In vitro metabolism and toxicity assessment of toxin T-514 (Peroxisomicine A1) of Karwinskia humboldtiana in microsomes and primary cultured hepatocytes.

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.

In vitro binding studies of the peroxisomicine A1-BSA and -HSA interactions.

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.

Cytotoxic hydroxyanthracenones from fruits of Karwinskia parvifolia.

Peroxisomicines A1 (1) and A2 (2) are cytotoxic hydroxyanthracenones isolated from the fruits of Karwinskia parvifolia. Peroxisomicine A1 exhibits selective cytotoxicity against several tumor cell lines. In the present investigation three minor constituents were obtained from the same fruit extracts: peroxisomicine A3 (3) and isoperoxisomicines A1 (4) and A2 (5). Compounds 3 and 5 are novel and their structures were determined by spectroscopic methods and molecular modeling. Compounds 1-5 were significantly cytotoxic against hepatoma cell lines.

Biologically active anthracenones from roots of Karwinskia parvifolia.

Tullidinol, a neurotoxin isolated from fruits and roots of several Karwinskia species, was resolved for the first time into two stereoisomers. This was achieved by means of preparative HPLC from roots of Karwinskia parvifolia. Structural assignments were made on the basis of spectroscopic data, including long range correlations as well as geometry optimization by means of semi-empirical methods.

Experimental acute intoxication with ripe fruit of Karwinskia humboldtiana (Tullidora) in rat, guinea-pig, hamster and dog.

Previous acute toxicity studies with 'Tullidora' in mice showed mainly hepatic and pulmonary complications. We tested samples in rat, guinea-pig, hamster and dog, searching for the mechanism of death. A single oral preparation of ripe fruit of Karwinskia humboldtiana was given to all animals. Clinical signs and histopathology were reproduced in all species except dogs. We propose a selective toxicity of the toxins of this fruit to the lungs and liver.

Extraction and quantification of toxins from Karwinskia humboldtiana (tullidora).

There is no mention in the literature of any analytical method to quantify toxins present in fruits of Karwinskia humboldtiana. It is recommended that thin layer chromatography and densitometry in situ be used, since it is rapid, sensitive and specific. Great variations in the toxin content of fruits collected in different regions of the state of Nuevo Leon (Mexico) were found, showing the importance of applying an analytical method before beginning toxicological experiments.

Experimental intoxication with fruit and purified toxins of buckthorn (Karwinskia humboldtiana).

To determine the extent of extraneural lesions in buckthorn poisoning, 180 CD1 mice were administered either green or ripe fruit or toxins T-544 or T-514 obtained from the fruit of the plant and were observed over a period of three weeks. Marked weakness, hyporeflexia, hair bristling, ptosis, spinal deformity, weight loss and dypsnea were prominent signs. Mortality in mice given green fruit was 100% at all doses; with toxin T-514 the mortality was 100% at 45 mg/kg. One hundred and sixty-two necropsies were performed and major lesions were found in liver and lung. The pulmonary lesions consisted of progressive vascular congestion and hemorrhage. Alterations in liver consisted of congestion, hemorrhage, hepatocyte degeneration, central zone necrosis and acute diffuse necrosis. Green fruit was more toxic than ripe fruit and T-514 was more active than T-544.

The glycoconjugates from Trypanosoma Cruzi, purification and composition of a glycoprotein.

1. The complex of glycoconjugates extracted with phenol-water from culture epimastigotes of a non-infective line (To) of Trypanosoma cruzi, derived from the Tulahuén strain showed only two components by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. 2. The slower migrating glycoconjugate, a glycoprotein of molecular weight 45, 000 was purified to apparent homogeneity. 3. A content of 76% protein, 17% neutral sugars and 6% glucosamine was analysed. Galactose and mannose occur in a ratio of 3:1 as determined by gas-liquid chromatography. A trace of glucose was detected in some samples.