Biomedical subjects

Y Q Xia

Publications and source records attributed to Y Q Xia.

15 recordsLinked to original sources

PubMed2004

Adsorptive separation of hemoglobin by molecularly imprinted chitosan beads.

A simply hemoglobin (Hb) molecularly imprinted polymer (MIP) was prepared using Hb as the imprinted molecule, acrylamide as the functional monomer and cross-linked chitosan beads as the supporting matrix. The MIP was achieved by entrapment of the selective soft polyacrylamide gel in the pores of the cross-linked chitosan beads by letting acrylamide monomer and the protein diffuse into the pores of chitosan beads before starting the polymerization. The chitosan beads were freed from the surrounding polyacrylamide gel by washing. The Langmuir and Freundlich adsorption models were applied to describe the equilibrium isotherms. Langmuir analysis showed that an equal class of adsorption was formed in the MIP and the adsorption equilibrium constant and the maximum adsorption capacity were evaluated. The MIP has much higher adsorption capacity for Hb than the non-imprinted polymer with the same chemical composition, and the MIP also has a higher selectivity for the imprinted molecule. The MIP can be reused in an easy way and the reproduction coefficient was approximately 100% at low concentration.

Acrylamide↗

PubMed2001

Glycosylasparaginase activity requires the alpha-carboxyl group, but not the alpha-amino group, on N(4)-(2-Acetamido-2-deoxy-beta-D-glucopyranosyl)-L-asparagine.

Glycosylasparaginase catalyzes the hydrolysis of the N-glycosylic bond in N(4)-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-L-asparagine in the catabolism of N-linked oligosaccharides. A deficiency, or absence, of enzyme activity gives rise to aspartylglycosaminuria, the most common disorder of glycoprotein metabolism. The enzyme catalyzes the hydrolysis of a variety of asparagine and aspartyl compounds containing a free alpha-carboxyl group and a free alpha-amino group; computational studies suggest that the alpha-amino group actively participates in the catalytic mechanism. In order to study the importance of the alpha-carboxyl group and the alpha-amino group on the natural substrate to the reaction catalyzed by the enzyme, 14 analogues of the natural substrate were studied where the structure of the aspartyl group of the substrate was changed. The incremental binding energy (DeltaDeltaGb) for those analogues that were substrates was calculated. The results show that the alpha-amino group may be substituted with a group of comparable size, for the alpha-amino group contributes little, if any, to the transition state binding energy of the natural substrate. The alpha-amino group position acts as an "anchor" in the binding site for the substrate. On the other hand, the alpha-carboxyl group is necessary for enzyme activity; removal of the alpha-carboxyl group or changing it to an alpha-carboxamide group results in no hydrolysis reaction. Also, N-acetyl-D-glucosamine is not sufficient for binding to the active site for efficient hydrolysis by the enzyme. These results provide supporting evidence for a proposed intramolecular autoproteolytic activation reaction for the enzyme. However, the results raise a question as to an important role for the alpha-amino group in the catalytic mechanism as indicated in computational studies.

Acetylglucosamine↗

PubMed2001

Parallel extraction columns and parallel analytical columns coupled with liquid chromatography/tandem mass spectrometry for on-line simultaneous quantification of a drug candidate and its six metabolites in dog plasma.

A method with parallel extraction columns and parallel analytical columns (PEC-PAC) for on-line high-flow liquid chromatography/tandem mass spectrometry (LC/MS/MS) was developed and validated for simultaneous quantification of a drug candidate and its six metabolites in dog plasma. Two on-line extraction columns were used in parallel for sample extraction and two analytical columns were used in parallel for separation and analysis. The plasma samples, after addition of an internal standard solution, were directly injected onto the PEC-PAC system for purification and analysis. This method allowed the use of one of the extraction columns for analyte purification while the other was being equilibrated. Similarly, one of the analytical columns was employed to separate the analytes while the other was undergoing equilibration. Therefore, the time needed for re-conditioning both extraction and analytical columns was not added to the total analysis time, which resulted in a shorter run time and higher throughput. Moreover, the on-line column extraction LC/MS/MS method made it possible to extract and analyze all seven analytes simultaneously with good precision and accuracy despite their chemical class diversity that included primary, secondary and tertiary amines, an alcohol, an aldehyde and a carboxylic acid. The method was validated with the standard curve ranging from 5.00 to 5000 ng/mL. The intra- and inter-day precision was no more than 8% CV and the assay accuracy was between 95 and 107%.

Alcohols↗

PubMed2001

Glycosylasparaginase inhibition studies: competitive inhibitors, transition state mimics, noncompetitive inhibitors.

Glycosylasparaginase catalyzes the hydrolysis of the N-glycosylic bond between asparagine and N-acetylglucosamine in the catabolism of N-linked glycoproteins. Previously only three competitive inhibitors, one noncompetitive inhibitor, and one irreversible inhibitor of glycosylasparaginase activity had been reported. Using human glycosylasparaginase from human amniotic fluid, L-aspartic acid and four of its analogues, where the alpha-amino group was substituted with a chloro, bromo, methyl or hydrogen, were competitive inhibitors having Ki values between 0.6-7.7 mM. These results provide supporting evidence for a proposed intramolecular autoproteolytic activation reaction. A proposed phosphono transition state mimic and a sulfo transition state mimic were competitive inhibitors with Ki values 0.9 mM and 1.4 mM, respectively. These results support a mechanism for the enzyme-catalyzed reaction involving formation of a tetrahedral high-energy intermediate. Three analogues of the natural substrate were noncompetitive inhibitors with Ki values between 0.56-0.75 mM, indicating the presence of a second binding site that may recognize (substituted)acetamido groups.

Acetylglucosamine↗

PubMed2000

Peripheral electric stimulation inhibits morphine-induced place preference in rats.

Conditioned place preference (CPP) is a commonly used model to detect rewarding effect of drugs. To observe the effect of peripheral electric stimulation (PES) on morphine-induced CPP, we trained the rats with morphine in a CPP paradigm. Twelve hours before the testing phase, rats were given PES via stainless-steel needles with frequencies of 2, 100, or 2/100 Hz, respectively. PES of 2 and 2/100 Hz significantly decreased CPP in morphine-trained animals in a naloxone reversible manner, while PES of 100 Hz, foot shock, needle insertion, or plain restraining, showed no effect. Thus, PES with a low-frequency component (2 Hz) could specifically inhibit the expression of morphine-induced CPP, presumably via activation of opioid receptors.

Acupuncture Therapy↗

PubMed2000

Ternary-column system for high-throughput direct-injection bioanalysis by liquid chromatography/tandem mass spectrometry.

As a continuation of our efforts to improve our high-flow on-line bioanalytical approach for high-throughput quantitation of drugs and metabolites in biological matrices by high-performance liquid chromatography (LC) and tandem mass spectrometry (MS/MS), we have developed a ternary-column on-line LC/MS/MS system with dual extraction columns used in parallel for purification and an analytical column for analysis. The advantage of the dual extraction column system is that sample analysis can take place in one of the extraction columns while the other column is being equilibrated. Thus, the equilibration time does not add to the run time, hence shortening the injection cycle time and increasing the sample throughput. Moreover, the use of two extraction columns in parallel increases the number of samples that can be injected before the system fails due to an overused extraction column. Such a system has successfully been used to develop and validate a positive ion electrospray LC/MS/MS bioanalytical method for the quantitative determination of a guanidine-containing drug candidate in rat plasma. The system used for this work utilized two Oasis HLB extraction columns (1 x 50 mm, 30 microm), one C18 analytical column (3.9 x 50 mm, 5 microm), a ten-port switching value and a tandem mass spectrometer. The on-line analysis was accomplished by the direct injection of 10 microL of the sample, obtained by mixing a rat plasma sample 1:1 with an aqueous internal standard solution. Selected reaction monitoring (SRM) was utilized for the detection of the analyte and internal standard. The standard curve range was 1.00-200 ng/mL. The intra- and inter-day precision and accuracy were within 6.6%. The on-line purification step lasted for only 0.3 min and total run time was only 1.6 min.

Animals↗

PubMed2000

Bioanalytical method validation design for the simultaneous quantitation of analytes that may undergo interconversion during analysis.

In the analysis of post-dose biological samples for quantitative determination of two analytes that can potentially undergo interconversion, it is essential to minimize the interconversion during the multiple steps of the bioanalytical method. However, even after optimizing the conditions of each step, some interconversion may be unavoidable. Even then, a method can be developed for the accurate simultaneous determination of the two analytes in post-dose biological samples if the composition, in terms of the ratio of the concentrations of the two analytes, of the calibration standards and quality control (QC) samples are selected judiciously, in relation to the composition of the unknown samples to be analyzed. As an example of such interconverting analytes, a delta-hydroxy acid compound (analyte 1) and its delta-lactone (analyte 2) were selected as model compounds that can potentially undergo interconversion. The effects of changing the relative concentrations of the two analytes in QC samples vis-à-vis the calibration standards on the performance of the method under conditions were investigated where: (a) the interconversion between the two analytes was minimized; (b) the conversion of analyte 2 to analyte 1 was enhanced; (c) the interconversion between the two analytes was enhanced. The results showed that the method performance, as measured by the accuracy and precision of the QC samples, was not acceptable when the ratio of concentration of analyte 1 to that of analyte 2 in the QC samples was different from that in the calibration standards and the conditions used facilitated the conversion of one analyte to the other. However, when the relative concentration of the two analytes in the QC samples was identical to that of the calibration standards, the method performance was acceptable under all three conditions of interconversion. This was because the same degree of interconversion took place in the QC samples and calibration standards. The purpose of QC samples in bioanalytical methods is to gauge how the method will perform for the analysis of post-dose test samples and hence, ideally, the relative concentrations of the analytes in QC samples, should be selected to mimic the anticipated concentrations in the test samples. However, the relative concentrations of the analytes in test samples may not be known a-priori, or may change from sample to sample; therefore, it is not always possible to construct QC samples that exactly mimic the relative concentrations of analytes in the test samples. Thus, in order to cover the variety of test samples, the method should include, in addition to QC samples that contain the analytes at the same relative concentration as in the calibration standards, QC samples with relative concentrations that are different from those in the calibration standards, including those that contain only analyte 1 and only analyte 2. In addition, the conditions adopted for the method should favor the minimization of the conversion of the analyte that is expected to be the major component in the post-dose test samples.

Chemistry, Pharmaceutical↗

PubMed1999

A versatile system of high-flow high performance liquid chromatography with tandem mass spectrometry for rapid direct-injection analysis of plasma samples for quantitation of a beta-lactam drug candidate and its open-ring biotransformation product.

A bioanalytical method has been developed and validated for quantitation of a beta-lactam drug candidate and its open-ring biotransformation product utilizing high-flow high-performance liquid chromatography (HPLC) for on-line purification of plasma samples and electrospray tandem mass spectrometry for detection and quantitation. The HPLC system used two columns: an Oasis column (1 x 50 mm, 30 microm) as the on-line extraction column and a conventional C18 column (2 x 50 mm, 5 microm) as the analytical column. Each plasma standard or quality control (QC) sample (50 microL) was mixed with 50 microL of a working solution of the internal standard in aqueous 0.5 M ammonium acetate (pH 4.0). Portions (10 microL) of these samples were then injected into an Oasis column with a mobile phase consisting of 100% aqueous 1 mM formic acid at a high flow rate (4.0 mL/min), with the effluent from the Oasis column directed to waste and not to the mass spectrometer. After the purification step, the Oasis column effluent was directed to the analytical column and the mass spectrometer and the analytes were eluted with methanol/aqueous 1 mM formic acid (70:30) at a flow rate of 1.0 mL/min. The total analysis time was 1.6 min per sample. The standard curve range was 0.980 to 250 ng/mL. The accuracy, inter-day precision and intra-day precision were within 10% for both compounds.

Biotransformation↗

PubMed1999

A simple liquid-liquid extraction with hexane for low-picogram determination of drugs and their metabolites in plasma by high-performance liquid chromatography with positive ion electrospray tandem mass spectrometry.

Four sensitive, specific and accurate methods, based on high-performance liquid chromatography (HPLC) with positive ion electrospray tandem mass spectrometry (MS/MS) coupled with liquid-Liquid extraction (LLE), have been developed and validated for the low-picogram determination of two drug candidates and a metabolite (compounds I-III) in human, monkey and rat plasma. In the LLE procedure, hexane or a mixture of hexane and methyl t-butyl ether was used to isolate these compounds from plasma of the different species after basification of each biological sample with sodium carbonate. The reconstituted extracts were then injected into a positive ion electrospray LC/MS/MS system for the quantitative analysis. The lower limit of quantitation of the methods ranged from 20 to 200 pg/mL. The use of hexane for the LLE proved to be simple, rapid and reproducible, and provided very clean extracts with little interference. The inter- and intra-day precision for the four methods was within 9%, and the accuracy was in the range 94-107%. The effect of pH on the isomerization of I (E-isomer) to its Z-isomer (II) showed that the rate of isomerization increased with decrease in pH and that there was no isomerization at pH >/=6.

Animals↗

PubMed1998

The use of high-flow high performance liquid chromatography coupled with positive and negative ion electrospray tandem mass spectrometry for quantitative bioanalysis via direct injection of the plasma/serum samples.

Two bioanalytical methods have been developed and validated utilizing high flow high performance liquid chromatography (HPLC) for on-line purification of plasma and serum samples and electrospray tandem mass spectrometry for detection and quantitation. Each plasma or serum sample, after mixing with an aqueous solution of the internal standard, was injected into a small diameter (1 x 50 mm) column packed with large particles of OASIS (30 microns), with a 100% aqueous mobile phase at a high flow rate (3-4 mL/min). The combination of the high linear speed (6-8 cm/s) of the aqueous mobile phase and the large particle size resulted in the rapid passage of the proteins and other large biomolecules through the column while the small-molecule analytes were retained on the column. During this purification period, the HPLC effluent was directed to waste. After the purification step, the HPLC mobile phase was rapidly changed from 100% aqueous to < or = 100% organic, the flow was reduced to 0.5-0.8 mL/min, and the column effluent was directed towards the mass spectrometer. The small molecule analytes were eluted during this period. In the method developed and validated for the quantitative determination of compound I in rat plasma (method A), the same OASIS column (1 x 50 mm, 30 microns) served as the purification and analytical (elution) column. In the method developed for the simultaneous determination of pravastatin and its positional isomer biotransformation product (SQ-31906) in human serum (method B), the purification column was connected to a conventional C18 analytical column (3.9 x 50 mm, 5 microns) to achieve the required chromatographic separation between the two isomers. For method A, where 50 microL of rat plasma mixed 1:1 with water containing the internal standard was injected, the standard curve range was 1 to 1,000 ng/mL. For method B, where 200 microL of a human serum sample mixed 4:1 with water containing the internal standard was injected, the standard curve range was 0.5 to 100 ng/mL. The total analysis time for each method was < or = 5 min per sample. The accuracy, inter-day precision and intra-day precision were within 10% for both methods.

Animals↗

PubMed1988