Prenylated proteins and lymphocyte proliferation: inhibition by d-limonene related monoterpenes

The aim of the present study was to explore the role of post-translational isoprenoid modification of cellular proteins in the proliferation of human lymphocytes. We here report that treatment of phytohemagglutinin-stimulated peripheral blood mononuclear cells with monoterpenes including d-limonene, perillic acid and perillyl alcohol (0.5-5 mM) which selectively inhibit the isoprenylation of 21-26-kDa proteins resulted in a dose-dependent inhibition of DNA synthesis. Cell cycle analysis revealed that perillic acid arrested cells in G1 and prevented cells from entering S phase in a manner similar to that induced by the specific 3-hydroxy-3-methylglutaryl-CoA reductase inhibitor, compactin. However, unlike compactin, the perillic acid-induced effects on lymphocyte proliferation were not prevented by addition of mevalonate. We also examined the incorporation of [3H]mevalonate into proteins in resting and phytohemagglutinin-stimulated lymphocytes during the first 30 h of culture. While in unstimulated lymphocytes radioactivity was predominantly incorporated into a cluster of 21-26-kDa proteins, mitogenic stimulation was associated with a striking increase in [3H]mevalonate incorporation into a protein (approximately 68 kDa) with migration characteristics similar to that of nuclear lamin B. Treatment of phytohemagglutinin-stimulated lymphocytes with 5 mM d-limonene, 2.5 mM perillic acid or 1.25 mM perillyl alcohol strongly suppressed [3H]mevalonate-labeling of proteins to a degree that correlated with the level of DNA synthesis inhibition. These findings suggest that those mevalonate-derived products required for lymphocyte proliferation may include one or more isoprenylated proteins and that the isoprenylation of these proteins is required for cell cycle progression.     

Analysis of suckling-induced changes in adenohypophyseal prolactin concentration in the lactating rat by three assay methods

Suckling-induced changes in adenohypophyseal prolactin (PRL) concentrations in lactating rats were measured in two experiments by three assays: a disc electrophoretic assay (DEA), a bioassay (BA) and a radioimmunoassay (RIA). Substantial discrepancies among the three assays were found in the absolute amounts of pituitary PRL measured and in the changes in adenohypophyseal PRL concentration that were recorded between suckling intervals. The results indicate that in dynamic states of secretory activity the correspondence among BA, RIA and DEA estimates of pituitary PRL concentration is poor.     

高河菜脂溶性化学成分分析

[目的]分析高河菜脂溶性化学成分,为高河菜的开发利用提供试验依据.[方法]采用气相色谱-质谱联用(GC-MS)技术分离鉴定脂溶性成分,用峰面积归一化法计算各成分相对百分含量.[结果]共鉴定出38种组分,占总成分含量的80.80%,其中植醇(26.49%)、乙基胆甾醇(10.65%)、2-甘油-亚麻酸脂(7.84%)和亚麻酸甲酯(7.16%)含量较高.[结论]高河菜脂溶性成分主要为萜类、脂肪酸酯、甾醇和烷烃类化合物,为高河菜的科学利用提供了试验依据.     

Pharmacokinetics of pentazocine and its occupancy of opioid receptors in rat brain

In order to assess quantitatively the pharmacodynamic process of pentazocine (PTZ), time courses of its plasma concentration and of the occupation of specific opioid receptors in the brain were investigated after intravenous (i.v.) administration of PTZ to rats. The plasma concentration of PTZ was determined by HPLC and the pharmacokinetic parameters were analyzed using nonlinear least-squares analysis. Measurement of ex vivo receptor occupation was made by comparing the specific [3H]naloxone (opioid receptor antagonist) binding in vitro to the crude P2-synaptosomal fractions between vehicle-treated rats (control) and PTZ-treated rats. Following the i.v. administration of PTZ, the occupancy of specific opioid receptors decreased rapidly until 10 min, depending on the two pharmacological doses (2.5 and 10 mg/kg). The results strongly suggest the fast binding kinetics of PTZ in terms of its association with and dissociation from specific opioid receptor sites in the brain in addition to its fast rate of disappearance from the brain compartment. Furthermore, we demonstrated that the time profile of receptor occupancy correlated well (r = 0.8650) with that of the unbound concentration in plasma until 120 min after the i.v. administration of PTZ to rats.     

Development and utilization of the rat lymphocyte hprt mutation assay

Much of the recent progress in the field of genetic toxicology has come from an increased understanding of the molecular and cellular biology of the mammalian organism. Most prominent has been the ability to detect and quantify somatic mutation and relate the nature of the mutation to the specific type of chemical damage. Building upon the foundation of the human lymphocyte hypoxanthine guanine phosphoribosyl transferase (hprt) system, and later, the mouse hprt system, methods for the detection and quantification of hprt mutations in rat lymphocytes were developed. These methods are described in this report as is the ongoing validation of the assay. Additionally, the characterization of the recovered mutants and a comparison of the mutation spectrum in the rat lymphocyte system to the spectrum in cancer genes, such as H-ras and p53, and the spectrum in transgenic systems, such as lacI, are included. The development of the rat lymphocyte hprt system and validation of the assay at the molecular level, provide an effective and reliable measure of genetic damage in an in vivo system which is readily comparable to measurement of genetic damage in the human.     

Application of the in vitro rat hepatocyte micronucleus assay in genetic toxicology testing

The investigation of micronuclei in mitogenic stimulated hepatocytes in vitro is a quite new area of research. Nevertheless, a relatively large database comprising more than 40 tested compounds of various classes has been generated up to now. This paper reviews the available data for the in vitro rat hepatocyte micronucleus assay, showing a sensitivity of this assay in identifying mutagens and genotoxic liver carcinogens of about 85%. Additionally, all of the tested non-carcinogens gave negative results. The use of primary hepatocytes instead of permanently dividing mammalian cell lines for the investigation of micronucleus induction has several advantages. (1) The broad spectrum of metabolizing enzymes expressed in primary hepatocytes ensures an adequate activation of most xenobiotics. (2) No transfer of activated metabolites via the culture medium is necessary in this system, since the metabolizing cells are the target cells themselves. (3) Whilst in experiments with permanently dividing cells the use of S9-mix restricts the treatment period with the test compounds to 2-6 h in the hepatocyte micronucleus assay continuous treatment of up to 48 h is possible. Investigations with the pyrrolizidine alkaloids retrorsine, monocrotaline and isatidine, strong mutagens and liver carcinogens, clearly showed that at least for isatidine a prolonged exposure period is essential to detect its mutagenic potential. This compound gave positive results in rat hepatocytes but not in V79-cells/S9-mix cultures. (4) The results obtained with the hepatocyte micronucleus assay are in good agreement with the genotoxic profiles of most of the compounds tested. Only three polycyclic aromatic hydrocarbons led to 'false-negative' results, since they strongly inhibited hepatocyte proliferation and thereby prevented micronucleus formation. (5) Hepatocytes are target cells of special interest when compounds are investigated which act specifically in the liver. Especially for hepatocarcinogens classified as non-genotoxins in standard genotoxicity tests or for chemicals showing DNA-repair induction in hepatocytes but no mutagenicity in standard tests, the hepatocyte micronucleus assay can contribute to clarify the situation. (6) The rat hepatocyte micronucleus assay can be performed easily and without great efforts in parallel to the in vitro hepatocyte DNA repair test (UDS-test), using the same hepatocyte batches. (7) Similar to the two versions of the UDS-test, the hepatocyte micronucleus assay can be performed following an in vivo-in vitro protocol. In order to further validate the hepatocyte micronucleus assay, as a next step controlled interlaboratory studies should be initiated.     

土壤中挥发性有机物的GC-MS测定

利用索氏提取-气相色谱-质谱联用技术,对某废弃化工厂原厂区土壤中可能存在的挥发性有机污染物进行定性和定量分析;利用索氏提取进行土样中有机物的提取,用十四烯配制标准曲线,以质谱全扫描方式的数据进行定性,选择离子监测的数据进行外标法定量,提高检测的灵敏度和准确度.依照美国环保局对环境分析的质量保证和质量控制指标对分析结果进行检测,发现方法的检出限为0.024mg·kg-1,回收率为95.84%,结果符合要求.     

Pharmacokinetics of and CYP1A induction by pyridine and acetone in the rat: interactions and effects of route of exposure

1. Male Sprague-Dawley rats were exposed to either pyridine, acetone or a combination of both compounds by either intraperitoneal administration (100 mg/kg pyridine or 400 mg/kg acetone) or whole-body inhalation (200 ppm pyridine or 1000 ppm acetone). Plasma and tissue levels of both compounds were determined by gas chromatography/mass spectrometry. 2. Both chemicals were well distributed in the tissues examined following either route of exposure, with concentrations in the order kidney > liver > plasma > lung. 3. Plasma half-life of pyridine was 7 h following a single 100 mg/kg dose of the compound, and 8 h following the last dose of a 3-day, 8 h/day exposure to a 200 ppm inhalation dose of the compound. 4. Plasma half-life of acetone was 4 h and was independent of the route of exposure. 5. The pharmacokinetics of pyridine was not affected by co-exposure to acetone. Similarly, the pharmacokinetics of acetone was not affected by co-exposure to pyridine. 6. Ethoxyresorufin O-deethylase activity in lung and liver and methoxyresorufin O-demethylase activities in liver were induced by pyridine but not by acetone at the doses examined. Pyridine-induced ethoxyresorufin O-deethylase activity was higher following inhalation exposure than following i.p. administration of pyridine but did not parallel tissue levels of the compound.     

Pharmacokinetics and organ distribution of cationized colchicine-specific IgG and Fab fragments in rat

Pharmacokinetics of cationized goat colchicine-specific polyclonal immunoglobulin G (IgG) and antigen binding fragment (Fab) (cIgG and cFab, respectively) were studied in male adult Sprague-Dawley rats and compared with those of the native proteins (nIgG and nFab). All proteins were radioiodinated by the Iodogen method, and kinetics were investigated following trichloroacetic acid (TCA) precipitation or immunoprecipitation. Deiodination and catabolism were more pronounced with the cationized than the native proteins, especially for cFab. Both cIgG and cFab in plasma decreased more rapidly than nIgG and nFab. The elimination half-lives were 52.9 and 81.8 h for cIgG and nIgG, respectively. In addition, there was a 74-fold increase in the volume of distribution and a 114-fold increase in the systemic clearance of cIgG compared with nIgG. For cFab, the volume of distribution and systemic clearance were increased 6.4- and 3.5-fold, respectively. Organ uptake of cIgG and cFab was markedly increased compared with that of nIgG and nFab, especially in kidney, liver, spleen, and lung. Renal clearance of cIgG and cFab was also increased 30- and 10-fold compared with that of nIgG and nFab, respectively. The present data suggest that cationization of colchicine-specific IgG and Fab fragments increased the organ distribution and greatly altered their pharmacokinetics. Nevertheless, the smaller molecular size of Fab versus IgG did not enhance the distribution and clearance of cFab. These data pave the way for evaluating the biological efficacy of these more tissue-organ-interactive antibodies.     

气相色谱-质谱法测定普洱茶挥发油的化学成分

[目的]为普洱茶产品质量标准的制定提供依据.[方法]采用水蒸气回流法提取普洱茶中的挥发油,并对其进行气相色谱-质谱分析,采用面积归一化法对各组分进行定量.[结果]从普洱荼挥发油中共鉴定出50种化合物,其主要成分为n-棕榈酸,其次为(Z,Z)-9,12-十八碳二烯酸、顺式芳樟醇、植醇、十四烷、1,2.3-三甲氧基苯、1,2-二甲氧基苯、己醛、二十一烷和苯甲醛,以上成分占挥发油相对含量的91.43%.[结论]普洱茶挥发油中主要含醇类、酸类、醛类、烃类、酯类、杂氧类、芳香族、杂环类等7类化合物.     

Nuclear estradiol receptor in the adult rat uterus: a new exchange assay

A protamine exchange assay has been developed to measure uterine nuclear estrogen receptor in mature rats exposed to estradiol (E). After ovariectomized-adrenalectomized mature rats are injected with E, estrogen receptor (RnE) is extracted from uterine nuclei with 0.6 M potassium chloride, diluted, and quantitatively precipitated with protamine sulfate. The precipitate is subjected to a ligand exchange with radiolabeled estradiol (E), with or without unlabeled diethylstilbesterol, to determine nonspecific binding. At 37 degrees C complete exchange of E for E in RnE is observed at 2.5 h; virtually no receptor degradation occurs up to at least 5 h. Exchange does not occur at 4 degrees C. Using the protamine assay, the depletion of cytoplasmic estrogen receptor (Rc) and the accumulation of RnE were studied at various doses of E at specific time points. Increasing doses of E result in a decrease of Rc with an equal increase of RnE. At the highest dose of E (10 mug) Rc is completely depleted within 10 min, by 6 h it is 25% replenished, and by 24 h returns to slightly above control levels. Within 10 min after the injection, RnE increases to 80-90% of the original cytoplasmic level of receptor (approximately 2-3 pmol/mg of DNA or approximately 1.5 pmol/100 mg of uterus). At 6 h RnE is 75% depleted and it is completely absent at 24 h. The protamine assay permits precise quantitative studies of nuclear estrogen receptor and avoids the problems of receptor degradation and excessive nonspecific binding often found in exchange reactions at elevated temperatures.     

Pharmacokinetics of the penicillins in man

The purpose of this article is to review and summarise those aspects of the pharmacokinetic behaviour of the penicillins that may be of particular interest to the clinician. While these antibiotics differ markedly in their acid stability and oral absorption, misleading inferences may be drawn from simple inspection of the maximal serum concentrations produced by a given dose administered orally. A more accurate picture emerges when serum protein binding and intrinsic activity of the drugs are taken into account. All of the penicillins are readily and actively secreted by the renal tubles and most are eliminated, almost completely unchanged, in the urine. The majority are excreted in small quantities in the bile, but this is a major route for elimination of nafcillin from the body. Distribution of the penicillins in 'non-specialised' sites is excellent. In contrast, penetration of the central nevous system and eye are poor, and of the prostate, minimal. Inflammation reduces the barries to penetration of these areas. However, quantitative data related to this phenomenon in man are few. Probenecid actively competes with the 'export' pump of the meninges and renal tubular cells. This results in an increase in concentrations of the penicillins in the blood and cerebrospinal fluid. The effect of this agent on active secretion of these antibiotics from the eye and biliary tract is minimal. While elimination of the penicillins from the body takes place largely via renal excretion, penicillin V and oxacillin are extensively degraded as well. In contrast to the situation with respect to 'natural' and 'broad-spectrum' penicillins, the serum half-life of the isoxazolyl congeners and nafcillin is only minimally prolonged in the presence of renal failure. These agents are only weakly haemodialyzable, while the other penicillins are rapidly removed from the circulation by this procedure.     

Pharmacokinetics of acyclovir in the cat

To investigate the detoxification of bromobenzene-induced hepatic lipid peroxidation by Oenanthe javanica DC, the hepatic lipid peroxide level and the activities of enzymes responsible for production and removal of epoxide were studied. The level of lipid peroxide elevated by bromobenzene was significantly reduced by the methanol extract (250 mg/kg) and persicarin (5 mg/kg). The methanol extract and persicarin administered daily over 4 weeks before intoxication with bromobenzene did not affect the activities of aminopyrine N-demethylase, aniline hydroxylase, and glutathione S-transferase. Epoxide hydrolase activity was decreased significantly by bromobenzene, which was restored to the control level by pretreatment with persicarin. However, the identical pretreatment with isorhamnetin and hyperoside did not change the enzyme activity or lipid peroxide level. The results suggest that the reduction of bromobenzene-induced hepatic lipid peroxidation by O. javanica under our experimental conditions is effected through enhancing the activity of epoxide hydrolase, an enzyme removing bromobenzene epoxide. In addition, the bioactive component of this plant responsible for the detoxification of bromobenzene, at least in part, is thought to be persicarin.     

Pharmacokinetics of the enantiomers of verapamil after intravenous and oral administration of racemic verapamil in a rat model

Verapamil is a chiral calcium channel blocking drug which is useful clinically as the racemate in treating hypertension and arrhythmia. The published pharmacokinetic data for verapamil enantiomers in the rat model are limited. Utilizing a stereospecific high-performance liquid chromatographic (HPLC) assay, the enantiomeric disposition of verapamil is reported after intravenous (1.0 mg kg-1) and oral (10 mg kg-1) administration of racemic verapamil to the rat model. After intravenous administration the systemic clearance of R-verapamil was significantly greater than that of S-verapamil; 34.9 +/- 7 against 23.7 +/- 3.7 mL min-1 kg-1 (mean +/- SD), respectively. After oral administration, the clearance of R-verapamil was significantly greater than that of S-verapamil, 889 +/- 294 against 351 +/- 109 mL min-1 kg-1, respectively. The apparent oral bioavailability of S-verapamil was greater than that of R-verapamil, 0.074 +/- 0.031 against 0.041 +/- 0.011, respectively. These data suggest that the disposition of verapamil in the rat is stereoselective; verapamil undergoes extensive stereoselective first-pass clearance after oral administration and the direction of stereoselectivity in plasma is opposite to that observed in the human.     

Pharmacokinetics of CDDP by the route of administration in patients with ovarian cancer

Seventy patients with local squamous cell carcinoma of the esophagus were treated between 1981 and 1990 with preoperative chemotherapy, surgical resection, and possible postoperative radiation therapy and/or chemotherapy. Chemotherapy included two cycles of 5-fluorouracil (1000 mg/m2) by continuous intravenous infusion on days 1-4 and cisplatin (100 mg/m2) on day 4. Complete clinical response (CCR) was achieved in 28 (41%) patients, partial clinical response (PCR) in 17 (25%), and no response in 23 (34%). Fifty-five (81%) patients were resected, 6 (9%) were explored, and 7 (10%) were unable to have surgery. Microscopic analysis of 55 resected patients showed 50 (91%) with active tumor, 1 (2%) with necrotic tumor, and 4 (7%) with a pathological complete response to chemotherapy. Twenty-six of the 55 resected patients (47%) had no gross evidence of disease at the time of surgical inspection. Median overall survival was 21.86 months (range 2-107 months) for all patients and 26.71 months (range 2-107 months) for resected patients. Actuarial 5-year survival rate was 31% for all patients and 39% for resected patients. Prolonged survival correlates with complete clinical response to chemotherapy, low pathological stage of disease, and successful resection of the lesion.     

Pharmacokinetics and metabolic disposition of 14C-metronidazole-derived radioactivity in rat after intravenous and intravaginal administration

Urinary metabolites and the pharmacokinetics of radioactivity derived from 14C-metronidazole (14C-MTZ) were determined after intravenous (iv) or intravaginal (ivg) administration of 10 mg/kg to adult rats. Following iv or ivg administration, the disappearance of 14C from blood followed the kinetics of a two-compartment open-system model. The blood half-lives of 14C during the beta-phase were 10.9 +/- 1.6 and 13.6 +/- 4.2 hr, after iv and ivg administration, respectively. After ivg application, the MTZ-derived radioactivity was detected in tail blood at 5 min, peaked at 1 hr, declined rapidly to 6 hr and more slowly thereafter. The vaginal absorption half-life of 14C-MTZ was 0.28 +/- 0.09 hr. About 12% of the administered dose remained in the vagina after 1 hr and 1.5% after 24 hr. At 24 hr, the tissue distribution and concentration of 14C were similar in iv and ivg dosed rats, the highest 14C concentration being present in the kidneys and lowest in the fat. The percentages of the dose excreted in 24 hr in the urine and feces were 58 and 15 after iv administration, compared to 37 and 40 after the ivg route, respectively. Unchanged 14C-MTZ and five of its metabolites were detected in the urine irrespective of the route of administration. The results show that metronidazole is rapidly absorbed through the vaginal mucosa of the rat and the metabolism and excretion of this chemotherapeutic agent are influenced by the route of administration.