LC-MS/MS测定人血浆中对乙酰氨基酚及其人体药动学和生物等效性研究

目的 建立一种快速、灵敏的高效液相色谱-串联质谱（HPLC-MS/MS）方法以测定人血浆中对乙酰氨基酚浓度,并应用于两种对乙酰氨基酚制剂的人体药代动力学和生物等效性研究。方法 以替硝唑为内标,200μL血浆样品经5倍于其体积的乙酸乙酯液液萃取,再经Waters XBridge? C18柱等度洗脱分离后导入串联质谱,以正离子多反应监测模式进行定量分析,对乙酰氨基酚和内标的选择性反应离子对分别是m/z 152→110和248→121。方法经验证后应用于19名健康受试者单剂量空腹口服两种对乙酰氨基酚制剂500mg后药代动力学和生物等效性的研究。结果 血浆中对乙酰氨基酚在0.1～8.0 μg·mL-1范围内线性良好（r2 > 0.99）,最低检测限为 0.1 μg·mL-1,提取回收率为91.0%～98.7%,日内和日间准确度分别为98.8%～111.3% （精密度：CV ? 9.03%）和94.9%～102.6% （精密度：CV ? 10.68%）。生物等效性试验中,受试制剂与参比制剂的主要药代动力学参数Cmax、AUC0-t和AUC0-∞ 几何均值比的90%置信区间分别为83.50%～105.79%,94.25%～101.54%和93.24%～101.02%,均落在生物等效可接受标准80.00%～125.00%范围内。结论 所建立测定人血浆中对乙酰氨基酚浓度的HPLC-MS/MS法具有快速灵敏、回收率高、选择性好的特点,适用于对乙酰氨基酚片人体药代动力学和生物等效性研究。受试制剂与参比制剂在人体内吸收速度和程度相似,两种制剂生物等效。     

A combination of radiotherapy, nitric oxide and a hyperoxygenation sensitizing protocol for brain malignant tumor treatment

Brain malignant tumor such as glioblastoma is a challenging medical and surgical problem. In spite of surgery, radiotherapy and chemotherapy, the prognosis is still very poor. The limitations of currently available treatment modalities to cure or significantly prolong and improve the quality of life should stimulate rigorous research and studies to combat brain malignant tumors. While precision radiotherapy to reduce tumor size and ameliorate symptoms is still the standard of care, tumor sensitivity to radiation is compromised by low oxygen tensions and a necrotic tumor center. We propose to take advantage of the fact that elevated oxygen increases sensitivity of tumor cells to radiation. A specific application of hyperbaric oxygen (HBO(2)), using nitric oxide (NO) donors and inducers (such as L-arginine, dinitrite or tocopheryl succinate) and ascorbic acid to dilate blood vessels, should permit oxygen tensions in the range of 1000 mmHg to diffuse into the cells and thus increase sensitivity to radiation. This should permit doses that are low enough to cause the death of tumors cells yet minimize injury to brain tissue near the tumor and induced neurological sequelae.     

Adhesion of platelets to surface-bound fibrinogen under flow

After platelet activation, fibrinogen mediates platelet-platelet interactions leading to platelet aggregation. In addition, fibrinogen can also function as a cell adhesion molecule, providing a substratum for adhesion of platelets and endothelial cells. In this report, we studied the adhesion of platelets to surface-immobilized fibrinogen under flow in different shear rates. Heparinized whole blood containing mepacrine-labeled platelets was perfused for two minutes at various wall shear rates from 250 to 2,000 s-1 in a parallel plate flow chamber. The number of adherent fluorescent platelets was quantitated every 15 seconds with an epifluorescent videomicroscope and digital image processing system. When compared with platelet adhesion and aggregation seen on glass surfaces coated with type I bovine collagen, a significant increase in platelet adhesion was observed on immobilized fibrinogen up to wall shear rates of 800 s-1. The adherent platelets formed a single layer on fibrinogen-coated surfaces. Under identical conditions, no significant adhesion was observed on fibronectin- or vitronectin-coated surfaces. Although platelet adhesion to collagen was substantially inhibited by the platelet inhibitors prostaglandin E1 and theophylline, these inhibitors had no effect on platelet adhesion to fibrinogen. Platelets adhered to recombinant homodimeric wild-type (gamma 400-411) fibrinogen, but not to the recombinant homodimeric gamma' variant of fibrinogen. Platelet adhesion to recombinant fibrinogen with RGD to RGE mutations at positions alpha 95-97 and alpha 572-574 was similar to that with plasma-derived fibrinogen. These results show that platelets adhere to fibrinogen-coated surfaces under moderate wall shear rates, that the interaction is mediated by the fibrinogen 400-411 sequence at the carboxy-terminus of the gamma chain, and that the interaction is independent of platelet activation and the RGD sequences in the alpha chain.     

The development of cellular immunity to Epstein-Barr virus after allogeneic bone marrow transplantation

Epstein-Barr virus-induced lymphoproliferative disease (EBV-LPD) is a potentially lethal complication during the first 6 months after allogeneic bone marrow transplantation (BMT). To determine whether deficiencies of EBV-specific cellular immunity contribute to EBV-LPD susceptibility and distinguish patients at risk, we performed limiting dilution analysis to quantify anti-EBV cytotoxic T-lymphocyte precursor (CTLp) frequencies in 26 recipients of unmodified or T-cell-depleted (TCD) grafts from EBV-seropositive donors. At 3 months post-BMT (n = 26), only five patients had EBV CTLp frequencies in the range of seropositive normal controls, irrespective of the type of transplant administered. By 6 months post-BMT, 9 of 13 patients tested had EBV CTLp frequencies within the normal range. The time period in which these patients had deficient cellular immunity to EBV corresponds to the period in which we have observed EBV-LPD in most prior patients. One patient with a low EBV CTLp frequency at 4 months post-BMT developed an EBV-LPD. Within 2 weeks of receiving an infusion of donor peripheral blood mononuclear cells (PBMC) providing less than 1,200 EBV- specific cytotoxic T-cell precursors, populations of EBV-specific CTL in the circulation were restored to levels detected in normal seropositive adults. Concurrently, the patient achieved a regression of the EBV-LPD, which has been sustained without further therapy. These studies indicate that recipients of both unmodified and TCD marrow grafts have profound deficiencies of EBV-specific T cell-mediated immunity early posttransplant, and that the period of risk for EBV-LPD closely corresponds to this interval of severe deficiency. Treatment of one patient with EBV-LPD with marrow donor-derived PBMC induced a rapid expansion of EBV-specific cytotoxic T-cell populations that occurred contemporaneously with the clinical regression of disease.     

对产前母体苯巴比妥给药以预防针对新生儿和胎儿溶血病的婴儿换血

胎儿和新生儿溶血病(HDFN)可导致严重的围产期疾病,包括高胆红素血症及其引起的核黄疸。血红素分子解离为胆红素并产生免疫介导的溶血。它与白蛋白结合后转运到肝细胞,再由尿苷二磷酸葡萄糖醛酸转移酶(UGT)酶家族作用而葡萄糖醛酸化。对于人类,这种代谢途径中主要的酶是胆红素-     

Risk factors for development of dehydration in young children with acute watery diarrhoea: a case-control study

In a case-control study to understand the risk factors for development of life-threatening dehydration, a total of 379 children comprising 243 cases (moderate or severe dehydration) and 136 controls (non or mild dehydration) up to 2 years of age suffering from acute watery diarrhoea were studied. By univariate analysis, the presence of vibrios in stool, withdrawal of breast feeding during diarrhoea, not giving fluids, including oral rehydration solution (ORS), during diarrhoea, frequent purging (> 8/ day), vomiting (> 2/day) and undernutrition were identified as risk factors. However, by multivariate analysis after controlling for confounders, withdrawal of breast feeding during diarrhoea (odds ratio (OR) = 6.8, p < 0.00001) and not giving ORS during diarrhoea (OR = 2.1, p < 0.006) were identified as significant risk factors. The confounding variables which also contributed significantly to increasing the risk were age (≤ 12 months; OR = 2.7, p = 0.001), frequent purging (> 8/day; OR = 4.1, p < 0.00001), vomiting (> 2/day; OR = 2.4, p = 0.001) and severe undernutrition (%median <60 weight-for-age of Indian Academy of Paediatrics classification; OR = 3.1, p = 0.001). We feel that these findings will be useful for Global and National Diarrhoeal Diseases Control Programmes for formulating intervention strategies for preventing death due to diarrhoeal dehydration.     

Urinary nitrite excretion and urinary variables in patients with primary nocturnal frequency of micturition: effects of indomethacin suppositories

Urinary nitrite excretion was measured in patients with primary nocturnal frequency of micturition (PNFM) and in normal individuals. Effects of indomethacin suppository on urine volume and other urinary variables were evaluated. The study comprised seven patients with PNFM and seven healthy control (age range 30–45 years). Nitrite was assayed in spot morning urine samples; urine volume, urine osmolality and electrolytes, serum osmolality and electrolytes and functional bladder capacity (FBC) were assayed. Both groups were then given 100 mg of indomethacin suppository daily for a maximum of 10 days and urinary variables were re-evaluated during day 10. Results showed that urinary nitrite excretion of patients with PNFM was greater than that of the normal subjects (230±62 umol/l vs. 42±30 umol/l, P<0.05). The mean (SD) 24 h urine volume and osmolality, the night urine volume and osmolality, serum osmolality, FBC, creatinine clearance, fractional excretion of sodium (FENa), fractional excretion of potassium (FEK), and urinary excretion of glucose and potassium were lower in patients with PNFM as compared with normal individuals, although not statistically significantly so, except for FBC that was significantly lower in the patients. Urinary excretion of sodium, calcium, chloride, phosphorus, magnesium, day-night urinary volume ratio, spot morning osmolality, nocturnal index, and nocturnal polyuria index were higher in patients with PNFM. Indomethacin decreased the 24 h urinary volume by 21%, creatinine clearance by 12%, osmolar clearance by 14% and urinary protein excretion by 38% in the patients. These variables decreased by 26, 45, 17 and 12% respectively in the healthy subjects, whereas 24 h urinary protein excretion increased mildly by 9%. Indomethacin increased day-night urinary volume ratio by 73% in the healthy subjects. It might be concluded that urinary nitrite excretion, urinary excretion of sodium, chloride, phosphorus, calcium, and magnesium increased and FBC decreased in patients with PNFM; Indomethacin decreased urinary volume, FENa, FEK, osmolar clearance, and free water clearance in the healthy subjects and the patients. These might explain the mechanism of action of indomethacin to reduce frequency of voiding. The possible interaction of prostaglandin and NO in the pathogenesis of PNFM is discussed.     

Investigating the antimicrobial activity of natural honey and its effects on the pathogenic bacterial infections of surgical wounds and conjunctiva

Antimicrobial activities of 10-100% (wt/vol) concentrations of new honey, stored honey, heated honey, ultraviolet-exposed honey, and heated stored honey were tested against common human pathogens, including Escherichia coli, Entrobacter cloacae, Pseudomonas aeruginosa, Shigella dysenteriae, Klebsiella sp., Haemophilus influenzae, Proteus sp., Staphylococcus aureus, Streptococcus hemolyticus group B, and Candida albicans. Antimicrobial activity of honey was tested in acidic, neutral, or alkaline media. These were compared with similar concentrations of glucose in nutrient broth. Surgical wounds were made on the dorsum of mice and infected with S. aureus or Klebsiella sp. The wounds were treated with local application of honey four times a day or appropriate antibiotics and compared with control values. Bacterial conjunctivitis due to E. coli, Proteus sp., S. aureus, Klebsiella sp., and P. aeruginosa was induced in rats. Conjunctival application of honey four times a day or appropriate antibiotics was used for treatment and compared with control values. Growth of all the isolates was completely inhibited by 30-100% honey concentrations. The most sensitive microbes were E. coli, P. aeruginosa, and H. influenzae. Glucose showed less antimicrobial activity than honey, and many microbes showed positive culture even in 100% glucose. Heating to 80 degrees C for 1 hour decreased antimicrobial activity of both new and stored honey. Storage of honey for 5 years decreased its antimicrobial activity, while ultraviolet light exposure increased its activity against some of the microorganisms. Antimicrobial activity of honey was stronger in acidic media than in neutral or alkaline media. Single doses of honey used to prepare the 60% concentration in nutrient broth were bacteriocidal for P. aeruginosa and bacteriostatic for S. aureus and Klebsiella sp. during certain periods. Local application of raw honey on infected wounds reduced redness, swelling, time for complete resolution of lesion, and time for eradication of bacterial infection due to S. aureus or Klebsiella sp. Its potency was comparable to that of local antibiotics. Honey application into infective conjunctivitis reduced redness, swelling, pus discharge, and time for eradication of bacterial infections due to all the isolates tested.