OBJECTIVE: To explain that receptor-ligand binding assay is an important drug screening method, which can quickly and inexpensively study the interactions between the targeted receptor and the potential ligands in vitro and provide information about the relative binding affinity of ligand-receptor. METHODS: The correlative literature and abroad were collected and analyzed. RESULTS AND CONCLUSION: The concept of MS binding assay based on mass spectrometric quantification of nonlabeled markers represents a promising alternative to conventional radioligands binding without inherent drawbacks. The technique will play an important role in the design and screening of receptor-targeting pharmaceuticals.

A chiral high-performance liquid chromatography method was developed for the simultaneous determination of ibuprofen enantiomers in dog plasma. It was used to study the pharmacokinetics in the Beagle dog after intravenous administration of racemic-ibuprofen, S-ibuprofen and R-ibuprofen. Ketoprofen was chosen as the internal standard. After a simple precipitation using methanol as the precipitating solvent, both analytes and IS were separated on a Kromasil 100-5CHI-TBB chiral column (250 mm x4.6 mm, 5 μm) with isocratic elution using acetonitrile - 20 mmol x L(-1) phosphate buffer (pH 3.0, containing 5% methanol) (6 : 4) as the mobile phase. The detection wavelength was 220 nm. Liner calibration curves for both of the ibuprofen enantiomers were over the concentration range from 0.5 to 50 μg x mL(-1) with a lower limit of quantification of 0.5 μg x mL(-1), the accuracies were all in standard ranges. The intra- and inter- assay precisions were all below 7%. The recovery rate was 93.1% to 100.4%. The experiments proved that the method was simple, rapid and sensitive. It can be used in the quantitative determination of ibuprofen enantiomers in dog plasma. The method was used to determine the concentration of ibuprofen enantiomers in Beagle dog plasma after a single intravenous administration of racemic-ibuprofen, S-ibuprofen and R-ibuprofen (9 mg x kg(-1)) and the pharmacokinetics parameters were calculated based on the concentration-time curves. The C(max) of S-ibuprofen in Beagle dog plasma after a single intravenous administration of racemic-ibuprofen, S-ibuprofen and R-ibuprofen were 30.8 ± 4.7, 46.1 ± 5.9 and 20.0 ± 2.6 μg x mL(-1), respectively. In terms of the exposure of active ingredient, it revealed a significant difference between the administration of S-ibuprofen and the other two groups. The systematical R- to S- chiral inversion was discussed. Comparing the pharmacokinetic parameters at different doses, chiral inversion were 70.1% ± 36.6% and 76.4% ± 36.2%, respectively, after intravenous administration of racemic- and R-ibuprofen. This study provides a theoretical basis for the safety of ibuprofen formula of injection drug.
Animals ; Chromatography, High Pressure Liquid ; Dogs ; Ibuprofen ; blood ; pharmacokinetics ; Stereoisomerism

OBJECTIVE: To establish a size exclusion chromatography(HPSCE) method for the determination of the polymers in meropenem for injection. METHODS: The analysis was carried out using a TSKgel G2000SWXL column(7.8 mm×30 cm, 5 μm). The mobile phase was pH 7.0 phosphate buffer solution(0.005 mol•L-1 disodium hydrogen orthophosphate-0.005 mol•L-1 sodium dihydrogen phosphate, 61:39, V/V). The flow rate was 0.8 mL•min-1. The detection wavelength was 254 nm. The column temperature was 25 ℃. RESULTS: The polymers were completely separated from meropenem. The calibration curve of meropenem was linear within the range of 2.54-50.83 μg•mL-1(r=0.999 6, n=6). The specificity of the HPSEC was checked by switching the effluent of each peak on the GFC to a C18 column and analyzing the effluent by LC-MS. CONCLUSION: This method is simple, accurate, sensitive, reliable and applicable for the determination of the polymers of meropenem, and can be applied to the drug quality evaluation.

OBJECTIVE: To study the plasma protein binding rate of decapeptide in several species. METHODS: Ultrafiltration was employed to separate the free and bound decapeptide, acetonitrile was used to precipitate protein, then the plasma concentration of decapeptide by HPLC-MS/MS. RESULTS: The plasma protein binding rates of decapeptide at low, middle and high concentrations (50.0, 100.0, and 800.0 ng·mL-1) were (95.9±1.1)%, (97.40±0.7)% and (94.9±0.6)% in SD rats,(96.8±0.8)%, (97.8±0.2)% and (96.9±0.5)% in Beagle dogs, and (97.3±1.0)%, (98.6±0.2 )% and (96.2±0.9)% in humans, respectively. The average plasma protein binding rate was 96.2% after subcutaneous administration at 200 μg·kg-1 in SD rats. And the average plasma protein binding rate was 97.1% after intramuscular injection at 320 μg·kg-1 in Beagle dogs. CONCLUSION: Decapeptide has potent binding ability to plasma protein in several species. The plasma protein binding rate of decapeptide is independent of its plasma concentrations.

The aim of this study was to find platelet specific autoantibodies against glycoproteins in myelodysplastic syndrome (MDS) and to explore its role in pathogenesis of MDS. The plasma autoantibodies against GP IIb/IIIa and GP Ib/IX were measured by using a modified monoclonal antibody specific immobolization platelet antigens assay (MAIPA). Absorbance greater than mean value plus tripled standard deviation recorded from the normal controls were regarded as positive. The results indicated that the total positive rate in patients with MDS was 16.67% (5/30), the total positive rate in patients with ITP was 46.67% (14/30), the difference between MDS group and ITP group was significant (P < 0.05). It is concluded that partial patients with MDS have plasma specific autoantibodies against platelet GP II b/III a and GP Ib/IX, indicating correlation of thrombocytopenia of patients with immune factors and the autoantibody-mediated platelet destruction may be involved in the pathogenesis of MDS. It provides a new basis for immunosuppression therapy for MDS.
Adolescent ; Adult ; Aged ; Antibodies ; immunology ; Antigens, Human Platelet ; immunology ; Autoantibodies ; biosynthesis ; immunology ; Female ; Humans ; Male ; Middle Aged ; Myelodysplastic Syndromes ; complications ; immunology ; Platelet Glycoprotein GPIIb-IIIa Complex ; immunology ; Platelet Glycoprotein GPIb-IX Complex ; immunology ; Thrombocytopenia ; etiology ; immunology

This paper developed a sensitive and specific liquid chromatography-electrospray ionization mass spectrometry (HPLC-MS/MS) method for the determination of decapeptide LXT-101 in Beagle dog plasma. Plasma samples spiked with internal standard (IS) were treated with acetonitrile to precipitate the protein. Selected reaction monitoring (SRM) using the precursor --> product ion combinations of m/z 472.1-->587.9 and m/z 502.8-->633.8 were used to quantify LXT-101 and IS, respectively. The linear calibration curves were obtained in the concentration range of 0.5 - 500.0 ng x mL(-1). The limit of quantification (LOQ) was 0.5 ng x mL(-1). The inter-day and intra-day precision (RSD) across three validation run over the entire concentration range was below 10.9%, and the accuracy (RE) was within +/- 1.8%. The main pharmacokinetic parameters of LXT-101 after muscle injection of 20 microg x kg(-1) were as follows, AUC(0-t): (176.8 +/- 116.7) microg x h x L(-1), MRT(0-t): (2.52 +/- 0.53) h, T(1/2): (1.4 +/- 0.3) h; CL: (0.16 +/- 0.09) L x h(-1) x kg(-1), and Vd: (0.30 +/- 0.16) L x kg(-1), respectively. The method is proved to be specific, sensitive and suitable for the investigation of LXT-101 pharmacokinetics in Beagle dog.
Animals ; Antineoplastic Agents ; administration & dosage ; blood ; pharmacokinetics ; Area Under Curve ; Chromatography, High Pressure Liquid ; Dogs ; Gonadotropin-Releasing Hormone ; antagonists & inhibitors ; Injections, Intramuscular ; Male ; Oligopeptides ; administration & dosage ; blood ; pharmacokinetics ; Spectrometry, Mass, Electrospray Ionization

AIMTo study the effect of curcumin on the expression of prostate specific antigen (PSA).

METHODSAXSYM system-chemical luciferase method was used to examine the content of PSA in prostate cancer cell lines, LNCap after treated with different doses of curcumin. pGL3-PSA luciferase expression vector, containing 640 bp DNA of PSA gene 5' promoter region was constructed and transfected into LNCap cell with lipofectin. Through detecting the activity of luciferase, the effect of curcumin on the promoter of PSA was studied. Western blotting was used to detect expression of androgen receptor (AR) in LNCap cell with different concentrations of curcumin.

RESULTSThe expression of PSA was inhibited and activity of luciferase was reduced by curcumin. There was also significant difference in AR expression as shown by Western blotting experiment after treatment of different doses of curcumin.

CONCLUSIONThrough inhibiting AR expression, curcumin reduced the function of PSA promoter and inhibited PSA protein expression.

Antineoplastic Agents ; pharmacology ; Cell Line, Tumor ; Curcumin ; pharmacology ; Humans ; Luciferases ; metabolism ; Male ; Promoter Regions, Genetic ; drug effects ; Prostate-Specific Antigen ; genetics ; metabolism ; Prostatic Neoplasms ; immunology ; metabolism ; pathology ; Receptors, Androgen ; metabolism

OBJECTIVETo investigate the quantitative and qualitative changes of TCRVα24(+)Vβ11(+) natural killer T (NKT) cells from bone marrow (BM) of aplastic anemia (AA) after in vitro stimulation of α-galactosylceramide (α-Galcer).

METHODSNKT cells in the bone marrow mononuclear cells (BMMNCs) from either AA patients or healthy controls were enumerated with flow cytometry. BMMNCs were cultured in RPMI1640 medium supplemented with either α-Galcer and rhIL-2 or α-Galcer, rhIL-2 and rhG-CSF. The proliferative capacity of NKT cells was determined by NKT cell numbers before and after in vitro culture. Expression of intracellular IFNγ and IL-4 in activated NKT cells was analyzed with flow cytometry.

RESULTSIn AA group, the percentage of NKT cells in BMMNCs was (0.19 ± 0.09)%. Addition of rhG-CSF into the α-Galcer/rhIL-2 culture medium resulted in significantly reduced expansion of NKT cells (67.45 ± 29.42-fold vs 79.91 ± 40.56 fold, P < 0.05). Meanwhile, addition of rhG-CSF reduced IFNγ positive NKT cells \[(37.45 ± 7.89)% vs (62.31 ± 14.67)%, P < 0.01\] and increased IL-4 positive NKT cells \[(55.11 ± 12.13)% vs (27.03 ± 9.88)%, P < 0.01\]. In healthy control group, the percentage of NKT cells in BMMNCs was (0.25 ± 0.12)%. Addition of rhG-CSF into the α-Galcer/rhIL-2 culture medium also significantly reduced expansion of NKT cells (97.91 ± 53.22-fold vs 119.58 ± 60.49-fold, P < 0.05), reduced IFNγ positive NKT cells \[(28.65 ± 10.63)% vs (50.87 ± 12.66)%, P < 0.01\], and increased IL-4 positive NKT cells \[(66.53 ± 14.96)% vs (31.11 ± 10.07)%, P < 0.01\].

CONCLUSIONCompared to those from healthy controls, BMMNCs from AA patiants have a reduced fraction of NKT cells, which possesses a decreased potential to expand in vitro in response to α-Galcer stimulation, and produce more IFNγ(+) NKT1 cells. rhG-CSF, in combination with α-Galcer, confers polarization of NKT cells towards IL-4(+) NKT2 subpopulation.

Anemia, Aplastic ; metabolism ; Bone Marrow ; metabolism ; Humans ; Interleukin-4 ; metabolism ; Killer Cells, Natural ; cytology ; Natural Killer T-Cells

OBJECTIVEAnalysis the viral pathogenic spectrum for patients with fever and respiratory tract infection syndrome in Shaanxi province during 2010 and investigate the molecular epidemiology characteristics of respiratory syncytial virus.

METHODSA total of 208 patients' pharyngeal swabs were collected based on surveillance definition from January 2010 to January 2011 and screened for sixteen human respiratory virus types/subtypes by Qiaxcel-based multiplex reverse transcription-PCR assay, including HRV,HCoV, Flu, HPIV, ADV, HRSV, HMPV and HBoV and investigate molecular epidemiology of HRSV by sequencing and phylogenetic analysis of the C-terminal second hypervariable region of the G gene.

RESULTS109 out of 208 specimens (53%) were positive for one or more viruses. HRSV(42. 2%) was the dominant pathogen detected, followed by Flu(24. 5%), PIV(20%), HRV(13.6%) and ADV( 10.9%),there were also 8 strains of HCoV, 5 strains of HMPV and 3 strains of HBoV detected. The results showed that 22 specimens were positive for two or more viruses, PIV (14/22) was the most frequently detected viral agent among co-infection specimens, and the highest incidence of mixed infection is aged 15-39 years group (P < 0.05). The overall viral detection rate was no related to age. In addition to Flu, HMPV and PIV, other viruses (HRV, HBoV, HCoV, ADV, RSV) mainly infected 0 to 4 years old children. Among 46 HRSV positive specimens, 42 HRSV-A strains clustered into NA1 genotype and two HRSV-B strains clustered into two genotypes, BA9 and GB2.

CONCLUSIONHRSV is the dominate pathogen collected from patients with fever and respiratory tract infection syndrome in Shaanxi and HRSV A is the predominant subtype. For most viruses, infection was most prevalent among children aged <4 years. PIV was the most common pathogen in co-infection.

Adolescent ; Adult ; China ; epidemiology ; Coinfection ; virology ; Female ; Fever ; epidemiology ; virology ; Genotype ; Humans ; Male ; Phylogeny ; Respiratory Syncytial Virus Infections ; epidemiology ; virology ; Respiratory Syncytial Virus, Human ; isolation & purification ; Respiratory Tract Infections ; epidemiology ; virology ; Young Adult