Detection of prostaglandins by high-performance liquid chromatography after conversion to p-(9-anthroyloxy)phenacyl esters.

p-(9-Anthroyloxy)phenacyl bromide (panacyl bromide) undergoes rapid reaction with the carboxyl group of prostaglandins in the presence of N,N-diisopropylethylamine in acetonitrile-tetrahydrofuran (4:1). The resulting prostaglandin panacyl esters are strongly uv absorbing with a lambda max at 253 nm and an epsilon of 174,280 in acetonitrile. The lower limit of detection of prostaglandins was approximately 200 pg with uv detection (254 nm) and about 30 pg with fluorescent detection (exitation 253 and emission 445 nm) using normal-phase HPLC. The reactivity of panacyl bromide with 23 prostaglandins as well as prostaglandins released by human lung tissues was investigated.

Physical model approach to gastrointestinal absorption of prostaglandins II: In situ rat intestinal absorption of dinoprost.

In situ absorption studies with dinoprost in the rat jejunum were carried out using a modified Doluisio technique. The absorption rate was first order. There was a sigmoidal decrease in the rate with increasing buffer pH (from 3.5 to 9.5), which strongly indicated the partitioning of weak acid species into the lipoidal membrane. An asymptotic minimum rate was attained from buffer pH 7.5 to 9.5, operationally indicative of transport of anions across aqueous pores. The importance of the aqueous diffusion layer on the mucosal side of the membrane was evident; rates at pH 3.5 and 4.5 were faster at high agitation hydrodynamics in the lumen solution. Preliminary studies showed that there was no metabolism in the lumenal solution and that metabolism occurred within the membrane. The transport mechanism involved simultaneous passive diffusion and bioconversion in the membrane because (a) a 1.5 X 10(4)-fold range in dinoprost concentration (0.014-210 microM) showed no saturable carrier-mediated tendency on the rate, (b) iodoacetic acid and indomethacin did not inhibit the absorption rate, and (c) the shape of the absorption-pH profiles was suggestive of passive diffusion. The prostaglandin did not have apparent adverse membrane and vascular effects under the conditions employed. The quantification and factorization of the physically meaningful transport parameters were accomplished using the physical model previously described. The permeability coefficients of the aqueous diffusion layer for the oscillation and static hydrodynamic situations were 0.8 X 10(-4) and 1.7 X 10(-4) cm/s, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Physical model approach to gastrointestinal absorption of prostaglandins III: In situ rat intestinal absorption of dinoprostone.

In situ absorption studies with dinoprostone in the rat jejunum were carried out to provide a quantitative mechanistic insight of the absorption process. The variables included buffer pH (3.5-9.5), buffer capacity, hydro-dynamics in the lumen, and concentration. The disappearance kinetics from the lumen was first order. The rate decreased with increasing pH in a sigmoidal manner and reached a minimum at about pH 9. These results indicate the effects of the partitioning of nondissociated species in the lipoidal membrane and transport across aqueous pores. The rate was higher with the higher degree of agitation of the lumenal solution. Between two hydrodynamic situations, the differences in the rates were large at pH 4.5 where the transport was largely aqueous diffusion controlled and then tended to become smaller with increasing pH where the transport became effectively membrane controlled. The 15-oxo- and 13,14-dihydro-15-oxo metabolites of dinoprostone were found. The physical model was applied to quantify the permeability coefficients of the aqueous diffusion layer and the aqueous pores of the membrane and the effective membrane transport-bioconversion permeability coefficient at various pH values. The overall absorption dinoprostone was similar to that of the less lipophilic dinoprost reported earlier and also more rapid. Hence, baseline absorption studies were completed with two major reference prostaglandins from which estimations of intestinal absorption can be made for their analogues and derivatives.

Prostaglandin prodrugs III: Synthesis and biological properties of C9- and C15-monoesters of dinoprost (prostaglandin F2 alpha).

Methods are described for the synthesis of dinoprost C9- and C15-monoesters using protective groups. Esters at C9 were synthesized by acylation of dinoprost 11,15-bis(tetrahydropyran-2-yl)ether followed by acid-catalyzed protective group removal. Esters at C15 were synthesized by initial formation of the protected intermediate, dinoprost 9,11-n-butylboronate, followed by acylation and hydrolytic protective group removal. Many esters were active in vivo in the hamster antifertility screen. Plasma hydrolysis studies showed that the C15-esters were more readily cleaved than the C9-esters. In vivo studies in the rat showed that both the C9- and C15-esters resulted in urinary excretion of 5 alpha, 7 alpha-dihydroxy-11-ketotetranorprosta-1,16-dioic acid in amounts comparable to those obtained after dosing with dinoprost, indicating that ester hydrolysis occurred in vivo.

Prostaglandin prodrugs. I: Stabilization of dinoprostone (prostaglandin E2) in solid state through formation of crystalline C1-phenyl esters.

Dinoprostone para-substituted phenyl esters were synthesized in attempt to improve the solid-state stability of the parent prostaglandin. A phenol series covering a wide melting-point range was employed, and a linear relationship was observed between the phenol melting points and the resulting prostaglandin C1-ester melting points. The crystalline esters showed improved solid-state stability over the parent compound, and many esters were biologically active.

Prostaglandin prodrugs. II: New method for synthesizing prostaglandin C1-aliphatic esters.

A new method for synthesizing C1-aliphatic esters of dinoprost and dinoprostone without using hydroxyl protective groups is described. Reaction of the prostaglandin with an alkyl halide in the presence of the sterically hindered amine N,N-diisopropylethylamine proceeds smoothly to give C1-esters in various solvents at ambient or slightly elevated temperatures. Polar solvents were strongly catalytic, and even the hindered tert-butyl esters were synthesized by employing solvents such as dimethylformamide or dimethyl sulfoxide. Biological evaluation in the hamster antifertility assay showed that some esters maintained high bioactivity.

Resolution of prostaglandin p-nitrophenacyl esters by liquid chromatography and conditions for rapid, quantitiative p-nitrophenacylation.

The p-nitrophenacyl esters of a number of closely related and isomeric prostaglandins were resolved by HPLC on a microparticulate silica gel column (Zorbax-Sil, DuPont). Ten F-series prostaglandin analogs, eight E-series prostaglandin analogs, the isomeric 15(R)- and 15(S)-methyl prostaglandins of the E- and F-series and, lastly, PGA2 and PGB2 were chromatographed under conditions generating 2,000 to 7,000 theoretical plates. Conditions are described for quantitative conversion of prostaglandins to p-nitrophenacyl esters in less than 6 minutes at room temperature. Linear peak height and peak area plots were obtained for in-situ esterified PGE2 p-nitrophenacyl ester over the range of 0.4 - 3.1 mug. The lower limit of detection of this ester is about 1 ng. A linear relationship is observed between silica gel TLC 1/Rf values and HPLC retention times as predicted by theory.

Liquid chromatography of clindamycin 2-phosphate on triethylaminoethyl cellulose.

The separation of clindamycin 2-phosphate from clindamycin 3-phosphate, clindamycin 4-phosphate, clindamycin B 2-phosphate, and lincomycin 2-phosphate was achieved by liquid chromatography on triethylaminoethyl cellulose using a 254-nm monitor. The compounds have low molar absorptivities at 254 nm (smaller than 17), and UV detection is made possible by the high capacity support triethylaminoethyl cellulose. Linear peak height response versus concentration allows rapid quantitation of clindamycin 2-phosphate.

Separation of prostaglandin A2 and prostaglandin B2 by ion-exchange liquid chromatography.

PIP: 2 closely related prostaglandins (PGs), A2 and B2, were separated by ion-exchange liquid chromatography. PGA2 and PGB2 are an isomeric pair of PGs which show little resolution on thin-layer chromatography; the pair can be resolved as gas-liquid chromatography, but use of this method requires protection of the C9-carbonyl group. Ion-exchange liquid chromatography, in contrast, requires no protective derivatization to achieve separation of PGA2 and PGB2 from PGE2. 2 different column types were used, and their results compared favorably. In 1 experiment, a triethylaminoethyl cellulose column resolved the 2 species; in another experiment, a strong anion-exchange pellicular support was successful. Both columns were stable and could be used to monitor the stability of PGE2 by following the appearance of PGA2 and PGB2, using the peak height method (i.e., a plot of micrograms injected vs. peak height). Triethylaminoethyl cellulose gave somewhat better resolution of PGA2 and PGB2 as compared with the pellicular strong anion-exchange column, but the advantage of this effect is offset by the tediousness of packing columns of triethylaminoethyl cellulose.^ieng

Prostaglandins.

PIP: Prostaglandins are analogs of the parent 20 carbon prostanoic acid. They are divided into 4 series: PGE; PGF; PGA; PGB, according to the presence of functionalities in the cyclopentane structure. Biosynthesis of prostaglandins were first independently reported by Bergstrom et.al. and van Dorp et.al. who showed that certain w-6-unsaturated fatty acids were biological precursors of prostaglandins. Later, various investigators reported the biosynthesis of prostaglandins of the different series. The 2 most widely used routes of prostaglandins synthesis are 1) the Corey cyclopentyl-lactone route, and 2) the bicyclohexane route. The synthesis is divided into 1) naturally occuring primary prostaglandins and 2) the prostaglandin analogs and derived prostaglandins. Because of the general instability of natural prostaglandins in the basic and acidic milieu, various preparations of prostaglandins and chemically stable dosage forms have been developed. Various methods used in analyzing prostaglandins include: 1) TLC; 2) GLC; 3) spectral methods; 4) radioimmunoassay; and 5) enzymatic assay. In vitro and in vivo studies on the metabolism and catabolism of various prostaglandins showed that they are rapidly metabolized in various animal systems and humans. The major routes for this metabolic transformation are: 1) oxidation of the secondary C15 hydroxy group; 2) reduction of the C13 double bond; 3) B-oxidation; 4) w-hydroxylation; and 5) w-oxidation. Prostaglandins produce a wide range of biological effects on animal and human systems (the reproductive; gastrointestinal; respiratory; and cardiovascular systems). The biological actions of prostaglandins on animal and human reproductive tissue vary depending on the particular prostaglandin studied and hormonal state of the tissue. Certain prostaglandins can decrease the tonus, frequency, and amplitude of spontaneous contractions of human uterine strips while other prostaglandins can cause contraction of isolated myometrial strips. Prostaglandins are widely used in labor induction; induction of therapeutic abortion; and fertility control. Prostaglandins have also been found to either increase or decrease cyclic AMP; inhibit ADP-induced platelet aggregation; lower blood pressure in animals; affect lipid and carbohydrate metabolism, and prevent adjuvant arthritis.^ieng