α-Hydroperoxy diethyl peroxide, an unusual natural compound isolated from an ascidian (Phallusia mammillata): acute toxicity in DDY mice

α-Hydroperoxy diethyl peroxide, a novel compound found in the tunic of ascidians, has two peroxide moieties per molecule. Since ascidians are a widely served food item in Japan, human exposure to this compound potentially exists in the seafood preparation industries. No toxicological data have so far been published on this compound, and so we determined the intraperitoneal 6-day LD₅₀ in mice and conducted histopathological examinations. The 6-day LD₅₀, was found to be 199 with 95% confidence limits of 126–314 . Histopathological examination revealed necrosis induced in a variety of cells that had been directly exposed to the compound. These cells included hepatocytes, parenchymal pancreatic cells and fat cells. It is concluded that direct contact with this compound is likely to elicit cellular necrosis of various organs. The specific toxicological effects are probably dependent on the route of exposure.     

1-Adrenoceptor subtype involved in the positive inotropic response to phenylephrine in rat atria

Phenylephrine produced positive inotropic responses in isolated rat right and left atria. The responses were competitively inhibited by ₁-adrenoceptor antagonists (prazosin, WB4101 and HV723) with relatively low affinities (pA₂ values close to 8.0). Chloroethylclonidine had no significant effect on the responses to phenylephrine. These results suggest that the positive inotropic response to phenylephrine in rat atria is mediated through ₁-adrenoceptors which cannot be defined by the _1A, _1B subclassification.     

Effect of atropine on cyanide-induced acute lethality in mice

The effects of atropine on acute lethality induced by cyanide were investigated in mice. The LD₅₀ value of cyanide (s.c. injection) was 8.4 (7.6–9.3) mg/kg. However, the LD₅₀ value of cyanide (s.c.) was significantly increased by 1.5-fold when atropine (32 mg/kg) was injected s.c. in mice. Furthermore, the combined administration of atropine (32 mg/kg). Ca²⁺ (500 mg/kg) and sodium thiosulfate (1 g/kg) tremendously increased the LD₅₀ value by 5.6-fold in mice although sodium thiosulfate or Ca²⁺ alone increased the LD₅₀ 2.5- or 1.5-fold. On the other hand, although the LD₅₀ value of cyanide (intracerebroventricular injection (i.v.t.)) was 52.0 (47.4–57.0) μ/brain, the LD₅₀ value of cyanide (i.v.t.) was significantly increased by 1.3- or 1.61-fold in mice 10 min after s.c. injection of atropine (32 mg/kg) or Ca²⁺ (500 mg/kg). Furthermore, the combined administration of atropine and Ca²⁺ increased the LD₅₀ value of cyanide by 2.1-fold. These results suggest that atropine inhibits cyanide-induced acute lethality and promotes the antagonistic effect of thiosulfate and Ca²⁺ in mice.     

Activity and gene expression profile of certain antioxidant enzymes to microcystin-LR induced oxidative stress in mice

Microcystins are cyclic heptapeptide toxins produced by certain strains of Microcystis aeruginosa and microcystin-LR (MC-LR) is the most toxic among the 70 variants isolated so far. These toxins have been implicated in both human and livestock mortality. In the present study we investigated the microcystin-LR induced oxidative stress in mice in terms of its effect on activity and gene expression profile of certain antioxidant enzymes and expression of heat shock protein-70 (HSP-70). Mice were treated with 0.5 LD₅₀ (38.31 μg/kg) and 1 LD₅₀ (76.62 μg/kg) and the biochemical variables were determined at 1, 3, 7 days and 15, 30, 60 and 120 min post-exposure for 0.5 and 1 LD₅₀ dose, respectively. A significant time-dependent increase in HSP-70 expression over control was observed at 1 LD₅₀ dose. The toxin induced significant increase in liver body weight index, hepatic lipid perxoidation and depletion of GSH levels at 1 LD₅₀ compared to control group. There was significant decrease in the activity of antioxidant enzymes glutathione peroxidase (GPX), superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR) and glutathione-S-transferase (GST) at 1 LD₅₀. Except catalase, there was no effect on other antioxidant enzymes at 0.5 LD₅₀ dose. In contrast to activity of antioxidant enzymes the gene expression profile did not show any significant difference compared to control at 1 LD₅₀. GR showed significant decrease in expression at 1, 3 and 7 days in animals dosed with 0.5 LD₅₀ MC-LR. The results of our in vivo study clearly show the oxidative stress induced by MC-LR, and a correlation with activity and regulation at gene expression level of antioxidant enzymes.     

Experimental evidence for toxic damage induced by a dimeric anthracenone: diast T-514 (peroxisomicine A2)

Dimeric anthracenones obtained from the genus Karwinskia (Rhamnaceae) are characteristic compounds isolated from the plants of this species. Previous toxicity studies demonstrated Diast T-514 to be toxic to animals in experimental settings. Diast T-514 extracted and characterized from Karwinskia parvifolia, was studied in CD1 mice. The LD₅₀ for this compound was determined. Animals were testedwith Diast T-514 following enteral and parenteral administration. An LD₅₀ dose by both oral and intraperitoneal administration showed selective damage to target organs.     

Effects of sarin on the operant behavior of guinea pigs

The present study evaluated the dose–response effects of subacute exposure to sublethal doses of the organophosphorus (OP) chemical warfare nerve agent (CWNA) sarin (GB) on the operant behavior of guinea pigs. Dietary restricted guinea pigs, trained to respond for food under a progressive ratio (PR) schedule of reinforcement, were injected five times per week (Monday–Friday) for 2 weeks with fractions (0.1, 0.2, and 0.4) of the established LD₅₀ of GB (42 μg/kg). Changes in body weight, whole blood (WB) acetylcholinesterase (AChE) levels, and operant performances were monitored over the 2 weeks of GB exposure and for an additional 2 weeks following the termination of exposures. There were dose-related changes in body weight and WB AChE levels throughout the exposure and post-exposure periods. Several parameters of PR performance were disrupted during exposure to 0.4 LD₅₀ GB, however, concurrent weight loss indicated the presence of overt toxicity. PR performance recovered following the termination of exposures. Lower doses (0.1 and 0.2 LD₅₀) of GB failed to produce reliable effects on operant performance during the exposure period. Overall responding decreased during exposure to 0.4 LD₅₀ GB, resulting in reduced response rates and break points. The decrease in overall response rates was attributed to an increase in pausing since there was no decrease in running rate. Motor effects of 0.4 LD₅₀ GB were evident as an increase in the proportion of lever press durations ≥ 1.0 s. In the present study, doses of GB lower than 0.4 LD₅₀ produced no marked alteration of operant performance in guinea pigs, although WB AChE levels were maximally inhibited to 20% of control.     

In vitro cytotoxicity tests for the prediction of acute toxicity in vivo

Investigations of the use of in vitro cytotoxicity tests for the prediction of acute toxicity in vivo have been reviewed with particular emphasis on those studies that have been published during the past 5 years. Numerous cell types, endpoints and exposure periods have been used in cytotoxicity tests, although these appear generally to have little effect on the resulting correlation between in vitro IC₅₀ values and in vivo LD₅₀ values. The in vitro data correlate better with rodent parenteral (ip or iv) LD₅₀ values than with oral LD₅₀ values due to kinetic considerations. For certain groups of related chemicals (e.g. antitumour compounds, metal salts), and for some sets of unrelated chemicals, the in vitro data correlate very well with LD₅₀ values. However, while cytotoxicity tests are useful for screening chemicals for their intrinsic and relative toxicities, it is impossible to tell whether predictions based on cytotoxicity data alone would be sufficiently accurate for labelling and classifying a new chemical according to its likely acute toxicity in vivo. The in vitro endpoints need to be of greater relevance to the possible mechanisms of chemically-induced acute toxicity in vivo than most of those that are used at present.     

1-Acid glycoprotein column in the high-performance liquid chromatographic analysis of some groups of chiral drugs

The usefulness of the ₁-acid glycoprotein (Chiral—AGP) column in the analysis of chiral drugs and related materials is illustrated in three different fields. Excellent separations of the enantiomers of pharmacologically active -ethyl benzhydrols such as 3-trifluoromethyl--ethyl benzhydrol (flumecinol) and 11 other derivatives are described. The enantiomers of N-protected amino-acid esters or amides can also be separated. The effect of the derivatization of the free -amino group on the resolution of the enantiomers is exemplified by the formylation of alanine benzyl ester. As an example of the use of the method for the estimation of the optical purity of chiral drugs, the determination of the “cis(−)” impurity in “cis(+)” diltiazem is presented.     

度洛西汀对小鼠急性毒性及遗传毒性的研究

目的:探讨度洛西汀对成年雄性小鼠的急性毒性及遗传毒性。方法:采用小鼠急性毒性试验观察致死率及半数致死量(LD₅₀)。采用小鼠骨髓微核试验、小鼠精子畸变试验及生殖与淋巴器官重量指数检测方法,将小鼠均分为6组,即阴性对照组、阳性对照组,度洛西汀1、2、3和4组(分别给予度洛西汀5、10、20和40 mg/kg),观察度洛西汀不同剂量对小鼠精子畸变率、骨髓微核率及生殖与淋巴器官重量指数的影响。结果:度洛西汀急性毒性试验结果表明,灌胃给予度洛西汀的LD₅₀为188.3 mg/kg。微核试验与精子畸变试验结果表明,度洛西汀灌胃剂量为5、10和20 mg/kg时(度洛西汀1、2和3组)均未诱发小鼠精子畸变率和骨髓微核率的改变,与阴性对照组相比,差异均无统计学意义(P>0.05);度洛西汀4组小鼠的精子畸变率、骨髓微核率均高于阴性对照组,差异有统计学意义(P<0.05)。度洛西汀1、2、3和4组小鼠的生殖与淋巴器官重量指数与阴性对照组相比,差异均无统计学意义(P>0.05)。结论:灌胃给予度洛西汀的LD₅₀为188.3 mg/kg,安全性高。度洛西汀以5、10和20 mg/kg的剂量灌胃对小鼠无生殖与遗传毒性,当灌胃剂量达到40 mg/kg时对雄性小鼠有轻微的潜在遗传毒性。     

Acute and subacute toxicity and efficacy of S-nitrosylated captopril, an ACE inhibitor possessing nitric oxide activities

The toxicity and efficacy of S-nitrosocaptopril (CapNO), a novel vasodilator possessing the capacities of both a nitric oxide donor and an angiotensin converting enzyme (ACE) inhibitor, were examined in rodents. In single-dose acute toxicity studies in ICR mice, the median lethal dose (LD₅₀) for CapNO was 674±94 mg/kg (iv) and 2078±100 mg/kg (po), whereas for oral captopril was 4286±173 mg/kg. S-nitrosoglutathione, containing the same S-nitroso moiety as CapNO, showed an LD₅₀ equal to CapNO when the values were expressed by the mol/kg. The cause of acute death by the high doses of CapNO was lethal hypotension. In the subacute toxicity studies, oral CapNO was well tolerated in normotensive and hypertensive rats at doses up to 500 mg/kg/day for 3 months, except for considerable reductions in food consumption and growth rate observed in the 500 mg/kg/day group. Serum chemistry and hematology tests performed in the subacute toxicity studies revealed no adverse effects of oral CapNO except for a significant decrease in cholesterol levels in hypertensive SHR rat. At autopsy, no histopathological changes in major organs were observed over the subacute period. Administration of a therapeutic dose of CapNO (iv, 250 μg/kg which produced 25% decreases in blood pressure) revealed no changes in the hematological parameters. Subchronic treatment of SHR and SS/Jr rats with oral CapNO (50 mg/kg/day) significantly reduced mean arterial pressure to the normotensive level. Considering the absence of adverse effects of CapNO in the subchronic toxicity study, CapNO appears to be a safe drug for further clinical trials, but particular caution must be taken because it can cause hypotension when overdosed.     

Tier-2 studies on monocrotaline immunotoxicity in C57BL/6 mice

Monocrotaline (MCT) is a member of a class of naturally occurring phytotoxins known as pyrrolizidine alkaloids, and is a toxicological concern to both man and his livestock. The purpose of these studies was to evaluate the effect of a 14-day oral MCT (0–100 mg/kg per day) exposure on the functional integrity of various immunocyte effector systems in C57BL/6 mice, as well as to investigate potential mechanisms for its immunotoxicity. Decreases in lymphoid organ weights and cellularity, and resident peritoneal exudate cell (PEC) number were only observed after exposure to the highest dose of 100 mg/kg MCT. This dose also inhibited NK cell cytotoxicity, while the total number of NK lytic units per spleen was decreased (−53%) after exposure to 50 mg/kg MCT. Following i.p. injection of SRBC, the percentage of PEC macrophages containing engulfed SRBC was significantly increased in MCT-exposed mice, while the percentage of large vacuolated (activated) macrophages was decreased in a dose-dependent manner. Exposure to MCT significantly decreased the total number of Ig+ cells without altering the number of CD4+ and CD8+ cells. The antibody responses (PFC/10⁶ spleen cells) to two T cell-independent antigens, TNP-LPS and DNP-Ficoll, were significantly decreased at all MCT doses, and the degree of suppression of both responses was identical at coincident doses. MCT exposure (25 mg/kg) significantly suppressed the blastogenic response to the T cell mitogen concanavalin A (−38%), and to the B cell mitogen lipopolysaccharide (−58%). These results indicate that exposure to MCT can alter the functional integrity of various immune effector responses in a dose-dependent manner, and suggest that the B cell may be a relatively more sensitive target of MCT immunotoxicity compared to T cells, macrophages and NK cells.     

Toxicokinetics and toxicity of thioacetamide sulfoxide: a metabolite of thioacetamide

Thioacetamide (TA) is bioactivated by CYP2E1 to TA sulfoxide (TASO), and to the highly reactive sulfdioxide (TASO₂), which initiates hepatic necrosis by covalent binding. Previously, we have established that TA exhibits saturation toxicokinetics over a 12-fold dose range, which explains the lack of dose–response for bioactivation-based liver injury. In vivo and in vitro studies indicated that the second step (TASO → TASO₂) of TA bioactivation is less efficient than the first one (TA → TASO). The objective of the present study was to specifically test the saturation of the second step of TA bioactivation by directly administering TASO, which obviates the contribution from first step, i.e. TA → TASO. Male SD rats were injected with low (50 mg/kg, ip), medium (100 mg/kg) and high (LD₇₀, 200 mg/kg) doses of TASO. Bioactivation-mediated liver injury that occurs in the initial time points (6 and 12 h), estimated by plasma ALT, AST and liver histopathology over a time course, was not dose-proportional. Escalation of liver injury thereafter was dose dependent: low dose injury subsided; medium dose injury escalated upto 36 h before declining; high dose injury escalated from 24 h leading to 70% mortality. TASO was quantified in plasma by HPLC at various time points after administration of the three doses. With increasing dose (i.e., from 50 to 200 mg/kg), area under the curve (AUC) and C_max increased more than dose proportionately, indicating that TASO bioactivation exhibits saturable kinetics. Toxicokinetics and initiation of liver injury of TASO are similar to that of TA, although TASO-initiated injury occurs at lower doses. These findings indicate that bioactivation of TASO to its reactive metabolite is saturable in the rat as suggested by previous studies with TA.     

Acute oral toxicity test and assessment of rhynchophylline in mice

As the main alkaloid constituent of Uncaria species, rhynchophylline has drawn extensive attention in recent years for its antihypertensive and neuroprotective activities. However, toxicity study of the rhynchophylline is still lacking. In the present study, oral acute toxicity of rhynchophylline was conducted in Kunming mice. The mice were orally treated with 520.00, 442.00, 375.70, 319.34 and 271.44 mg/kg of rhynchophylline for 14 d. The general behavior, body weight changes, toxic reaction, and death were recorded, and histopathological analyses were performed. The acute toxicity was evaluated by the assessment of the median lethal dose (LD₅₀). The acute toxicity study showed that no significant difference was found in the body weight of the mice in the control group and those in the drug group. However, the mice treated with rhynchophylline showed obvious abnormal symptoms and mortality. The median lethal dose (LD₅₀) of orally administered rhynchophylline was 308.08 mg/kg. The histopathological results showed that the mice in the high-dose rhynchophylline group displayed toxic effects in the brain, liver, lung, and kidney. The results of the current study indicated that rhynchophylline could not be taken at a high dose. Collectively, our current findings provided a strong basis for further clinical investigation.     

Toxicity of the dialyzable fraction of the venom of the yellow scorpion, Leiurus quinquestriatus, to the migratory locust

Dialysis was carried out on redissolved freeze-dried venom of the scorpion Leiurus quinquestriatus H. and E. The mortality rate of Locusta migratoria migratorioides R. & F. injected with the dialyzable portion was used for calculating the regression equation and the LD₅₀. It was found that the LD₅₀ per female locust (mean weight 1·8 g) was that amount which was dialyzed out of 161 μg of whole venom, which means that the dialyzable portion has 2·67 per cent of the lethality of the whole venom. The toxic dose of the dialyzed venom was found not to differ from that of the whole venom.     

Biological evaluation of N-octyl-O-sulfate chitosan as a new nano-carrier of intravenous drugs

An amphiphilic chitosan derivate, N-octyl-O-sulfate chitosan (NOSC) was prepared by octylation of amino group at C-2 position and sulfonylation at C-6 position. Micelle formed by NOSC has great capability in solubilization of water-insoluble drug paclitaxel. Enormous attention was attracted by the potential application of NOSC as a new drug delivery system. Tritium labeled NOSC (³H NOSC) was injected by tail vein at dose of 13.44 mg/kg in mice; kidney retained the maximum amount of NOSC all the time even after 24 h following the injection. Pharmacokinetic parameters (the area under the plasma concentration–time curve, maximum plasma concentration, apparent plasma half-life of distribution phase and elimination phase, mean residence time, apparent volume of distribution, total body clearance) were obtained by fluorometric method in rats. The results showed a linear pharmacokinetics proceeding of FITC-NOSC in vivo. 75.4 ± 11.6% ³H NOSC of dose was excreted in urine over a 7-day period, urinary excretion was the predominant way of excretion of NOSC compared with bilary or fecal pathway. A series of safety studies consisted of acute toxicity study, intravenous stimulation study, injection anaphylaxis study, hemolysis study and cell viability assay were performed to warrant the biocompatibility of the NOSC as intravenous materials. The LD₅₀ value of NOSC administrated by i.v. and i.p. were calculated as 102.59 and 130.53 mg/kg, respectively. No intravenous stimulation, injection anaphylaxis, hemolysis and cytotoxicity were observed in the safety studies. The tissue distribution, pharmacokinetics, excretion and safety study were persuasive for the potential application of NOSC as a new drug carrier.     

Limited immunotoxic potential of technical formulation of the herbicide atrazine (AAtrex) in mice

Immunotoxicity of the technical atrazine formulation, AAtrex, was examined in C57B1/6 female mice following a sublethal exposure to equivalent 1/2–1.64 LD₅₀ doses of the herbicide. Animal weight was not affected by the herbicide exposure. No dose-related changes could be concluded for fluctuations in organ weight, changes in the spleen cell number and cell viability. Furthermore, cytofluorometric studies showed no significant changes in the frequency of L3T4-positive and Lyt-2-positive T-cells. Functional in vitro assays of mitogen activation showed no marked effects of AAtrex exposure on lymphocyte stimulation by lipopolysaccharide (LPS), phytohemagglutinin (PHA) and concanavalin A (Con-A). In addition, sublethal exposure to AAtrex did not affect interleukin-2 (IL-2) production by splenic cells. Furthermore, no doserelated effect could be concluded from a transient suppression of a primary humoral IgM response to sheep erythrocytes (SRBC) as well as from a transient inhibition of a specific T-cell response to alloantigens in mixed lymphocyte reaction (MLR). Exposure to equiv. 1/2−1/16 LD₅₀ doses augmented phagocytic activity of peritoneal macrophages, without any visible AAtrex dose-related effect. Normal humoral and cellular responses were restored at 14–40 days after the herbicide exposure. Overall, transient and reversible immunosuppression of humoral-mediated and cell-mediated responses and activated macrophage phagocytic activity could not be attributed to the direct chemical-related effect of sublethal exposure to AAtrex.     

Experimental envenoming of mice with venom from the scorpion Centruroides limpidus limpidus: differences in mortality and symptoms with and without antibody therapy relating to differences in age, sex and strain of mouse

C57Bl/6J and BALB/cAnN inbred strains of mice differed significantly in mortality and symptoms when intoxicated subcutaneously with one LD₅₀ of venom from Centruroides limpidus limpidus. Higher mortality was observed in C57Bl/6J than in BALB/cAnN. Also, C57Bl/6J mice more quickly developed muscular and respiratory collapse whilst BALB/cAnN mice were hyperactive before dying. Also, the symptoms in the survivors lasted for 24 h in C57Bl/6J and for 2 h in BALB/cAnN. The age and sex of mice were also related to mortality: younger mice were more resistant than older mice and females were more susceptible than males, especially in the younger groups. Antivenom (horse F(ab′)₂) administration 5–10 min after envenoming of mice with one LD₅₀ rescued 60% of BALB/cAnN and 52% of C57Bl/6J mice, respectively. Results indicate that genetic background, gender and age differences are of consequence in the pathogenesis of C. limpidus scorpion envenomation in mice, and that timely treatment with active antivenom F(ab′)₂ saves a significant fraction of intoxicated mice without statistically significant distinction of strains.     

An assay of the toxicity of the Atlantic puffer fish, Spheroides maculatus

Skin, muscle, liver, and gonadal extracts of the Atlantic puffer, Spheroides maculatus were assayed for toxicity in white mice. Extracts were injected intraperitoneally and were administered in the amounts of 1 ml per 20 g of body weight. The LD₅₀ of a composite skin extract, administered intraperitoneally in progressive doses per 20 g of body weight, was determined for white rats, white mice, chicks and frogs (Rana pipiens), while a LD₅₀ of a composite muscle extract was determined for the pinfish, Lagodon rhomboides.     

Design,synthesis and anti-tumor activities of cyclic phosphoramidate mustard-quinazoline conjugates

A series of new cyclic phosphoramidate mustard-quinazolineconjugates were designed and synthesized based on the drug candidate EMB-3, a multi-target-directed ligand against tumor cells, and their anti-tumor activities were evaluated on breast cancer and lung cancer cells. Compound 6d exhibited the best anti-tumor performance with IC₅₀ = 0.6 μM(8-fold of EMB-3) on BT474breast tumor cells. Compound 6d inhibited epidermal growth factor receptor (EGFR, biomarker for NSCLC) and human epidermal growth factor 2 (HER2, biomarker for breast cancer) with IC₅₀ of 18 nM and 78 nM, respectively. The preliminary pharmacokinetic study revealed that 6d was more stable than EMB-3 during the in vivo metabolism. A single dose per os (PO) administration of 6d in rat model (10 mg/kg) resulted in a moderate t_1/2of 1.7 h. These results indicated that compound 6d was a potential lead compound for the treatment of breast cancer.     

Zootoxicological properties of venom L-amino acid oxidase

L-amino acid oxidase prepared from Crotalus adamanteus venom after the method of and does not contribute to the production of the profound fall in systemic arterial pressure usually seen following injection of crude Crotalus venom, nor does it provoke any early deleterious changes in the dependent variables of the cardiovascular system.

The enzyme has no effect on neuromuscular transmission in the mammal. Its LD₅₀ is 9·13 (7·83–10·35) mg per kg test animal body weight.