The synthesis of chiral synthons by means of microbial and enzymatic function and its application to the synthesis of complex, bioactive substances, the aglycone of venturicidins A and B are reviewed. 1) The syntheses of chiral synthons having two chiral centers are described based on the microbial asymmetric reduction of α-methyl β-keto esters or enzymatic asymmetric hydrolysis of α-methyl β-acetoxy esters. 2) The new chiral synthons synthesized in 1) have been successfully applied for the total syntheses of oudemansins A and B. 3) Purification and properties of the asymmetric reduction enzyme of α-methyl β-keto esters in Saccharomyces cerevisiae or Saccharomyces fermentati were investigated. 4) Formal total synthesis of (-)-indolmycin is described based on the asymmetric hydrolysis of α-acetoxy ester with lipases. 5) Enzymatic hydrolysis in organic solvents for kinetic resolution of water-insoluble α-acyloxy esters with immobilized lipases is described. 6) The first total synthesis of the aglycone of venturicidins A and B has been successfully achieved from the new chiral synthon based on the developed method in 1).
Separation and determination of bile acids in biological fluids by high-performance liquid chromatography are reviewed. The capacity ratios of bile acids on an ODS column were affected by the number, position and configuration of the hydroxyl group on the steroid nucleus, and the chromatographic behavior was markedly influenced by the pH of a mobile phase according to the conjugated form at C-24. A new pre-column derivatization reagent, 1-anthroyl nitrile, was developed and applied to the analysis of bile acids in biological fluids. Bile acids were derivatized through the 3α-hydroxyl group into the corresponding esters, separated on an ODS column, and monitored by a fluorescence detector with detection limit of 20 fmol. The sensitive method for the determination of bile acids in biological materials by gas chromatography (GC) in combination with negative ion chemical ionization (NICI) mass spectrometry is also described. Of various derivatives for the carboxyl group, the pentafluorobenzyl (PFB) ester provided the highest value of the ratio of the negative to positive ion current. A characteristic carboxylate anion [M-181]- was produced as the most abundant ion by the loss of the PFB group in NICI. PFB esters of bile acids were further derivatized into the dimethylethylsilyl ethers and then separated by GC. The detection limit was 2 fg when the characteristic anion was monitored in the NICI mode. The preparation of 18O-labelled bile acids, as the internal standard for the trace analysis or the tracer for the metabolic study, was developed. Finally, the clean-up procedure for bile acids in biological fluids was investigated. The combined use of solid-phase extraction with a Sep-pak C18 or Bond Elut cartridge and group separation on a lipophilic ion-exchange gel, piperidinohydroxypropyl Sephadex LH-20, was found most effective for this purpose.
From the leaves of Melicope triphylla MERR., three new flavonoids (1-3) were isolated, together with nine known flavonoids, 4', 5-dihydroxy-3, 3', 7-trimethoxyflavone (4), 5-hydroxy-3, 3', 4', 7-tetramethoxyflavone (5), 3, 3', 4', 5, 7-pentamethoxyflavone (6), 7-hydroxy-3, 3', 4', 5, 8-pentamethoxyflavone (7), 3, 3', 4', 5, 7, 8-hexamethoxyflavone (8), 5-hydroxy-3, 7, 8-trimethoxy-3', 4'-methylenedioxyflavone (9), 7-hydroxy-3, 5, 8-trimethoxy-3', 4'-methylenedioxyflavone (10), 3, 5, 7, 8-tetramethoxy-3', 4'-methylenedioxyflavone (11), and 5-hydroxy-3, 6, 7, 8-tetramethoxy-3', 4'-methylenedioxyflavone (12). The structures of 1, 2, and 3 were established as 5-hydroxy-3, 7-dimethoxy-3', 4'-methylenedioxyflavone, 5-hydroxy-7-isopentenyloxy-3, 8-dimethoxy-3', 4'-methylenedioxyflavone, and 4'-hydroxy-7-isopentenyloxy-3, 3', 5, 8-tetramethoxyflavone by their respective chemical and spectral data. The 20 flavonoids isolated from this plant were examined for the piscicidal activities.
For the purpose of clarifing the factors which affect the growth of Bupleurum falcatum L., the appearance of Bupleurum roots cultivated under 5 kinds of soils with or without ventilation and watering in the soil was compared. Main and hair roots grew up well in sandy loam soil, whereas main and lateral roots grew up in clay loam soil. The content of saikosaponins in the root was shown to be highest in stream or mountain sand. The number of blooming increased and time to flowering came earlier with soil ventilation. Gaseous phase in the three phase distribution of soil increased with ventilation or watering. The yield of Bupleurum root also increased with soil ventilation or watering in all kinds of soils. As a result, it is concluded that the soil ventilation is a useful method for the cultivation of B. falcatum owing to the change in the physical properties of soils.
During our screening of pharmacologically active principles from Indonesian medicinal plants by a hypothermic effect the methanol-extract of Brucea javanica (L.) MERR. has been found to exhibit a lethal toxicity to mice. The toxic components were isolated and identified with bruceoside A and B and yadanzioside F. Correction of 18C-nuclear magnetic resonance assignment was also mentioned.
Plasmid pKYM is a multicopy plasmid isolated from Shigella sonnei and multiplies stably in Escherichia coli. The plasmid encodes Rep protein which is essential for its multiplication and synthesizes cop ribonucleic acid (RNA) which is a short RNA complementary to the 5' region of rep m-RNA. This RNA controls the copy number and the incompatibility of the plasmid. The previous analysis located the promoters of rep m-RNA (PR) and cop RNA (PL) in the inc region. This report confirmed the presence of these promoters by analyzing the RNAs isolated from the cells carrying pKYM and those synthesized in vitro. The initiation sites of these transcriptions were also determined. Analysis of in vivo RNA suggested that the quantity of cop RNA whose size was about 90 nucleotides was larger than that of rep m-RNA and these RNAs easily formed RNA-RNA hybrid. The analysis also suggested that the synthesis of rep m-RNA was repressed by cop RNA and Rep protein itself.
A method of seeding and subsequent freeze-drying of aqueous cephalothin sodium (CET-Na) of supersaturated concentrations was employed in an experiment with a regular production scale. The freeze-dried plugs of non-uniformity in appearance were consequently obtained together with those of white in color and in high uniformity in appearance. Such plugs of irregular shape have been found to show a marked color development during storage, and the reconstitution time was also prolonged. Scanning electron microscopy, powder X-ray diffractometry, and thermogravimetry have all demonstrated that the plugs of irregular appearance show dense aggregates of fine crystals of a high non-crystallinity by an incomplete crystal growth. Investigation has then been made with supersaturated CET-Na solution, which has preliminarily received varying mechanical stresses in varying time courses of temperature, similarly to a regular production program, for the purpouse of achieving the reproducibility of such unacceptable plugs. It has been demonstrated that by a more prolonged storage at a lower temperature of CET-Na in supersaturated solution the plug may surely contain aggregates of more fine crystallites of a higher non-crystallinity, accordingly have a higher irregularity in appearance and exhibit a more marked deterioration in pharmaceutical qualities.
Seeding and subsequent freeze-drying of aqueous cephalothin sodium (CET-Na) solution of supersaturated concentrations, which preliminarily received an excess of mechanical stress, have been reported to make the product unacceptable in pharmaceutical elegance and in chemical stability upon storage. In order to find out the cause, the CET-Na solution that preliminarily received mechanical stress has been examined as to its crystallization behavior during freezedrying process, by using polarizedlight cryomicroscopy, scanning electron microscopy, cryodifferential scanning calorimetry, refractometry, viscositometry, and surface tensiometry. Aqueous CET-Na solution of supersaturated concentrations, to which an excess of mechanical stress was given, has been demonstrated to exhibit the following features : in the process of seeding and subsequent freeze-drying, the solution shows an incomplete crystallization at -40°C ; in the crystal growth stage at -5°C, crystals are found to be difficult to grow sufficiently from the seeds as their nuclei. These findings indicate that the supersaturated solution has changed to an unusual state upon receiving such stress, the state being different from the state of untreated solution in that the solution thus treated shows physico-chemical properties capable of easy crystallization through a simple step of freezing. The change as such in physico-chemical properties has also been estimated to have resulted from a structural change of associated molecules of CET-Na in its supersaturated solution into a condensed state quite similarly to that of liquid crystals.
Degradation and excretion of Sizofiran (SPG), an anti-tumor polysaccharide, were studied in rats after a single or multiple administration. After a single intravenous injection of [14C] SPG (3 mg/kg), SPG distributed in the liver was degradated at very slow rate to SPG-like substances (SPGLS) having lower molecular weight than that of SPG, while SPG in the spleen and mesenteric lymph node was metabolized at much slower rate than that in the liver. In the experiment with multiple subcutaneous administration, SPG was also found to be present mainly as SPGLS in the liver, but almost as an unchanged SPG in the spleen. SPG was excreted in the urine mainly as metabolites with a molecular weight of less than 10000. These results indicate that degradation of SPG to lower molecular weight-SPGLS is a prerequisite for efficient urinary excretion and the degradation occurs mainly in the liver.
Protective effects of Aloe arborescens (AA) on mouse skin injury induced by soft X-irradiation were examined. The mechanisms on radiation protection by measuring scavenge activity of activated oxygen, protective effects of nucleic acid, induction of antioxidative protein and so on were further investigated. Consequently a significant protective effect of skin injury was observed in AA S6-3-b. As the mechanisms of radiation protection in AA, the following matters were found. AA S6-3-b showed scavenge activity of hydroxyl radicals generated by Haber-Weiss reaction. AA S6-3-b suppressed the changes of activity in superoxide dismutase and gluthathione peroxidase at 7d after soft X-irradiation. Metallothionein was induced in the skin and liver against normal mice at 24 h after administration of AA S6-3-b.
This study is to investigate radioprotective effects of 23 Taiwan crude drugs on X-ray induced bone marrow death and skin injury in mice. Each methanol extract of these Taiwan crude drugs was injected intraperitoneally into ICR male mice at 6 weeks of age before irradiation. Mice were whole-body irradiated with a soft X-ray generator. Radiation factors of the two screening tests used were as follows : 70 kVp, 10 mA, 10 mm acrylate filter, 70R/min, 2100R for survival test, and 30 kVp, 10 mA, 190R/min, 1100R for protective test on skin injury. As a result of these studies, the survival effect was recognized in Solani Incani Herba and Orthosiphi Aristati Herba. On the other hand, Mimosae Herba, Canarii Radix, Bombacis Radix, Arecae Fructus, Hedyotidis Diffusae Herba and Cynomorii Caulis were shown to have significant protective potency on skin injury.