Neurochemical development of the brainstem catecholaminergic cell groups in rat

The postnatal development of tyrosine hydroxylase activity has been studied in the brainstem catecholaminergic cell groups (A1C1, A2C2, A5, A6, A7), involved in cardiorespiratory control. In rat, at birth and at postnatal days P3, P7, P14, P21 ant P68, we used a microdissection technique followed by in vivo measurement of the tyrosine hydroxylase (TH) activity, the rate-limiting enzyme in catecholamine synthesis. There is two successive marked increases in TH activity: at P3 in every catecholaminergic cell groups (A1C1, +225%; A2C2, +300%; A5, +190%; A6, +205% compared to birth) and during the third postnatal week with a peak of TH activity at P14 (A6, +90% above the P7 level) or at P21 (A1C1, +715%; caudal A2C2, +585%; rostral A2C2, +15%; A5, +445%; A7, +180% compared to P7). The data suggest the existence of two temporal windows during the neurochemical development of the catecholaminergic cell groups, which correspond to two metabolic transitions. The first one could be related to the intra-, extrauterine transition and the second one, to a deep energetic phase of maturation in the rat brain, closely related to the maturation of cardiorespiratory processes.     

导痰祛瘀口服液的工艺研究

目的:研究导痰祛瘀口服液的提取工艺。方法:采用 Ｌ９(３４)正交试验设计及Ｕ４７均匀试验设计,以大黄中的大黄素含量及石菖蒲中挥发油含量分别为指标进行试验。结果:较好渗漉条件为 ５５%乙醇渗漉１０倍量,５%ＮａＯＨ作润湿液,流速为３ｍｌ/ｍｉｎ;较好蒸馏条件为药材粉碎成粗粉,加水１６倍,蒸馏５ｈ。结论:研究所得工艺较合理。     

Comparison of subretinal fluid levels of two 0.3% ciprofloxacin-containing eye drops

Purpose : Two ophthalmic solutions of 0.3% ciprofloxacin eye drops are available in Turkey: Ciloxan and Siprogut. A previous study by the same authors was the first to report vitreous penetration of ciprofloxacin‐containing eye drops. The aim of the present study was to compare the levels of drug found in the subretinal fluid by the two products following local administration. Methods : Forty‐three patients undergoing conventional retinal detachment surgery received either Ciloxan (22 patients) or Siprogut (21 patients). Beginning 6 h before surgery, two drops of solution were instilled onto the operative eye every 30 min for the first 3 h and then hourly for the next 3 h. Subretinal fluid samples were collected 30 min after administration of the last dose and were assayed for ciprofloxacin levels using a method involving high‐ performance liquid chromatography with fluorometric detection. Results : The minimum and maximum subretinal fluid concentrations measured were 0.11 μg/mL and 0.65 μg/mL, respectively, with Ciloxan, and 0.08 μg/mL and 0.62 μg/mL, respectively, with Siprogut. There was no statistical difference between the subretinal fluid ciprofloxacin levels of the two products. The subretinal fluid drug levels attained by both products were below the minimum inhibitory concentrations of common ocular pathogens. Conclusions : Ciloxan and Siprogut can penetrate subretinal fluid. The ocular bioavailability of ciprofloxacin after local administration is equivalent for both pharmaceutical products.     

Simultaneous HPLC determination of nitrendipine and impurities of the process of synthesis

A method has been developed for separation of nitrendipine and its impurities of reaction partners and side reaction products by high-performance liquid chromatographic method on a RP-18 column and detection at 238 nm. The mobile phase composition that provided an acceptable nitrendipine resolution, in large excess and possible impurities, in a short elution time, is methanol:water (70:30) and pH 3. Linearity (r≥0.999), reproducibility (RSD=0.8–1.4%), determination limit (0.5–2%) and recovery (99.8–102.3) were validated and found to be satisfactory. This method enables monitoring of the process of synthesis, as well as the choice of the synthetic design.     

Application of conventional UV, photodiode array (PDA) and fluorescence (FL) detection to analysis of phenolic acids in plant material and pharmaceutical preparations

Free phenolic acids (PhAs) contained in methanolic extracts of Eleutherococcus senticosus roots and pharmaceutical preparations, deriving from this plant, were isolated by solid-phase extraction (SPE) and identified by reversed-phase high-performance liquid chromatography (RP-HPLC). To obtain precise, accurate and validated results of qualitative and quantitative analysis, ultraviolet (at λ=254 nm), photodiode array (at λ=254 and 280 nm) and fluorescence (at λ_Ex=230 or 265 nm and λ_Em=350 nm) detection was used. Additionally, the HPLC separation of PhAs on two different octadecyl sorbents: Hypersil™ (Shandon, UK) and Symmetry™ (Waters, USA) was performed. Eight PhAs: chlorogenic, protocatechuic, p-hydroxybenzoic, caffeic, vanillic, syringic, p-coumaric and ferulic, in different quantitative proportions, were identified, both in the roots of E. senticosus and pharmaceutical formulations examined.     

Identification and quantitation of sodium-thyroxine and its degradation products by LC using electrochemical and MS detection

High performance liquid chromatography (HPLC) was used in combination with an amperometric and mass spectrometric detection to elucidate and quantitate the degradation products and contaminants of the photo-sensitive Na-thyroxine. Using HPLC with amperometric detection, seven decomposition compounds were separated. These products, which occur mostly as contaminants, were then identified by a developed liquid chromatography-mass spectrometry technique. The same HPLC method was also employed to analyze Na-thyroxine and its degradation products in three commercially available brands of Na-thyroxine tablets.     

Assay of gamma-glutamylcysteine synthetase activity in Plasmodium berghei by liquid chromatography with electrochemical detection

This work describes a high-performance liquid chromatography (HPLC) method to determine γ-glutamylcysteine (γ-GC), the intermediate product of glutathione biosynthesis. Separation relies on isocratic reversed-phase chromatography using a Symmetry C18 HPLC column, particle size 5 μm, 4.6×250 mm i.d. The mobile phase is methanol–dibasic sodium phosphate (pH 6.6; 2.8 mM) (10:90, v/v) at the flow-rate of 0.5 ml/min and detection is operated electrochemically (+200 and +550 mV) with a pre-column derivatisation reaction using ortho-phthalaldehyde (OPA) as reagent. Under these conditions the calibration range of γ-GC was 0.3–10 μg/ml; the limit of quantification was 0.3 μg/ml; accuracy, expressed as %Bias, was <10 and precision (%CV) was <6. The proposed HPLC assay was used to quantitate the γ-glutamylcysteine produced by the γ-glutamylcysteine synthetase of the rodent malaria parasite Plasmodium berghei in an in vitro enzymatic assay.     

Comparison between computerized slow-stage and static liquid nitrogen vapour freezing methods with respect to the deleterious effect on chromatin and morphology of spermatozoa from fertile and subfertile men

The purpose of this study was to determine the negative effects (cryodamage) on human spermatozoa after freeze-thawing and to determine whether freeze-thawing of spermatozoa with a programmed slow freezer is better than freezing with liquid nitrogen vapour (rapid freezing) with regard to alterations in sperm chromatin and morphology in semen from fertile (donor) and subfertile, IVF/ICSI, patients. Ninety-five semen samples were obtained either from patients attending our IVF unit for treatment (n=34) or from donors (n=25) with proven fertility and normal sperm quality according to WHO guidelines. Each semen sample was divided into two parts after liquefaction and addition of the cryoprotectant. The first part was frozen using a programmed biological freezer and the second part was frozen by means of liquid nitrogen vapour. Smears were made before the freezing and after the thawing procedure to assess morphology (strict criteria) and chromatin condensation (Acridine Orange test). The mean percentage of chromatin condensed spermatozoa in the samples from donors (control group) was 92.4 +/- 8.4% before freezing and this decreased significantly (p < 0.0001) to 88.7 +/- 11.2% after freeze-thawing with the computerized slow-stage freezer and to 87.2 +/- 12.3% after using static liquid nitrogen vapour (p < 0.001). The corresponding values for semen obtained from patients was 78.9 +/- 10.3% before freezing which decreased to 70.7 +/- 10.8 and 68.5 +/- 14.8%, respectively (p < 0.001). On the other hand, the mean percentage of normal sperm morphology in the control group decreased from 26.3 +/- 7.5% before freezing to 22.1 +/- 6.4% (p < 0.0001) after thawing with the computerized slow-stage freezer and to 22.2 +/- 6.6% (p < 0.0001) after the use of static liquid nitrogen vapour. In the patient group, the mean percentage of normal morphology decreased from 11.7 +/- 6.1% after freezing with the biological freezer to 9.3 +/- 5.6% and to 8.0 +/- 4.9% after freezing with static liquid nitrogen vapour. This study demonstrates that chromatin packaging and morphology of human spermatozoa decrease significantly after the freeze-thawing procedure, not only after the use of static liquid nitrogen vapour but also after the use of a computerized slow-stage freezer. However, the chromatin of semen samples with normal semen parameters (donor sperm) withstand the freeze-thaw injury better than those with low quality semen samples. Therefore, the computerized slow stage freezer could be recommended for freezing of human spermatozoa, especially for subnormal semen samples, for example, ICSI and ICSI/TESE candidates and from patients with testicular tumours or Hodgkin's disease, in order to avoid further damage to the sperm chromatin structure.     

Immune hemolytic anemia caused by sensitivity to a metabolite of etodolac, a nonsteroidal anti-inflammatory drug

BACKGROUND: Immune hemolytic anemia can be caused by sensitivity to many different drugs. In some instances, the sensitizing compound can be identified by in vitro testing, but results are often negative. One reason for this is that a drug metabolite formed in vivo can be the sensitizing agent, but the responsible metabolites have rarely been identified at a chemical level. This report describes a patient who developed severe, Coombs-positive hemolytic anemia on two occasions after taking the nonsteroidal anti-inflammatory drug etodolac. Studies were performed to characterize etodolac metabolites to which this patient was sensitive. CASE REPORT: Serum was tested for antibody in the presence and absence of drug using conventional methods and urine from individuals taking etodolac as a source of drug metabolites. Urinary metabolites of etodolac were identified by high-pressure liquid chromatography analysis. Glucuronide conjugates of etodolac and the 6-OH metabolite of etodolac were synthesized in a rat liver microsomal system to obtain reference standards. RESULTS: The patient's serum gave only trace (+/-) reactions with normal RBCs in the presence of etodolac but reacted strongly (4+) in the presence of urine from an individual taking this drug. The active urinary metabolites were identified as etodolac glucuronide and 6-OH etodolac glucuronide. CONCLUSION: This patient appears to have experienced acute, severe immune hemolytic anemia on two occasions because of sensitivity to the glucuronides of etodolac and 6-OH etodolac. In patients suspected of having drug-induced immune hemolytic anemia, RBC-reactive antibodies can sometimes be detected by using urine from an individual taking the implicated medication as the source of drug metabolites in in vitro reactions. For patients who present with acute immune hemolysis, a careful history of drug exposure should be taken, and, where indicated, confirmatory testing should be performed to identify the sensitizing drug and prevent inadvertent reinduction of hemolysis at a later time.     

高效液相色谱法检测食品中氨基酸的方法研究

目的应用高效液相色谱法测定食品中氨基酸。方法使用带自动衍生功能的自动进样器进行在线衍生，梯度分离，利用二极管阵列一荧光检测器串联分析。器联用技术能有效提高定性的可靠性和定量的准确性。确，结果满意。结果在线衍生技术克服了离线手工衍生的偏差，双检测结论研究的方法测定重复性好，灵敏度高，定性定量准