1'-hydroxylase, erythromycin N-demethylase, acetaminophen UDP-glucuronosyltransferase, acetaminophen sulfotransferase, and tolbutamide 4-hydroxylase. Cynomolgus monkey had higher activity than humans and rhesus monkeys for S-mephenytoin 4'-hydroxylase erythromycin N-demethylase. Rhesus monkey and human cytosol displayed an apparent genetic polymorphism in the N-acetylation of isoniazid, whereas cynomolgus monkey cytosol did not. All other monkey activities were not significantly different than human. Dog subcellular fractions showed higher activity than humans for midazolam 1'-hydroxylase, erythromycin N-demethylase, acetaminophen UDP-glucuronosyltransferase, acetaminophen sulfotransferase, 17 alpha-ethinylestradiol sulfotransferase, and DCNB glutathione S-transferase. Furthermore, dog samples had significantly lower activity for coumarin 7-hydroxylase and 6-mercaptopurine methylase, and no detectable activity for tolbutamide 4-hydroxylase or isoniazid N-acetylase. All other activities were not significantly different from human. These results reveal minor differences between the cynomolgus and rhesus monkey in drug metabolism capacities in vitro, but both species are generally more metabolically active than humans in both phase I and phase II metabolism, whereas dogs had more diverse deviations from humans.

Animals↗
PubMed1996

Amyloid beta toxicity consists of a Ca(2+)-independent early phase and a Ca(2+)-dependent late phase.

Amyloid beta protein (A beta), which accumulates in the senile plaques in the brain of Alzheimer's patients, is cytotoxic to neurons. A modified 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, in which a yellow redox dye, MTT, is reduced to purple formazan, is very sensitive to the effect of A beta. In primary hippocampal cultures, inhibition of MTT reduction starts within 2 h after the addition of low concentrations of A beta and reaches a plateau in 12 h. This effect of A beta is not blocked by Ca2+ channel blockers or in Ca(2+)-free medium. In contrast, lactate dehydrogenase (LDH) release and trypan blue exclusion, which are indices of cell death, start 3 days after exposure to high concentrations of A beta and are blocked by Ca2+ channel blockers such as Co2+, nicardipine, and diltiazem. When A beta was washed out from the medium after 12 h, MTT reduction recovers and LDH release does not occur, suggesting that a long-lasting inhibition of the cellular redox system may be required to induce cell death. These observations demonstrate that A beta toxicity consists of two phases-a Ca(2+)-independent early phase and a Ca(2+)-dependent late phase- and that the early phase may be required to induce the late phase.

Amyloid beta-Peptides↗
PubMed1997

Crystal violet staining of Bordetella bronchiseptica colonies for differentiation of phase-I strains from variant strains in degraded phases.

After 2 days of growth on Brain heart infusion agar (BHIA) at 38 degrees C, phase-I colonies and degraded-phase colonies of Bordetella bronchiseptica could be differentiated by their ability to take up crystal violet (CV). Phase-I colonies in X mode, but not colonies in degraded phases (phases II, III, and rough) bound CV. Phenotypically-altered C-mode colonies (grown at 32 degrees C or lower temperatures) also lacked this ability. CV staining offers an easy method for the recognition of different colony types that appear identical when observed on BHIA.

Animals↗
PubMed1990

Acute-phase response of human hepatocytes: regulation of acute-phase protein synthesis by interleukin-6.

Human hepatocytes in primary culture were used as a model system to investigate the mechanism(s) involved in the induction of the acute-phase response in human liver. Hepatocytes were incubated with increasing amounts of recombinant human interleukin-1 beta, recombinant interleukin-6 and tumor necrosis factor-alpha. Synthesis of C-reactive protein was studied at the mRNA and protein levels. Only recombinant interleukin-6 was capable of inducing C-reactive protein-mRNA and C-reactive protein-protein synthesis. Also, fibrinogen and alpha-1-antitrypsin synthesis measured by immunoprecipitation with specific antisera increased in a dose-dependent, time-dependent manner, whereas albumin synthesis decreased to about 50% of controls. Maximal effects were observed at 100 to 300 units of recombinant interleukin-6/ml culture medium after 20 hr of incubation. Although the synthetic glucocorticoid dexamethasone slightly modulated the effect of recombinant interleukin-6, it was not an absolute requirement for the induction of acute-phase protein synthesis in human hepatocytes. In pulse-chase experiments it was shown that the time course of the disappearance of the acute-phase proteins from the cells and their appearance in the medium is not influenced by recombinant interleukin-6. This finding suggests that recombinant interleukin-6 exerts its regulatory effect on acute-phase protein synthesis at the pretranslational level.

Acute-Phase Proteins↗
PubMed2004

Response to flumazenil in the late luteal phase and follicular phase of the menstrual cycle in healthy control females.

RATIONALE: Animal studies of short-term progesterone administration and withdrawal model the natural increase and abrupt decrease in progesterone levels which occur in the late luteal phase (LP) of the human menstrual cycle (MC). Previously, studies in animals have shown that abrupt cessation of chronic or short-term progesterone administration results in pharmacological changes at the GABAA receptor, resulting in altered sensitivity to GABAA receptor neuromodulators such as benzodiazepines and flumazenil, a GABAA receptor antagonist. OBJECTIVES: This study's goal was to compare the response to flumazenil in the follicular phase (FP) and late LP in female healthy controls (HCs). We postulated that HC females would exhibit a greater psychological and somatic response to flumazenil in the late LP, a period of progesterone withdrawal, compared to the FP. METHODS: Twelve healthy females, without history of psychiatric disorder, were randomized to receive two injections of a 2 mg bolus injection of flumazenil (one in the late LP and one in the FP) and two injections of placebo (one in the late LP and one in the FP). Following injection, subjects were asked to rate the occurrence and intensity of panic symptoms on the panic symptom scale (PSS). RESULTS: A main treatment effect was detected for the PSS score response after flumazenil injection (P=0.008). However, there was no significant treatment-by-phase interaction observed (P=0.449). CONCLUSIONS: These findings indicate that MC phase did not affect the response to flumazenil in HC females. This result is contrary to our hypothesis of altered sensitivity to flumazenil in the late LP.

Adult↗
PubMed2006

Molecularly imprinted polymers for solid-phase extraction and solid-phase microextraction: recent developments and future trends.

Molecularly imprinted polymers (MIPs) are synthetic polymers having a predetermined selectivity for a given analyte, or group of structurally related compounds, that make them ideal materials to be used in separation processes. In this sense, during past years a huge amount of papers have been published dealing with the use of MIPs as sorbents in solid-phase extraction, namely molecularly imprinted solid-phase extraction (MISPE). Although the majority of these papers were restricted to describe the use of different templates for different applications, several attempts proposing new alternatives to minimize the inherent drawbacks of the preparation and use of MIPs (i.e. template bleeding, tedious synthesis procedure, etc.) have been reported. Thus, this paper does not pretend to be a collection of MISPE-related papers but to give an overview on the significant attempts carried out during recent years to improve the performance of MIPs in solid-phase extraction. In addition, the use of MIPs packed in high performance liquid chromatography (HPLC) columns for the direct injection of crude sample extracts and the preparation of imprinted fibres for solid-phase microextraction will be also discussed.

Combinatorial Chemistry Techniques↗
PubMed1989

Luteal phase deficiency after completely normal follicular and periovulatory phases.

Luteal phase defect (LPD) accounts for a significant proportion of reproductive disorders, however its etiology is still debated. A prospective study was performed on 37 ovulatory women to determine whether LPD can occur in cycles characterized by completely normal folliculogenesis. Criteria for normal folliculogenesis included: a gradual rise of serum estradiol, a luteinizing hormone (LH) surge, the presence of a dominant follicle that disappeared, an increase of serum progesterone, and normal serum levels of prolactin, testosterone, dehydroepiandrosterone sulfate, follicle-stimulating hormone, and LH. Thirty of 37 women fulfilled the above mentioned strict criteria and underwent endometrial biopsy in the late luteal phase. Seven of 30 (23%) demonstrated a delay in endometrial development and all had normal hormonal and ultrasonographic parameters of folliculogenesis and ovulation. Women with delayed endometrial development demonstrated slightly longer follicular phases (17.0 +/- 1.1 versus 14.5 +/- 0.3 days). Perfectly normal follicular and periovulatory events may be followed by deficient luteal phases.

Adult↗