Solid-phase supported synthesis: a possibility for rapid scale-up of chemical reactions.

The direct scale-up of a solid-phase synthesis has been demonstrated with 4-(2-amino-6-phenylpyrimidin-4-yl)benzamide and an arylsulfonamido-substituted hydroxamic acid derivative as examples. These compounds were obtained through combinatorial chemistry and solution-phase synthesis was used in parallel to provide a comparison. By applying highly loaded polystyrene-derived resins as the solid support, a good ratio between the product and the starting resin is achieved. We have demonstrated that the synthesis can be scaled up directly on the solid support, successfully providing the desired compounds easily and quickly in sufficient quantities for early development demands.

Application of coil centrifugal counter-current chromatography to the separation of macrolide antibiotic analogues III. Effects of flow-rate, mass load and rotation speed on the peak resolution.

As the third part of our methodology studies on the application of centrifugal counter-current chromatography to the preparative separation of macrolide antibiotic analogues, we have investigated the effects of various parameters on the retention of stationary phase and peak resolution. Our results show that the retention percentage of the stationary phase has linear relationships with both flow-rate at 1 to 3 ml/min and rotation speed at 100 to 700 rpm, but their correlation coefficients are negative (-1.000) and positive (0.9821), respectively. The peak resolution (Rs) is inversely proportional to the flow-rate (Fr) and mass load (Ml), but directly proportional to the rotation speed (Rrev). Their correlation coefficients in linear regression for the preparative separation in laboratory scale are -0.981 to -1.000 for Rs=a+bFr at flow-rates of 1 to 3 ml/min, -0.929 to -0.993 for Rs=a+bMl at mass loads of 12.5 to 100 mg, and 0.975 to 0.998 for Rs=a+bRrev at rotation speeds of 300 to 700 rpm, respectively. Preparative separation of six very closely related macrolide antibiotics, which belong to ascomycin and rapamycin analogues, has also been successfully achieved under optimized conditions.

Application of centrifugal counter-current chromatography to the separation of macrolide antibiotic analogues. I. Selection of solvent systems based on solubility and partition coefficient investigations.

As the first part of our studies on counter-current chromatography, the methodology for selecting suitable solvent systems was established based on detailed investigations of solubility and partition coefficients (log K) of macrolide antibiotic analogues. The solubility of two important macrolides, ascomycin and FK-506, was measured in a series of common solvents, where their polarities were ranked with dielectric constants. The partition coefficients of the two macrolides were compared in various binary, ternary, quaternary solvent systems. Hexane-tert.-butyl methyl ether-methanol-water system was selected based on suitable log K of solutes and hydrogen-bonding properties of solvents. In the further optimisation of composition proportions in the multicomponent solvent system, hexane-tert.-butyl methyl ether-methanol-water (1:3:6:5) showed the best solvent selectivity by giving the most prominent difference of partition coefficient (delta log K) between ascomycin and FK-506. With this solvent system, a baseline preparative separation of these two very closely related 23-membered macrolide antibiotics was successfully achieved by employing the newly introduced Quattro counter-current chromatograph.

Follow-up of skin manifestations in Yu-Cheng children.

We report an 11-year follow-up study on the dermatological manifestations of Yu-Cheng children born to mothers who were poisoned by polychlorinated biphenyl (PCB)-contaminated cooking oil between 1977 and 1979. Eighty-eight children born to these mothers, and 86 matched controls, were examined by one dermatologist. Chloracne scars were found in one patient, and there were nail abnormalities in about one-third of the exposed patients. Transverse grooves, irregular depressions, and koilonychia/nail flattening were significantly more frequent than in the control group, based on Fischer's exact test. This study indicates that in Yu-Cheng children the nail changes are the most persistent abnormality after PCB intoxication, and their occurrence may indicate developmental retardation of the fetal nail matrix. Such a finding might also suggest that PCBs remaining in the mother could exert an effect on nail growth in children born several years after the intoxication event.

Extrathoracic sarcoidosis in a Chinese man presenting with multiple, large plaques and tumors.

We report an unusual case of tumorous sarcoidosis in a 42-year-old Chinese man with extrathoracic sarcoidosis. He presented with multiple large plaques and tumors on the neck and extremities of a nine-year duration. Because of the atypicality of the cutaneous manifestation, the lack of intrathoracic lesions and the rarity of sarcoidosis in Taiwan, the case remained undiagnosed for nine years despite numerous examinations and tests. A subsequent skin biopsy revealed sarcoidal granulomas. The detection of cytoplasmic birefringent inclusions in the granulomas led to the final diagnosis. Originally thought to be foreign bodies, the inclusions were proven to be calcium oxalate by x-ray microanalysis and calcium oxalate stain. Lymph node and liver biopsies confirmed the diagnosis. The skin lesions in this case may have been precipitated by frequent harsh scrubbing to clean the skin. Sarcoidosis should be considered in the differential diagnosis in patients presenting with multiple cutaneous tumors. This case also illustrates that detection of calcium oxalate inclusions in granulomas, while not specific, can serve as a valuable clue in the diagnosis of cutaneous sarcoidosis.

Effects of retinoids on endothelial cell proliferation, prostacyclin production and platelet aggregation.

In addition to their anti-inflammatory and anti-cancer activities, retinoids have been shown to affect angiogenesis, endothelial proliferation and the process of wound healing. While peripheral vascular occlusion has not been observed as an adverse effect clinically, the effects of retinoids on prostacyclin production in endothelial cells and platelet aggregation are not known. We examined the effects of tretinoin, isotretinoin and etretinate (3.3 x 10(-8) to 3.3 x 10(-5) M) on cytotoxicity by 51Cr-release assay, growth and prostacyclin in bovine carotid endothelial cell cultures, and the aggregation of human platelets induced by ADP. All retinoids showed either no or only small effects on cytotoxicity and human platelet aggregation. Prostacyclin production was not significantly affected except for tretinoin and isotretinoin at 3.3 x 10(-5) M. Endothelial proliferation was affected by all three retinoids in a dose-dependent fashion; for tretinoin and isotretinoin an inhibitory trend was noted as the concentration increased but the reverse was true for etretinate. Retinoids at 3.3 x 10(-5) M induced alterations of typical endothelial morphology; the cells became fibroblastoid. The results of prostacyclin production and platelet aggregation in the present study are consistent with the absence of peripheral vascular occlusion as a side effect clinically.

Quantitative and morphological changes of Langerhans cells in Bowen's disease from patients with chronic arsenicism.

Langerhans cells (LCs) are considered to be responsible for the immunologic presentation of tumor-associated antigens and play a role in the elimination of neoplastic clones. Ultraviolet light B can cause dysfunction and loss of LCs. Both the number and dendritic morphology of LCs are known to be diminished in squamous cell carcinomas from sun-exposed skin. The effects of arsenics on LCs are unknown. Using an OKT-6 monoclonal antibody to stain intraepithelial LCs, we compared their number and morphology in Bowen's lesions and in the perilesional skin from sun-protected sites in ten patients with chronic arsenicism. There was a significant reduction in the numbers of LCs in the Bowen's lesions as compared to the perilesional skin specimens. Loss of dendrites was observed in all Bowen's lesions and in seven of the perilesional skin specimens. Ultrastructurally, the LCs showed an absence of dendrites, but the Birbeck granules were preserved. Since the specimens were not from sun-exposed skin in our study, the findings may be related to chronic arsenic intoxication. The morphologic alteration of LCs observed in the perilesional skin further suggests an arsenic-related systemic dysfunction of the LCs, which in turn may contribute to the development of skin cancers in these patients.

Study of peripheral T-cell subpopulations and lymphocyte proliferation in different phases of evolution of alopecia areata.

The current hypothesis on the pathogenesis of alopecia areata is immunologically mediated, yet no conclusive data has been documented. To clarify the changes in various phases of disease evolution, analysis of T lymphocyte subpopulations was carried out in the peripheral blood of 34 patients and 27 healthy normal subjects by means of OKT-3, OKT-4, OKT-8 and Ia1 monoclonal antibodies. Study of lymphocyte proliferation in response to various mitogens was further carried out in 27 patients and 25 healthy normal subjects. Significant predominance (p less than 0.01) of OKT-8 positive (cytotoxic/suppressor) cells and diminution (p less than 0.01) of Ia1 positive cells were noted in the whole alopecia group and in various subgroups of disease evolution. Since diminution (p less than 0.05) of OKT-4 positive (inducer/helper) cells was also noted in the stable phase, it was postulated that these activated T cells migrated from circulation to the local site of primary event. Hence, it was evident that the predominance of cytotoxic/suppressor cells was a relative change, and the role of inducer/helper cells in the pathogenesis was implicated. No enhancement of lymphocyte proliferation was observed.

Synthesis and beta-lactamase inhibitory activities of some clavulanic acid analogues.

Clavulanic acid analogs lacking the C-3 carboxyl group are potent inhibitors of both plasmid and chromosomally mediated beta-lactamases. They exhibit only low intrinsic anti-bacterial activity, but potentiate the activity of ampicillin and cephaloridine against beta-lactamase producing Escherichia coli and Enterobacter cloacae in vitro. No synergism was observed in beta-lactamase negative strains. The E. coli TEM 1 and the E. cloacae P99 enzymes are inhibited in a progressive and irreversible manner by these compounds.

(+/-)-2-Depentylperhydrohistrionicotoxin: a new probe for a regulatory site on the nicotinic acetylcholine receptor-channel.

(+/-)-2-Depentylperhydrohistrionicotoxin (4), several of its analogues, and N- and O-substituted derivatives were prepared and tested for their effects on the neuromuscular transmission of the frog sartorius muscle. Compound 4, its N-methyl derivative 5, the O-acetyl derivative 9, and the quaternary methiodides 19 and 20 blocked the indirectly elicited twitch. The oxidation of 4 and 5 to ketones 12 and 14 and their reduction to the epimeric alcohols 17 and 18 afforded materials with substantially reduced activity. N-Acetylation of 4 to 11 changed the course of the activity to a transient potentiation of muscle twitch. Both 4 and 5 were not very toxic to mice after subcutaneous administration. (+/-)-7-n-Butyl-1-azaspiro[5,5]undecan-8-one (12) epimerized readily at room temperature to afford the epimer 13, and preparation of the hydrochloride of its N-methylated derivative 14 was accompanied by a retro-Michael reaction, affording the 2-n-butyl-3-[4-(methylamino)butyl]cyclohexene-2-one (22). The strongly hydrogen-bonded alcohol 4 was analyzed as the hydrobromide by a single-crystal X-ray analysis, confirming its structure.