Gas chromatography of some prostaglandins

Summary The synthetic prostaglandin F_2α (PG_2α), used as an ovulation regulator, and abortion inducer, is contaminated with its Δ5-trans isomer. The ratio of the isomers was measured by capillary GC. The prostaglandin I₂ (PGI₂), a blood platelet aggregation inhibitor, easily decomposes to 6-ketoprostaglandin F_1α (6-keto PG_1α). Their ratio was measured by packed column GC. An unexpected, instantaneous methanol cleavage was observed during the GC analysis of the methyl ketal derivative of 6-keto PG_1α, forming a mixture of PGI₂ and Δ6-prostaglandin I₁ (Δ6-PGI₁). Presented at the 15th International Symposium on Chromatography, Nürnberg, October 1984     

Mobile Phase Selectivity in the Separation of Epimeric and Positionally Isomeric Triterpenoids by Reversed Phase High Performance Liquid Chromatography

The chromatographic behavior of 8 lanostanoid triterpenes including 4 pairs of C-3 epimers and 2 pairs of C-3/C-15 positional isomers was studied by reversed phase HPLC. A consistent elution sequence of these paired isomers in all combination of three distinct solvent systems was observed. The mobile phase selectivity to separate specific, paired isomers was mainly dependent on the polarity difference contributed by C-3/C-15 hydroxyl and acetoxyl groups. The stereochemistry at C-3 played more profound role in guiding the hydrophobic interaction with C₁₈ stationary phase in the separation of C-3/C-15 positional isomers.     

Simultaneous quantitative profiling of 20 isoprostanoids from omega-3 and omega-6 polyunsaturated fatty acids by LC–MS/MS in various biological samples

Isoprostanoids are a group of non-enzymatic oxygenated metabolites of polyunsaturated fatty acids. It belongs to oxylipins group, which are important lipid mediators in biological processes, such as tissue repair, blood clotting, blood vessel permeability, inflammation and immunity regulation. Recently, isoprostanoids from eicosapentaenoic, docosahexaenoic, adrenic and α-linolenic namely F₃-isoprostanes, F₄-neuroprostanes, F₂-dihomo-isoprostanes and F₁-phytoprostanes, respectively have attracted attention because of their putative contribution to health. Since isoprostanoids are derived from different substrate of PUFAs and can have similar or opposing biological consequences, a total isoprostanoids profile is essential to understand the overall effect in the testing model. However, the concentration of most isoprostanoids range from picogram to nanogram, therefore a sensitive method to quantify 20 isoprostanoids simultaneously was formulated and measured by liquid chromatography-tandem mass spectrometry (LC–MS/MS). The lipid portion from various biological samples was extracted prior to LC–MS/MS evaluation. For all the isoprostanoids LOD and LOQ, and the method was validated on plasma samples for matrix effect, yield of extraction and reproducibility were determined. The methodology was further tested for the isoprostanoids profiles in brain and liver of LDLR^−/− mice with and without docosahexaenoic acid (DHA) supplementation. Our analysis showed similar levels of total F₂-isoprostanes and F₄-neuroprostanes in the liver and brain of non-supplemented LDLR^−/− mice. The distribution of different F₂-isoprostane isomers varied between tissues but not for F₄-neuroprostanes which were predominated by the 4(RS)-4-F_4t-neuroprostane isomer. DHA supplementation to LDLR^−/− mice concomitantly increased total F₄-neuroprostanes levels compared to F₂-isoprostanes but this effect was more pronounced in the liver than brain.     

Development of a mass spectrometric hydroxyl‐position determination method for the hydroxyindole metabolites of JWH‐018 by GC‐MS/MS

One of the many issues of designer drugs of abuse like synthetic cannabinoids (SCs) such as JWH‐018 is that details on their metabolism has yet to be fully elucidated and that multiple metabolites exist. The presence of isomeric compounds poses further challenges in their identification. Our group has previously shown the effectiveness of gas chromatography‐electron ionization‐tandem mass spectrometry (GC‐EI‐MS/MS) in the mass spectrometric differentiation of the positional isomers of the naphthoylindole‐type SC JWH‐081, and speculated that the same approach could be used for the metabolite isomers. Using JWH‐018 as a model SC, the aim of this study was to differentiate the positional isomers of its hydroxyindole metabolites by GC‐MS/MS. Standard compounds of JWH‐018 and its hydroxyindole metabolite positional isomers were first analyzed by GC‐EI‐MS in full scan mode, which was only able to differentiate the 4‐hydroxyindole isomer. Further GC‐MS/MS analysis was performed by selecting m/z 302 as the precursor ion. All four isomers produced characteristic product ions that enabled the differentiation between them. Using these ions, MRM analysis was performed on the urine of JWH‐018 administered mice and determined the hydroxyl positions to be at the 6‐position on the indole ring. GC‐EI‐MS/MS allowed for the regioisomeric differentiation of the hydroxyindole metabolite isomers of JWH‐018. Furthermore, analysis of the fragmentation patterns suggests that the present method has high potential to be extended to hydroxyindole metabolites of other naphthoylindole type SCs in identifying the position of the hydroxyl group on the indole ring. Copyright © 2016 John Wiley & Sons, Ltd.     

Nano-LC-MS/MS for the quantitation of prostanoids in immune cells

Prostanoids, derivatives of arachidonic acid, are involved in inflammation and immune reactions. To understand the role of prostanoids produced by diverse immune cells, a highly sensitive quantitation method for prostaglandin E₂ (PGE₂), prostaglandin D₂ (PGD₂), 6-keto prostaglandin F_1α (6-keto PGF_1α), prostaglandin F_2α (PGF_2α), and thromboxane B₂ (TXB₂) by means of nano-liquid chromatography-tandem mass spectrometry has been developed. It was validated according to the guidelines of the Food and Drug Administration (FDA) in terms of linearity, precision, accuracy, recovery, stability, and lower limit of quantitation (LLOQ). The LLOQ were 25 pg/mL in the injected solution (75 fg on column (o.c.)) for PGE₂ and PGD₂ and 37.5 pg/mL (112.5 fg on column) for 6-keto PGF_1α, PGF_2α, and TXB₂, respectively. It was successfully applied to murine mast cells isolated from paws after zymosan injection and to CD4⁺ and CD8⁺ T lymphocytes from blood of sensitized versus non-sensitized mice in context of a delayed type hypersensitivity model. About 5,000 (T cells) to 40,000 (mast cells) cells were sufficient for quantitation. In the mast cells, the production of PGE₂ increased at a significantly higher extent than the synthesis of the other prostanoids. The T lymphocytes did not show any difference in prostanoid production, no matter whether they were obtained from sensitized mice or non-sensitized mice.     

Radioimmunoassay for Thromboxane B2

Abstract

A radioimmunoassay for a prostaglandin endoperoxide metabolite, thromboxane B₂, was developed. The antibodies were very specific for this compound, and the method had a sensitivity of 10 pg. Platelets from a human subject were aggregated by addition of collagen and the synthesis of prostaglandin endoperoxides with time was monitored by parallel assay of prostaglandin E₂, F_2α ^and thromboxane B₂.     

Preparation and Characterization of 61,6n-DI-O-(α-D-Galactopyranosyl)Cyclomaltooctaoses

ABSTRACT

Four positional isomers of 6¹,6ⁿ-di-O-(D-galactopyranosyl)cyclomaltooctaoses (cG₈s, γ-cyclodextrins) (n = 2?5) were chemically synthesized using the trichloroacetimidate method. The desired compounds having two α-(1→6)-linkages were isolated from a mixture of configurational isomers by HPLC, and their structures were confirmed by ¹³C NMR spectroscopy and FAB-high resolution mass spectra (HRMS). The elution behavior of their four positional isomers on an ODS column by HPLC is discussed.     

Total Synthesis of 15-Oxo-13,14-Dihydro-prostaglandin F2α, A Major Metabolite of Prostaglandin F2α

The synthesis of prostaglandin F_2α metabolite starting from the Corey lactone is described. The key step of the synthesis involves chemoselective Wittig reaction.     

Photochemische Reaktionen 94. Mitteilung. Vinyloge β-Spaltung bei Epoxy-enonen der Jonon-Reihe

Photoinduced Vinylogous β-Cleavage of Epoxy-enones of the Ionone Series The photochemistry of the α,β-unsaturated γ,δ-epoxy-enones 1–3 is determined by: (i) C(γ)-O-scission of the epoxide (vinylogous β-cleavage of Type A); (ii) C(γ)-C(δ)-cleavage of the oxirane (vinylogous β-cleavage of Type B); (iii) (E/Z)-isomerization of the enone chromophore. In contrast, 4 with tertiary C(β) shows no Type B cleavage. Type A cleavage is induced both by n,π*- and π,π*-excitation and arises probably from the T₁-state, but Type B cleavage is observed only on π,π*-excitation and represents presumably a S₂-reaction. On Type A cleavage 1–4 undergo 1,2-alkyl-shifts to 1,5-dicarbonyl compounds ( 15–18, 25–28, 34 and 35 ) or rearrange to dihydrofuranes ( 7 and 30 ). The isomerization 1→7 proceeds by a stereoselective [1,3]-sigmatropic shift. On Type B cleavage 1–3 isomerize to a bicyclic enol-ether ( 8, 29 ) or to a monocyclic enol-ether ( 9 ; product of a homosigmatropic [1,5]-shift) or undergo fragmentation to isomers such as allenes 10, 22 and 31 or cyclopropenes 11 and 21 . The non-isolated, unstable (Z)-epoxy-enones 14, 19, 24 and 38 isomerize by fragmentation to the furanes 12, 23, 33 and 39 respectively, on contact with traces of acid or by heating. However, for 19 and 4 , Type B cleavage may lead to the furanes 23 and 39 . On UV. irradiation of the epoxy-enone 4 the initially formed (E/Z)-isomers 34 and 35 yield on π,π*-excitation the enones 37 and 40 by a vinylogous β-fragmentation. In addition, on n,π*-excitation 34 isomerizes to 35 , which decarbonylates exclusively to the enone 37 . The reactions of 1–4 with BF₃ · O(C₂H₅)₂ were also studied (see appendix). The epoxy-enones 1 and 2 isomerize by an 1,2-alkyl shift in good yield to the 1,4-dicarbonyl compounds 79 and 81 , whereas 3 gives the 1,4-diketone 83 , and in small amounts the 1,5-diketone 84 . On the other hand, 4 is converted to the fluorohydroxy-enone 85 and to the 1,5-dicarbonyl product 34 , the only isomer in this series which is identical with one of the photoproducts.     

A novel synthesis of 2,3-dinor-6-oxo-prostaglandin F1α

The synthesis of 2,3-dinor-6-oxo-prostaglandin F_1α, the major metabolite of prostacyclin, from the prostaglandin lactone intermediate (2) is reported.     

Chromatographic separation of benzo(a)pyrene isomers on nematic liquid crystal GC and polymeric reversed-phase HPLC phases

Summary The use of two nematic liquid crystals (BABT and BPhBT) as GC stationary phases for the separation of monohydroxybenzo(a)pyrenes. as their TMS ethers, and monomethylbenzo(a)pyrenes was developed and compared with the separation of these isomers by HPLC using a polymeric ODS reversed-phase column. It was found that while HPLC and GC gave comparable separation of the hydroxy isomers, 10 out of 12 separated, better separation of the methyl isomers was obtained using HPLC. A simultaneous use of both HPLC and GC would resolve the twelve hydroxy isomers in about 70min. The results indicated that HPLC, using polymeric reversed-phase columns, is as powerful a tool as GC using nematic liquid crystal phases, for the separation of benzo(a)pyrene isomers. A discussion of the effect of solute length-to-breadth ratio on elution order is presented.Presented in part at the 1981 Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy, Atlantic City, NJ; paper No. 51.     

Separation of Methylated Purines by High Pressure Liquid Chromatography

Abstract

This paper reports a method for the separation and measurement of methylated purines of interest to carcinogenesis studies by high-pressure liquid chromatography (HPLC) following their column chromatographic isolation from collected urine samples. HPLC was evaluated on three different cation-exchange columns, with optimum conditions obtained on a Partisil 10-SCX column employing isocratic elution with 0.25M (NH₄)H₂PO₄ at pH 4.0. This column was also found to be useful for the separation of mono-methylguanine isomers. Application is shown to the analysis of rat urine following animal treatment with methyl methanesulfonate.     

Spin-spin coupling constant 3 J H,F as a reliable criterion for recognition of individual regioisomeric and tautomeric pairs of H(CF2)2-containing isoxazoles and pyrazoles

The spin-spin coupling constant ³ J _H,F of the H(CF₂)₂ group varies within 1.6—3.5 Hz for 5-R^F- and 3.8—4.5 Hz for 3-R^F-isoxazoles and pyrazoles in CDCl₃ and can serve as a reliable criterion for recognition of regioisomeric and tautomeric structures of H(CF₂)₂-containing heterocyclic compounds.     

Synthesis and molecular structures of heptafluoroisopropylated fullerenes: C60(i-C3F7)8, C60(i-C3F7)6, and C60(CF3)2(i-C3F7)2

One isomer of C₆₀(i-C₃F₇)₈, three isomers of C₆₀(i-C₃F₇)₆, and the first mixed perfluoroalkylated fullerene, C₆₀(CF₃)₂(i-C₃F₇)₂, have been isolated by HPLC from a mixture prepared by reaction of C₆₀ with heptafluoroisopropyl iodide in a glass ampoule at 260-290 °C. The molecular structures of the four new compounds have been determined by means of X-ray single crystal diffraction partially also by use of synchrotron radiation. Theoretical calculations at the DFT level of theory have been employed to rationalize the energetics of isomers and of C₆₀-R_f binding.     

Stereoselective ring-opening reactions with AcBr and AcCl. A new method for preparation of some haloconduritols

The actions of AcX (X=Br, Cl) on 7-oxa-bicyclo[2.2.1]hept-5-ene-2,3-diol diacetates and a transoid-epoxide prepared from the acetonide of cyclohexa-3,5-diene-cis-1,2-diol were studied. H₂SO₄-catalyzed cleavage of exo-cis-7-oxa-bicyclo[2.2.1]hept-5-ene-2,3-diol diacetate with AcCl gave (1α,2α,3α,6β)-6-chloro-4-cyclohexene-1,2,3-triol triacetate, from which the corresponding chloroconduritol was obtained by trans-esterification (MeOH/HCl). A similar reaction of the exo-diacetate with AcBr in the presence of H₂SO₄ resulted in bromine addition. The formation of bromine from the reaction of AcBr and H₂SO₄ was observed by independent experiments. H₂SO₄-catalyzed reaction of endo-cis-7-oxa-bicyclo[2.2.1]hept-5-ene-2,3-diol diacetate with AcX (X=Br, Cl) gave (1α,2α,3β,6β)-6-halo-4-cyclohexene-1,2,3-triol triacetates. The reaction of the transoid-epoxide with AcX (X=Br, Cl) with no catalyst gave also (1α,2α,3β,6β)-6-halo-4-cyclohexene-1,2,3-triol triacetates.     

Determination of 8‐epi PGF2α concentrations as a biomarker of oxidative stress using triple‐stage liquid chromatography/tandem mass spectrometry

F2‐isoprostanes are a family of prostaglandin F2‐like compounds that are formed by free‐radical‐catalyzed peroxidation of arachidonic acid. Several F2‐isoprostanes, but in particular 8‐epi PGF_2α, are widely used as oxidative stress biomarkers. An analytical method based on liquid chromatography with negative electrospray ionization (ESI) coupled to tandem mass spectrometric detection (LC/MS/MS) was developed for the determination of 8‐epi PGF_2α concentrations in human plasma, whole blood, erythrocytes and urine. 8‐epi PGF_2α‐d4, a stable isotope derivative of 8‐epi PGF_2α, was used as an internal standard (IS). A 50 µL sample was focused on‐column and separated on two 3 µm particle size SUPELCOSIL? ABZ+Plus HPLC columns (15 cm × 4.6 mm and 7.5 cm × 4.6 mm) connected in series. An Applied Biosystems 4000 Q TRAP LC/MS/MS system with ESI was operated in multiple reaction monitoring (MRM) mode with the precursor‐to‐product ion transitions m/z 353.4 → 193.1 (8‐epi PGF_2α), 357.4 → 197.1 (8‐epi PGF_2α‐d4), used for quantification. The assay was fully validated and found to have adequate accuracy, precision, linearity, sensitivity and selectivity. The mass limit of detection (mLOD) was 1 pg of analyte eluting from the column. The assay has been successfully applied to the analysis of human plasma, whole blood, erythrocytes and urine samples. Copyright © 2009 John Wiley & Sons, Ltd.     

Photochemical Reactions. Part 67. photochemistry of saturated aliphatic and cyclic β-keto sulfoxides (Cα?S)- and α-cleavage – A new case of photostereomutation

The photochemical behaviour of saturated aliphatic ( 2, 4 , and 5 ) and bicyclic ( 18 and 19 ) β-keto sulfoxides has been studied. Photostereomutation of the sulfoxide group was observed on irradiation of 4a, 4b, 18 , and 19 . Most likely an internal energy transfer from the excited carbonyl to the sulfoxide group is operating on direct irradiation of such compounds. Prolonged photolysis of an aliphatic β-keto sulfoxide, which is nonalkylated a t the α-carbon ( 2 ), yielded a product due to preferential (C_α-S)-cleavage ( 24 ). Mono- ( 4 ) and dialkylated- ( 5 , 6 , and 8 )analogues primarily afforded products due to α-cleavage ( 26–31 and 32 ). The carboxylic acid S-methylesters ( 26–31 ) were exclusively formed by an intermolecular path. Prolonged irradiation of the bicyclic β-keto sulfoxides 18 and 19 favored the formation of a desulfurized compound 34 due to initial ( C_α-S )-cleavage.     

Glass capillary column chromatography of mixed derivatives of primary prostaglandins, 6-keto prostaglandin F1α and thromboxane B2

This work describes the use of glass capillary columns (GCC) in the rapid concurrent analysis of primary prostaglandins (PGs) (e. g. PGE₂, PGE₂, PGF_2α) and other functionally significant metabolites of arachidonic acid (AA) such as TXB₂ and 6-keto PGF_1α. The use of a new multistep mixed derivatization approach that generates the methyl esters of n-butylboronate, pentafluorobenzyloxime, trimethylsilyl ether derivatives of these compounds remarkably simplifies the GC profiling of the three main pathways of AA metabolism (PGs, thromboxane, and prostacyclin). Furthermore, isomeric species giving very similar or identical mass spectral patterns can be easily identified by their relative retention times on high efficiency capillary columns.     

Analyses of a Mixture of Glucosyl-Cyclomaltoheptaoses Prepared on an Industrial Scale

ABSTRACT

A mixture of glucosyl-cyclomaltoheptaoses (β-cyclodextrins, βCDs) was prepared by glucoamylolysis of a mixture of maltosyl-βCDs which was produced on an industrial scale from maltose and β CD through the reverse action of Klebsiella pneumoniae pullulanase. Glucosyl-βCDs in the mixture were separated by HPLC on a reversed phase column and their molecular weights were measured by FAB-MS. In addition, the number of side-chains in each molecule was confirmed by methylation analysis and it was proved that the mixture comprised mainly of a monoglucosyl-βCD [G-β CD] and diglucosyl-β-CDs [(G)₂-βCDs], and as a minor component triglucosyl-β CDs [(G)₃-βCDs], and that G-, (G)₂-, and (G)₃-β CDs were produced in the ratios of 50%:45%:5%. The structures of three positional isomers of (G)₂-β CD were established by HPLC analysis of partial hydrolyzates, ¹³C NMR spectroscopy, and chemical synthesis. Four regioisomeric (G)₃-β CDs which could be isolated were characterized by ¹³C NMR spectroscopy.     

Total synthesis of PGF2α and 6,15-diketo-PGF1α and formal synthesis of 6-keto-PGF1α via three-component coupling

The asymmetric total synthesis of PGF_2α and 6,15-diketo-PGF_1α and formal synthesis of 6-keto-PGF_1α from a common key intermediate are described. The key intermediate, which has a chiral cyclopentane backbone possessing suitable functional groups with required stereochemistry for both side chains, was prepared from (R)-4-silyloxy-2-cyclopentenone through a three-component coupling reaction. The Wittig reaction, Nozaki-Hiyama-Kishi (NHK) coupling and cross metathesis completed the synthesis of PGF_2α, 6,15-diketo-PGF_1α and 6-keto-PGF_1α.