ABSTRACT
T-514 (Peroxisomicine A(1)) from Karwinskia humboldtiana is a dimeric hydroxyanthracenone with a highly selective cytotoxic effect on tumor cells. We evaluated the metabolism of this compound in two in vitro systems (liver microsomes and hepatocytes) and assessed the cytotoxicity of its metabolites on normal and tumor cells. Microsomes (12.5, 125 and 250 microg of protein/ml) and hepatocytes (1 x 10(6) cells/ml) were incubated with the toxin (25 microM) for 0.5, 1, 3, 6, 9, 12 and 24 h and the samples were examined using chromatographic analysis and UV spectra. Two metabolites (M1 and M2) were detected in the rat microsomes and one (M1) in the monkey microsomes. The retention times and UV spectra of the peaks were very similar to those of the toxin T-514. M1 was isolated and identified as a mixture of two isomers. The cytotoxicity of the metabolites was evaluated in Chang liver and Hep G2 cells but they did not show the selective cytotoxic effect on tumor cells seen in the original compound.
Subject(s)
Anthracenes/toxicity , Cytotoxins/toxicity , Hepatocytes/drug effects , Karwinskia/chemistry , Microsomes, Liver/drug effects , Animals , Cells, Cultured , Chromatography, High Pressure Liquid , Dogs , Dose-Response Relationship, Drug , Drugs, Chinese Herbal/toxicity , Guinea Pigs , Haplorhini , Hepatocytes/pathology , Humans , Magnetic Resonance Spectroscopy , Mass Spectrometry , Mice , Microsomes, Liver/metabolism , Rats , Species Specificity , Ultraviolet TherapyABSTRACT
In the present study we have analyzed the production of reactive oxygen species by toxin T-514 of the genus Karwinskia in vitro (primary liver cell cultures and microsomes), as well as their possible role in its cytotoxicity. The role of catalase and superoxide dismutase (SOD) as defense mechanisms against oxidative stress was also studied. Freshly isolated hepatocytes or microsomes were exposed to T-514 in the presence or absence of catalase and SOD. Cytotoxicity was determined by methylthiazoltetrazolium (MTT) reduction. Oxidative stress was evaluated by the dichlorofluorescein diacetate (DCFDA) fluorescent probe and the reduction of ferricytochrome c. Exposure of hepatocytes to toxin T-514 for 2-, 4-, 6- and 24-h periods resulted in a time- and concentration-dependent increase in the suppression of mitochondrial metabolic activity. T-514 induced the production of reactive oxygen species in both hepatocytes and microsomes. Catalase and superoxide dismutase had a protective effect against the cytotoxicity of T-514 in hepatocytes and also inhibited the production of oxygen reactive species in microsomes. The results indicate that oxidative stress mediated by reactive intermediates may be a mechanism by which T-514 induces its cytotoxic effect.
Subject(s)
Anthracenes/adverse effects , Karwinskia/chemistry , Oxidative Stress , Reactive Oxygen Species , Animals , Catalase/pharmacology , Cytotoxins , Hepatocytes , Liver/cytology , Male , Microsomes, Liver , Rats , Rats, Sprague-Dawley , Rats, Wistar , Superoxide Dismutase/pharmacologyABSTRACT
Peroxisomicine A1 is a potential antineoplastic substance extracted from plants of the genus Karwinskia. An RP-HPLC-DAD method was developed and validated for the separation and quantification of four isomers of this compound. These isomers coelute in the preparative procedure and are present at a proportion ranging between 3 and 5% in the peroxisomicine A1 purified in the laboratory. The desirability coefficient of the method described here was enhanced 140% with respect to the previously reported method.
Subject(s)
Anthracenes/analysis , Chromatography, High Pressure Liquid/methods , Reproducibility of ResultsABSTRACT
The efficacy of meso-2,3-dimercaptosuccinic acid (DMSA) was evaluated in workers occupationally exposed to lead (Pb; blood level >50 microg/dL). Ten men were given 600 mg of DMSA per orem daily, for five days. Pb concentrations of whole blood and urine were determined throughout therapy. Hematology analyses, blood chemistry, and urinalysis were obtained at the start of the study, at the end of the DMSA treatment, and at 72 hours after the administration of the final dose. DMSA therapy had no influence on hepatic, hematologic, or renal functions and was effective in decreasing the concentration of blood Pb in all the subjects without adverse drug reactions.
Subject(s)
Chelating Agents/therapeutic use , Lead Poisoning/drug therapy , Lead/urine , Occupational Diseases/drug therapy , Succimer/therapeutic use , Administration, Oral , Adult , Chelating Agents/administration & dosage , Chelating Agents/adverse effects , Humans , Lead/blood , Lead Poisoning/blood , Lead Poisoning/urine , Male , Occupational Diseases/blood , Occupational Diseases/urine , Succimer/administration & dosage , Succimer/adverse effects , Treatment OutcomeABSTRACT
Peroxisomicine A1 (PA1) is a dimeric hydroxyanthracenone isolated from fruits of plants belonging to the genus Karwinskia. Showing selective toxicity between malignant and benign cell lines, it is currently under screening as an antineoplastic agent. Very little is known about its mechanism of action. In the present work the extent of binding of this substance with Bovine Serum Albumin (BSA) and Human Serum Albumin (HSA) at pH 7.2 and 7.4 has been evaluated using the spectrophotometric method. Absorbance of PA1 was altered by the presence of albumin and this property was used to generate binding isotherms. The investigation was carried out at four different temperatures. The data were analyzed by assuming two types of binding sites. Results indicated that PA1 binds to both albumins at physiological pH, which is reflected by the affinity constants of the order of 10(5). There are two types of binding sites in the albumin for PA1; with the electrostatic forces being discarded, the hydrophobic and hydrogen bond are more probable. Binding with HSA is stronger than with BSA.
Subject(s)
Anthracenes/metabolism , Antineoplastic Agents, Phytogenic/metabolism , Animals , Cattle , Humans , Hydrogen-Ion Concentration , Kinetics , Protein Binding , Serum Albumin , Serum Albumin, Bovine , Spectrophotometry, Ultraviolet , ThermodynamicsABSTRACT
OBJECTIVE: To evaluate the effects of intravenous sodium 2,3-dimercaptopropane-1-sulfonate (DMPS, Dimaval) on urinary excretion of essential trace elements in subjects who received this chelating agent as a mercury challenge test. SUBJECTS: Eleven subjects sought medical attention due to concern with the toxicity of mercury released from dental amalgam fillings. DESIGN: The subjects were given DMPS 3 mg/kg intravenously. Spot urine samples were collected 1 hour before and 1 hour after the DMPS dose for laboratory analysis. In addition to mercury, the urinary excretion of copper, zinc, selenium, magnesium, manganese, molybdenum, chromium, cobalt, and aluminum were measured. RESULTS: A significant increase in urinary excretion of mercury (3- to 107-fold) was observed after the DMPS dose. The DMPS treatment led to a 2- to 119-fold increase in copper excretion; 3- to 43.8-fold in selenium excretion; 1.6- to 44-fold in zinc excretion; and 1.75- to 42.7-fold in magnesium excretion. The excretion of manganese, chromium, cobalt, aluminium, and molybdenum remained unchanged. CONCLUSIONS: In this study, an intravenous DMPS challenge test produced a significant increase in mercury excretion and also led to an increased excretion of copper, selenium, zinc, and magnesium.
Subject(s)
Chelating Agents/pharmacology , Mercury/urine , Metals/urine , Trace Elements/urine , Unithiol/pharmacology , Chelating Agents/administration & dosage , Chelating Agents/adverse effects , Dental Amalgam , Female , Humans , Male , Mercury Poisoning/diagnosis , Unithiol/administration & dosage , Unithiol/adverse effectsABSTRACT
Previously we demonstrated that peroxisomicine A1 (T-514), a plant toxin isolated from Karwinskia species, has a deteriorating effect on the integrity of peroxisomes of methylotrophic yeasts. Here we describe two strains of Hansenula polymorpha, affected in the normal utilization of methanol as sole source of carbon and energy due to peroxisomicine A1 treatment. The two strains isolated (L17 and RV31) grew poorly on methanol, apparently due to malfunctioning of their peroxisomes. Moreover, the cells displayed a high peroxisome turnover rate. We argue that the peroxisomicine A1 induced phenotype of both strains is due to a genomic mutation. Strain L17 was functionally complemented after transformation with a H. polymorpha genomic library. The complementing 2.8 kb DNA fragment did not contain a well-defined ORF and led us to speculate that it may contain regulatory sequences that, when present in multiple copies in the cell, result in a change of expression of specific genes, thus causing restoration of normal methylotrophic growth.
Subject(s)
Anthracenes/toxicity , Methanol/metabolism , Microbodies/drug effects , Pichia/metabolism , Plant Extracts/toxicity , Cloning, Molecular , Drug Interactions , Immunohistochemistry , Microbodies/chemistry , Microbodies/metabolism , Microscopy, Electron , Pichia/classification , Pichia/drug effects , Pichia/genetics , Pichia/ultrastructureABSTRACT
Peroxisomes are single-membrane-bound organelles present in almost all eukaryotic cells. Hypolipidemic agents such as clofibrate, herbicides and plasticizers induce an increase in the number and size of peroxisomes from mammalian cells. However, there is no evidence of drugs causing a decrease in the number of these organelles. In this paper, we report the effect in vivo of toxin T-514 extracted from the plant Karwinskia humboldtiana, now re-named peroxisomicine-A1, on hepatic peroxisomes from rats intoxicated with this compound. Rats were treated with a single dose of 25 mg/kg of peroxisomicine-A1 and at different times were killed by decapitation. For the peroxisomal counting, liver tissue sections from control and treated rats were processed for the localization of catalase in peroxisomes. The results of the quantitative analysis demonstrated a significant decrease in the number of liver peroxisomes from rats intoxicated with peroxisomicine-A1. This finding suggests that peroxisomicine-A1 as in yeast, causes a damage to mammalian peroxisomes. The diminution in the number of peroxisomes could be a consequence of damage to the organelle, which is further removed by an autophagic process.
Subject(s)
Anthracenes/toxicity , Cytotoxins/toxicity , Liver/ultrastructure , Microbodies/ultrastructure , Animals , Catalase/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Female , Liver/drug effects , Liver/enzymology , Male , Microbodies/drug effects , Microbodies/metabolism , Rats , Rats, Wistar , Time FactorsABSTRACT
Dimeric anthracenones have been isolated from toxic plants of the genus Karwinskia (Rhamnaceae). T 514 or peroxisomicine A1 is one of the anthracenonic compounds which produce irreversible and selective damage on the peroxisomes of yeast cells in vivo. In this paper we describe the effect of two structurally related anthracenones on cell viability and on the peroxisomes of the methylotrophic yeast Candida boidinii. As has been described for peroxisomicine A1, peroxisomicine A2 and T 544 caused a decrease in the viability of C. boidinii at all concentrations tested, and disruption of the peroxisomal membrane, T 544 showing the strongest effect. In C. boidinii cell death and peroxisomal damage seem to be independent events.
Subject(s)
Anthracenes/pharmacology , Antineoplastic Agents, Phytogenic/pharmacology , Microbodies/drug effects , Neurotoxins/pharmacology , Pyrans/pharmacology , Candida/drug effects , Candida/growth & development , Candida/metabolism , Microbodies/ultrastructure , Microscopy, Electron , Plant Proteins/pharmacologyABSTRACT
Peroxisomicine is a toxic compound isolated from plants of the genus Karwinskia (Rhamnaceae). This toxin produces irreversible and selective damage to the peroxisomes of yeast cells in vivo. Peroxisomicine also inhibits catalase activity in vitro, when using purified enzyme. This paper reports on the effect of peroxisomicine on liver catalase in tissue fragments, in situ, as well as in mice intoxicated with peroxisomicine, in vivo. The catalase activity was determined by biochemical and histochemical methods. In contrast with the reported findings in vitro, the results demonstrate that there is no inhibition of the activity of tissue catalase, and suggest that catalase in situ and in vivo is protected against the inhibitory effect of peroxisomicine by an unknown factor.
Subject(s)
Anthracenes/toxicity , Catalase/metabolism , Liver/enzymology , Neurotoxins/toxicity , Pyrans/toxicity , Amitrole/toxicity , Animals , Chemical Fractionation , Cytosol/drug effects , Cytosol/enzymology , Liver/drug effects , Liver/pathology , Mice , Microbodies/drug effects , Microbodies/enzymology , Microbodies/pathology , Organelles/drug effects , Organelles/enzymologyABSTRACT
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 tested with Diast T-514 following enteral and parenteral administration. An LD50 dose by both oral and intraperitoneal administration showed selective damage to target organs.
Subject(s)
Anthracenes/toxicity , Antineoplastic Agents, Phytogenic/toxicity , Administration, Oral , Animals , Anthracenes/administration & dosage , Chemical and Drug Induced Liver Injury , Dimerization , Female , Injections, Intraperitoneal , Liver Diseases/pathology , Mice , Mice, Inbred Strains , Organ Specificity , Polymers/administration & dosage , Polymers/toxicity , Pulmonary Edema/chemically induced , Pulmonary Edema/pathologyABSTRACT
CASE REPORT: A case of intravenous self-administration of elementary mercury is presented. The increase urinary excretion of mercury after treatment with 2,3-dimercaptopropane-1-sulfonate is reported on a 5-year follow-up. No biochemical abnormalities in hepatic or renal function nor clinical pulmonary malfunction have been detected, despite the persistence of metallic densities in the body. The only persistent symptoms are tremor and lower extremity weakness. Any long term benefits of 2,3-dimercaptopropane-1-sulfonate treatment remains to be determined.
Subject(s)
Antidotes/therapeutic use , Mercury Poisoning/etiology , Suicide, Attempted , Unithiol/therapeutic use , Hand/diagnostic imaging , Humans , Injections, Intravenous , Male , Mercury/analysis , Mercury/urine , Mercury Poisoning/therapy , Middle Aged , Radiography, Abdominal , Radiography, ThoracicABSTRACT
OBJECTIVE: To evaluate clinical symptoms and urinary mercury before and after chelation therapy in subjects with chronic cutaneous mercurous chloride (HgCl; calomel) exposure. SUBJECTS: Twelve women from 19-45 years who had used a facial cream which contained HgCl (5.9%) for 2 to 10 years. DESIGN: Twenty-four hour urine samples were collected for basal urine mercury. All the subjects received a 5-day cycle of oral sodium 2,3 dimercaptopropane-l-sulfonate (Dimaval capsules 100 mg) 200 mg/d on an outpatient basis. The urine mercury excretion was monitored 24 hours after the first dose and 72 hours after the last dose in eight subjects. RESULT: Exanthem and tremor were detected in two of 12 subjects. The range of urine mercury was 180 to 1876 micrograms/g creatinine. A significant increase in the urinary mercury excretion was observed in the first 24 hours after beginning sodium 2,3-dimercaptopropane-1-sulfonate. CONCLUSION: Chronic topical application of 5.9% HgCl cream was associated with clinical mercurialism in two subjects and with high urinary mercury level in all the cases. Sodium 2,3-dimercaptopropane-1-sulfonate was effective in increasing urine mercury.
Subject(s)
Chelating Agents/therapeutic use , Mercury Compounds/poisoning , Mercury Poisoning/drug therapy , Mercury/urine , Unithiol/therapeutic use , Administration, Oral , Adult , Chelating Agents/administration & dosage , Face , Female , Humans , Mercury Poisoning/urine , Middle Aged , Ointments/poisoning , Skin Absorption/drug effects , Spectrophotometry, Atomic , Unithiol/administration & dosageABSTRACT
Dimeric anthracenones were isolated from toxic plants of the genus Karwinskia (Rhamnaceae). T 514 or peroxisomicine A1 is one of these toxic compounds which produces an irreversible and selective damage on the peroxisomes of yeast cells in vivo. In this paper we now report the inhibitory effect in vitro of peroxisomicine A1 and other structurally related anthracenones on liver catalase activity. The peroxisomicine A1 produces a non-competitive inhibition with respect to H2O2 on bovine, dog, and mouse liver catalases. In the three cases Vmax was decreased whereas Km was unaffected. Other dimeric anthracenones of natural origin were also found to be inhibitors of bovine liver catalase. There is a relationship between structure and degree of inhibition of all anthracenonic compounds tested. Peroxisomicine A1 and peroxisomicine A2 caused the highest degree of inhibition (IC50 = 3.34 and 3.64 microM, respectively).
Subject(s)
Anthracenes/pharmacology , Catalase/antagonists & inhibitors , Liver/enzymology , Plants, Toxic , Animals , Cattle , Dogs , Kinetics , MiceABSTRACT
The ingestion of ripe fruit of the Karwinskia humboldtiana, a shrub commonly known as tullidora or coyotillo, produces an intoxication described in the literature as a symmetric flaccid paralysis of the hind limbs, progressive and ascendent, that in severe cases may cause bulbar paralysis and death. The cause of an acute accidental intoxication of an entire family is presented here, wherein ten out of thirteen members ingested the ripe fruit of the tullidora. Three died, the father and two daughters. For the first time the toxins determination in blood by thin layer chromatography method is described. This method supports the diagnosis with other polyradiculoneuritis such as poliomyelitis and the Guillain Barre's syndrome.
Subject(s)
Fruit/poisoning , Paralysis/etiology , Child , Child, Preschool , Family Health , Female , Humans , Male , Middle Aged , Muscle Hypotonia , Paralysis/blood , Toxins, Biological/bloodABSTRACT
Intoxication produced by Karwinskia humboldtiana presents a neurological picture similar to that of poliomyelitis, Guillain-Barré syndrome or other polyradiculoneuritis with which it is frequently confused. The purpose of this paper is to report the frequency of this intoxication, by means of the antecedent of ingestion of the fruit and the detection of toxins in blood using a thin layer chromatography method. One hundred fifty four samples of cases with acute flaccid paralysis from 18 states of the country were received. The antecedent of ingestion in 56 of them was corroborated and the detection was positive in 50 of these. In 98 patients there was not antecedent of ingestion and detection was negative in 95 of them. We estimated that the sensibility and specificity of detection method are 89% and 96.9% respectively.
Subject(s)
Plant Poisoning/epidemiology , Plants, Medicinal , Plants, Toxic , Rhamnus , Adolescent , Aged , Child , Child, Preschool , Diagnosis, Differential , Female , Humans , Incidence , Infant , Male , Mexico/epidemiology , Muscle Hypotonia/diagnosis , Muscle Hypotonia/epidemiology , Muscle Hypotonia/etiology , Paralysis/diagnosis , Paralysis/epidemiology , Paralysis/etiology , Plant Poisoning/complications , Plant Poisoning/diagnosis , Polyradiculoneuropathy/diagnosisABSTRACT
The purpose of this study was to determine the clinical efficacy of 2,3-dimercapto-1-propane sulfonic acid, Na salt, on the urinary excretion of mercury as well as its possible adverse effects. Ten men with occupational mercury exposure (urinary level of 50 micrograms/g creatinine or more) were assigned to receive 2,3-dimercapto-1-propane sulfonic acid p.o. (DIMAVAL capsules, 100 mg) 300 mg/d for five days. Informed written consent was obtained from each subject. Hematology analyses, blood, chemistry, and urinalysis were obtained at the start of the study, at the end of the 2,3-dimercapto-1-propane sulfonic acid treatment and 72 hours after the administration of the final dose of 2,3-dimercapto-1-propane sulfonic acid. Twenty-four-hour urine mercury levels were closely monitored throughout therapy. All data and measurements before and during drug doses were evaluated by analyses of variance. In all subjects mean urine mercury was significantly increased (p < .05) after pre-2,3-dimercapto-1-propane sulfonic acid treatment. One subject had a moderate hypersensitivity reaction (rash) to 2,3-dimercapto-1-propane sulfonic acid but no other toxic effects were observed.
Subject(s)
Chelating Agents/therapeutic use , Mercury Poisoning/drug therapy , Mercury/urine , Occupational Diseases/chemically induced , Unithiol/therapeutic use , Adult , Chelation Therapy , Creatinine/urine , Humans , Male , Mercury Poisoning/urine , Middle Aged , Occupational Diseases/urine , Prospective Studies , Unithiol/adverse effectsABSTRACT
Toxin T-514 of Karwinskia humboldtiana has been demonstrated to be hepatotoxic in vivo and in vitro. Recently a diastereoisomer of T-514 has been isolated. In the present study we have evaluated and compared the in vitro hepatoxicity of the diastereoisomer of T-514 using primary cultures of rat hepatocytes. Cytotoxicity was evaluated by release of cytoplasmic enzyme lactate dehydrogenase (LDH), and mitochondrial metabolic function (MTT reduction). The diastereoisomer was shown to be almost as hepatoxic in vitro as toxin T-514.
Subject(s)
Anthracenes/toxicity , Cytotoxins/toxicity , Liver/drug effects , Analysis of Variance , Animals , Animals, Newborn , Anthracenes/chemistry , Cells, Cultured , Cytoplasm/enzymology , Cytotoxins/chemistry , Dose-Response Relationship, Drug , L-Lactate Dehydrogenase/metabolism , Liver/cytology , Mitochondria, Liver/drug effects , Mitochondria, Liver/enzymology , Plants, Medicinal , Plants, Toxic , Rats , Rats, Sprague-Dawley , Rhamnus , StereoisomerismABSTRACT
Toxin T-514 is a dimeric anthracenone isolated from the Karwinskia humboldtiana (buckthorn) plant. Its potential anti-neoplastic effect was evaluated in vitro and the results obtained were compared with the effect of other known anti-cancer agents. Normal and malignant continuous cell lines were tested. After a 72-h exposure, neoplastic cells derived from hepatic, pulmonary and colonic tissues were more sensitive to toxin T-514 than normal cells from the corresponding organ. Hepatoma cells and colon adenocarcinoma CT50 values were < 10 micrograms/ml. Lung adenocarcinoma, undifferentiated bronchogenic cancer cells and small cell carcinoma CT50 values were < 20 micrograms/ml. All benign cell CT50 levels tested were > 113 micrograms/ml. This in vitro selective toxicity found with toxin T-514 was also seen with 5-fluororacil and mitomycin for colon adenocarcinoma and with epidoxorubicin for undifferentiated bronchogenic cancer cells.