Cytotoxic sesquiterpene lactones from Inula britannica

Cytotoxicity-guided fractionation of the flowers of Inula britannica led to the isolation of four sesquiterpene lactones, 4 alpha, 6 alpha-dihydroxyeudesman-8 beta, 12-olide (1), ergolide (2), 8-epi-helenalin (3), and bigelovin (4). Compound 1 was isolated as a new natural product. These compounds showed cytotoxicity against human tumor cell lines.     

Further lupane lactones from Kokoona ochracea

Additional new lupane lactones were isolated from the stem bark of Kokoona ochracea (Celastraceae). Their structures have been elucidated, through the application of 1D and 2D nmr spectroscopic methods, as 20,29-dihydroxy-3-oxolupan-30,21 alpha-olide (ochraceolide D) [1] and 28-hydroxy-3-oxolup-20(29)-en-30,21 alpha-olide (ochraceolide E) [2]. These compounds and the mono- and di-acetates of ochraceolide D (4 and 5, respectively) were evaluated for in vitro cytotoxic activity against P-388 murine lymphocytic leukemia cells and a panel of human cancer cell systems. Ochraceolide D [1] was significantly cytotoxic (ED50, 3.9 micrograms/ml) against human glioblastoma (U373) cells. Other compounds (4, 5, and 2) exhibited only a weak cytotoxic response in certain cancer cell lines.     

Lignans and sesquiterpene lactones from Artemisia sieversiana and Inula racemosa

Alternatively spliced isoforms of Na(+)-Ca2+ exchanger were found from various tissues and species. RT-PCR amplification was performed on the basis of our cloned mouse cardiac Na(+)-Ca2+ exchanger and four alternatively spliced isoforms of Na(+)-Ca2+ exchanger were identified. Three (NCX1.3, NCX1.4, and NCX1.12) of them were first identified in the heart, and one isoform (NCX1.12) was a novel spliced variant. These four spliced variants were present in the embryonic and adult atria and ventricles. Different cell types of the heart expressed different spliced isoforms of Na(+)-Ca2+ exchanger. Southern blot analysis indicated that the Na(+)-Ca2+ exchanger gene existed as a single copy in the mouse genome. Thus, the Na(+)-Ca2+ exchanger isoforms expressed in mouse heart are consistent with being produced by alternative splicing and they may have different functions in various cell types in the mouse heart.     

Antimycobacterial activity of substituted isosteres of pyridine- and pyrazinecarboxylic acids

Pyrazines and pyridines substituted with alkylated tetrazoles, esterified vinylogous carboxylic acids, and ketosulfides were synthesized as precursors of antimycobacterial agents which, after penetration of the mycobacterial cell wall, could be biotransformed by esterases or peroxidase-catalases. The expected products are tetrazoles, a vinylogous carboxylic acid, and CH-acidic ketosulfoxides, isosteres of pyrazinoic and nicotinic acids, which should inhibit mycobacterial growth when released inside the bacterial cell. The growth inhibitory activity of the synthesized compounds against the H37Rv strain of Mycobacterium tuberculosis was determined to assess the viability of this concept. It was shown that all of the compounds designed as lipophilic precursors were more active than the unmodified polar isosteres of pyrazinoic and nicotinic acids.     

The role of sesquiterpene lactones in contact hypersensitivity to some North and South American species of feverfew (Parthenium-Compositae)

Sixteen taxa of the genus Parthenium from North and South America and 20 sesquiterpene lactones derived from plants of the Compositae family were used for patch testing six volunteers who were known to be contact sensitive to plants and lactones of this family. The patterns of cross-sensitivity and structure-activity relationships of the allergenic lactones are evaluated and the clinical and ecological relevance is discussed.     

A SINE species from hippopotamus and its distribution among animal species

Thirty sequences of a short interspersed repetitive element (SINE) were isolated from genomic DNA of Hippopotamus amphibius (hippopotamus). RNA polymerase III split promoter sequence was observed in all of the 30 sequences; and poly(A)-like structure at 3'-end, as well as direct repeat flanking to the repetitive sequence in many of the 30 sequences. A comparison of the consensus sequence of the 30 sequences with sequences in a DNA database (DDBJ/GENBANK/EMBL) revealed 93% homology to the consensus sequence of a whale SINE, CHR-2, and 73% homology to mouse glutamic acid tRNA. Phylogenetic analysis of tRNA-related regions of the sequences with all of the mouse tRNAs revealed that glutamic acid tRNA was genetically closest to the hippopotamus SINE. In addition, the tRNA-related region of the consensus sequence was folded into a cloverleaf structure as with mouse glutamic acid tRNA. These findings led us to conclude that the SINE of hippopotamus was genetically related to a whale SINE, CHR-2 [the hippopotamus SINE was named CHR-2(hippo)] and was a retroposon derived from glutamic acid tRNA. Hipo53 and hipo95, which were the genetically most separated CHR-2(hippo) sequences in the present study, were used as a probe for dot-blot hybridization to examine the distribution of their homologous sequences among animal species. Although the distribution spectra of hipo53 and hipo95 homologous sequences in animal species differed to some extent, large amounts of both sequences were found in Hippopotamus amphibius and Globicephala macrorhynchus (whale); and small amounts in most of the animal species in Artiodactyla examined. These findings indicated that the hippopotamus and whale had more recently branched off from the clade that includes chevrotain and pecorans than the other animal species in the clade. The 30 CHR-2(hippo) sequences were aligned, and the substitution rates among the sequences were calculated with a different substitution rate model for transition and for transversion. The calculation combined with the mutation rate of the pseudogenes (r = 4.6 x 10(9)) indicated that CHR-2(hippo) sequences diversified at least 132 million years ago (Myr).     

Allergenic sesquiterpene lactones from Eupatorium cannabinum L. and Kaunia rufescens (Lund ex de Candolle)

From Eupatorium cannabinum L., a hitherto unknown alpha-methylene-gamma-butyrolactone, 3 beta-peroxyeucannabinolide, was isolated. This compound and eupatoriopicrin from the same plant showed a weak sensitizing capacity in guinea pigs. 2-oxoludartin and dehydroleucodin, isolated from Kaunia rufescens (syn. Eupatorium rufescens), were strong sensitizers in the same sensitizarian procedure.     

Choline esters and biogenic amines in the hypobranchial gland of 55 molluscan species of the neogastropod Muricoidea superfamily

As many as 55 neogastropod molluscs, all belonging to the Muricoidea superfamily, have been investigated for occurrence and contents, in their hypobranchial gland (HG), of choline esters and, subordinately, biogenic amines. Very high amounts of esters, strictly localized in the median area of the HG, were found in all dye-secreting molluscs. The choline esters were represented by murexine, dihydromurexine and senecioylcholine. A fourth ester, acryloylcholine, occurred in the HG of a single, non dye-secreting mollusc. All the compounds displayed potent neuromuscular blocking actions in all examined vertebrate and invertebrate species, as well as potent nicotinic actions. Muscarinic effects were either lacking or unimportant. In addition to choline esters the HG occasionally contained known and hitherto unknown biogenic amines: tyramine, octopamine, 5-hydroxytryptamine, histamine, urocanylhistamine and imidazole-propionylhistamine. The interest of extending the search of bioactive compounds to carnivorous, predatory molluscs other than those described in this paper and, more, extensively, to any molluscan species provided with 'venomous' glands or apparatuses, is emphasized.     

Altered transfer of cholesteryl esters and phospholipids in plasma from alcohol abusers

The net mass transfer (NMT) of cholesteryl esters (CEs), triglycerides (TGs), and phospholipids (PLs) between lipoproteins was measured after incubation of fresh plasma for up to 2 hours from 18 male alcohol abusers and 17 male volunteer control subjects. In alcohol abusers the mean value of CE NMT was 3.7 nmol.mL-1.h-1 from apolipoprotein B-containing lipoproteins (apoB-containing lipoproteins) to HDL and in control subjects 8.7 nmol.mL-1.h-1 from HDL to apoB-containing lipoproteins. The NMT of PL was higher in alcohol abusers than in control subjects (35.0 vs 11.6 nmol.mL-1.h-1 from apoB-containing lipoproteins to HDL, respectively), and plasma PL transfer protein (TP) activity was 33% higher (P < .05) in alcohol abusers than in control subjects. The lack of correlation between the NMTs and CETP and PLTP activities suggests that the NMT could more closely reflect the role of lipoprotein properties in reverse cholesterol transport in vivo, whereas in vitro activities reflect the total capacity of transfer but not its direction. The rate of CE NMT from HDL to apoB-containing lipoproteins was dependent on the VLDL TG concentration. Moreover, at low VLDL TG levels, the increased HDL cholesterol concentration in alcohol abusers reversed the direction of CE NMT. This situation could be reconstructed in the plasma of control subjects by adding autologous HDL or VLDL to mimic the lipoprotein profiles of the alcohol abusers. Addition of VLDL enhanced the CE NMT from HDL to apoB-containing lipoproteins, whereas addition of HDL had an opposite effect, and at higher HDL levels, even reversed the direction of CE NMT. In conclusion, the NMT of CE and PL in alcohol abusers differs from that in control subjects. The concentrations of HDL and VLDL seem to be the major determinants of the direction of CE NMT in alcohol abusers.     

Sesquiterpene lactones. Part XV. New cytostatic active sesquiterpene lactone from herb of Anthemis nobilis L

From fresh and dried herb (without inflorescences) of Anthemis nobilis L. a new sesquiterpene lactone C20H26O6 was isolated. This compound showed a cytotoxic activity at a level of ED50 HeLa -- 0-56 microgram/ml (1-5 x 10(-6) M), and ED50 KB -- 1.23 microgram/ml (3-5 x 10(-6) M), what qualifies it for further in vivo studies.     

Synthesis and biological activity of lactones en route to Altohyrtin A

Lactones 2 and 7 were synthesised and tested against six human tumour cell lines (Pancreas-a, BXPC-3), (Thyroid ca, KAT-4), (Thyroid ca, SW1736), (Lung-NSC, NCI-H460), (Pharynx-sq, FADU) and (Prostate, DU-145). Lactone 7 proved inactive, but lactone 2 displayed some activity against four of the six cell lines examined. Both lactones were converted into an intermediate 5 en route to Altohyrtin A.     

Studies on long chain cis- and trans-acyl-CoA esters and Acyl-CoA dehydrogenase from rat heart mitochondria

A quantitative counterimmunoelectrophoresis technique has been applied to evaluation of RNA antibodies in serum and unconcentrated CSF. Serum anti-RNA antibodies have been found at high dilutions in some patients with infectious diseases, subacute panencephalitis and multiple sclerosis. RNA antibodies in CSF are significantly linked to oligoclonal aspect, i.e. to a synthesis of IgG inside the CNS.     

Novel site-specific endonucleases from Brevibacterium species

New site-specific endonucleases BecAI and BecAII have been detected in Brevibacterium species A. Endonuclease BecAII free from contaminating nonspecific endonucleases, exonucleases, and phosphatases was isolated by column chromatography on phosphocellulose, heparin sepharose, and DNA cellulose. It recognizes and cleaves the 5'-GG decreases CC-3' sequence and is a true isoschizomer of HaeIII restriction enzyme. The other restriction endonuclease, BecAI, cleaves Ad2 DNA at least by 2 sites but not the DNA of phage lambda, T7, SV40, phiX174, and plasmides pBR322 and pUC19. The substrate specificity of BecAI indicates its appurtenance to the super rare restriction endonucleases.     

Isolation and identification of Aeromonas species from human stools

One thousand and three diarrhoeal stool samples were processed in our laboratory during the period 1996/1997 for the presence of enteric pathogens especially Aeromonas spp., which has emerged as a new agent causing diarrhoea. Ampicillin sheep blood agar was found to be the best medium for the isolation of Aeromonas spp. from stool specimens. Enteric pathogens were found in 200 (20%) stools, of which Aeromonas spp. was the second commonest pathogen isolated amounting to 21% of isolates. This study clearly indicates that Aeromonas spp. must be looked for in every diarrhoeal stool samples, specially in children below 10 years of age. Isolation and identification is cost effective and easy, if the given protocol is observed.     

Clerodane diterpenoids, long chain esters of coumaric acid and other compounds from Baccharis myrsinites

PURPOSE: To compare 1-year survival and acute toxicity rates between an accelerated hyperfractionated (AH) radiotherapy (1.6 Gy b.i.d.) to a total dose of 54.4 Gy vs. an accelerated fractionation (AF) of 30 Gy in 10 daily fractions in patients with unresected brain metastasis. METHODS AND MATERIALS: The Radiation Therapy Oncology Group (RTOG) accrued 445 patients to a Phase III comparison of accelerated hyperfractionation vs. standard fractionation from 1991 through 1995. All patients had histologic proof of malignancy at the primary site. Brain metastasis were measurable by CT or MRI scan and all patients had a Karnofsky performance score (KPS) of at least 70 and a neurologic function classification of 1 or 2. For AH, 32 Gy in 20 fractions over 10 treatment days (1.6 Gy twice daily) was delivered to the whole brain. A boost of 22.4 Gy in 14 fractions was delivered to each lesion with a 2-cm margin. RESULTS: The average age in both groups was 60 years; nearly two-thirds of all patients had lung primaries. Of the 429 eligible and analyzable patients, the median survival time was 4.5 months in both arms. The 1-year survival rate was 19% in the AF arm vs. 16% in the AH arm. No difference in median or 1-year survival was observed among patients with solitary metastasis between treatment arms. Recursive partitioning analysis (RPA) classes have previously been identified and patients with a KPS of 70 or more, a controlled primary tumor, less than 65 years of age, and brain metastases only (RPA class I), had a 1-year survival of 35% in the AF arm vs. 25% in the AH arm (p = 0.95). In a multivariate model, only age, KPS, extent of metastatic disease (intracranial metastases only vs. intra- and extracranial metastases), and status of primary (controlled vs. uncontrolled) were statistically significant (at p < 0.05). Treatment assignment was not statistically significant. Overall Grade III or IV toxicity was equivalent in both arms, and one fatal toxicity at 44 days secondary to cerebral edema was seen in the AH arm. CONCLUSION: Although a previous RTOG Phase I/II report had suggested a potential benefit in patients with limited metastatic disease, a good Karnofsky performance status, or neurologic function when treated with an AH regimen, this randomized comparison could not demonstrate any improvement in survival when compared to a conventional regimen of 30 Gy in 10 fractions. Therefore, this accelerated hyperfractionated regimen to 54.4 Gy cannot be recommended for patients with intracranial metastatic disease.     

Fatty acid ethyl esters and HepG2 cells: intracellular synthesis and release from the cells

Fatty acid ethyl esters (FAEE), esterification products of fatty acid and ethanol, have been implicated as mediators of ethanol-induced organ damage. To understand the molecular and cellular events in FAEE synthesis and secretion, we developed a system in which HepG2 cells synthesize and release FAEE into the culture medium upon incubation with ethanol. The synthesis of FAEE was observed within 5 min of the addition of ethanol, with a plateau for FAEE synthesis after 2 h of incubation. It was also observed that FAEE are synthesized by both a microsomal FAEE synthase, which preferentially uses fatty acyl-CoA as a substrate, and a cytosolic FAEE synthase, which accepts both unesterified fatty acid and fatty acyl-CoA as substrates with a slight preference for fatty acyl-CoA. Although the kinetics of cellular FAEE synthesis await further characterization, the intracellular fatty acid substrate appears to be derived principally from glycerolipids and other esters. FAEE were released into the culture medium by a mechanism independent of the vesicular transport pathway. Lipoprotein particles and albumin were found to be carriers of FAEE after FAEE secretion from the cell.     

Novel Psychrobacter species from Antarctic ornithogenic soils

Ornithogenic soil is derived from the deposition of the fecal matter of various species of birds and is a major source of nutrient input in the Antarctic marine ecosystem. A significant proportion of microbiota of ornithogenic soil collected from an Adélie penguin colony in eastern Antarctica (Vestfold Hills ice-free zone) consisted of gram-negative, coccoid bacteria identified on the basis of their phospholipid ester-linked fatty acid and lipid class profiles as Psychrobacter strains. Phenotypic, genotypic, and 16S ribosomal DNA phylogenetic analyses revealed that the Antarctic psychrobacters belonged to three distinct groups. Comparisons with Psychrobacter immobilis and Moraxella phenylpyruvica reference cultures isolated from fish, seawater, poultry, and human clinical specimens revealed the relationships of these groups within the genus Psychrobacter. Two of the groups represent the following two novel species: Psychrobacter urativorans sp. nov. (type strain, strain ACAM 534) and Psychrobacter frigidicola sp. nov. (type strain, strain ACAM 304). The third group of strains included members of the previously described species P. immobilis (Juni and Heym 1986). In addition, M. phenylpyruvica (B?vre and Henriksen 1967) is renamed Psychrobacter phenylpyruvicus comb. nov. (type strain, strain ACAM 535) on the basis of 16S ribosomal DNA phylogenetic data. In general, the genus Psychrobacter could be differentiated from the related genera Moraxella and Acinetobacter by the fact that the members of the genus Psychrobacter are psychrotolerant or psychrophilic and halotolerant, which reflects the ubiquitous distribution of the genus in both marine and terrestrial environments. On the basis of the results of this and previous studies, the genus Psychrobacter is the predominant genus in ornithogenic soils in Antarctica and is diverse.     

Effects of macrocyclic lactones on ingestion in susceptible and resistant Haemonchus contortus larvae

The effects of ivermectin and ivermectin aglycone on pharyngeal uptake of a carbohydrate substrate (3H-inulin) were measured in larvae of a macrocyclic lactone (ML)-susceptible isolate and 2 ML-resistant isolates of Haemonchus contortus. The resistant isolates showed a tolerance (in terms of the concentration of compound required to reduce feeding to 50%) toward ivermectin of approximately 4.5- and 9-fold and toward ivermectin aglycone of approximately 14-fold, compared to the susceptible isolate. This indicates that susceptible and resistant isolates can be readily distinguished on the basis of the sensitivity of pharyngeal uptake to MLs. The use of various metabolic inhibitors in this assay system did not reveal the nature of the resistance mechanism. Pretreatment of resistant larvae with inhibitors of multidrug resistance mechanisms (P-glycoprotein and multidrug resistance protein) and detoxification enzymes (monooxygenases, esterases, and glutathione transferases) did not reduce their level of tolerance to the ivermectin aglycone.     

A simple method for extracting crude sesquiterpene lactones from Compositae plants for skin tests, chemical investigations and sensitizing experiments in guinea pigs

The increased populatiry of surfing has produced a marked augmentation in the incidence of ear canal exostosis. However, when it becomes moderately severe, I prefer to call it "hyperostosis." Exposure to cold ocean water for many years can be an important etiologic factor in hyperostosis. There is a serious risk, and a high incidence of tympanic membrane perforations during the removal of large external canal hyperostosis. This injury can be prevented by placing a sheet of Silastic against the tympanic membrane beforehand. I describe the method. Serious degrees of hyperostosis, causing transient hearing loss and otitis externa, are increasingly common in coastal towns, where cold-water surfing is a popular year-around sport.     

Relevance of culturing Candida species from intravascular catheters

The significance of isolating Candida species from intravascular catheters was examined retrospectively. Our findings demonstrate that these organisms are encountered among patients with proven invasive candidiasis or as an isolated finding. Febrile patients with yeasts in two or more additional sites may be at risk for invasive cadidiasis and may benefit from treatment.