Simultaneous determination of 6β- and 6α-hydroxycortisols and 6β-hydroxycortisone in human urine by stable isotope dilution mass spectrometry

A capillary gas chromatographic–mass spectrometric method for the simultaneous determination of 6β-hydroxycortisol (6β-OHF, 6β,11β,17α,21-tetrahydroxypregn-4-ene-3,20-dione), 6α-hydroxycortisol (6α-OHF, 6α,11β,17α,21-tetrahydroxypregn-4-ene-3,20-dione) and 6β-hydroxycortisone (6β-OHE, 6β,17α,21-trihydroxypregn-4-ene-3,11,20-trione) in human urine is described. Deuterium-labelled compounds, 6β-[1,1,19,19,19-²H₅]OHF (6β-OHF-d₅), 6α-[1,1,19,19,19-²H₅]OHF (6α-OHF-d₅) and 6β-[1,1,19,19,19-²H₅]OHE (6β-OHE-d₅) were used as internal standards. Quantitation was carried out by selected-ion monitoring of the characteristic fragment ions ([M-31]⁺) of the methoxime–trimethylsilyl (MO–TMS) derivatives of 6β-OHF, 6α-OHF and 6β-OHE. The sensitivity, specificity, precision and accuracy of the method were demonstrated to be satisfactory for measuring 6β-OHF, 6α-OHF and 6β-OHE in human urine.     

Determination of serum unconjugated estrone, estradiol-17β and estriol during pregnancy by selected ion monitoring

A simple, rapid and highly specific method by selected ion monitoring (SIM), using 9α,11α-[²H₂]estrone, [2,4-²H₂]estradiol-17β and 2,4-[²H₂]estriol as internal standards, was developed for the determination of serum estrogens during pregnancy. Serum samples were submitted to a simple extraction procedure and were analysed after formation of the trifluoroacetic anhydride derivative. The inter-assay coefficients of variation for estrone, estradiol-17β and estriol were 3.73%, 3.42% and 3.49%, respectively. The results obtained by SIM were compared with analysis performed using radioimmunoassay.     

Biosynthesis of 20-hydroxyecdysone in Ajuga hairy roots: fate of 6α- and 6β-hydrogens of lathosterol

The fate of 6α- and 6β-hydrogens of lathosterol during the transformation into 20-hydroxyecdysone was chased by feeding [3α,6β-²H₂]- and [3α,6α-²H₂]-lathosterols to hairy roots of Ajuga reptans var. atropurpurea. The behavior of 6β-hydrogen, which mostly migrated to the C-5 position of 20-hydroxyecdysone, was in agreement with that of C-6 hydrogen of cholesterol. The results strongly supported the view that cholesterol and lathosterol are first metabolized into 7-dehydrocholesterol, which is then converted into 20-hydroxyecdysone via 7-dehydrocholesterol 5α,6α-epoxide in the hairy roots.     

The partial characterization of 1-oxygenated steroids from urine of a hypertensive newborn child

1. Four substances from the urine of a hypertensive newborn girl were partially characterized and shown to be 17α-hydroxy-5β-pregnane-1,3,20-trione, 3α,17α-dihydroxy-5β-pregnane-1,20-dione, 3α,17α,20α-trihydroxy-5β-pregnan-1-one and 5β-pregnane-1β,3α,17α,20α-tetrol. 2. The characterization rested mainly on R_M analysis of the substances and their derivatives by glycol fission, providing evidence for position and degree of substitution and for steroidal character. Supporting evidence was provided by chemically specific location reactions. 3. Certain problems in the manipulation of these β-disubstituted steroids are discussed.     

Polar corticosteroids in human neonatal urine; Synthesis and gas chromatography-mass spectrometry of ring a reduced 6-hydroxylate corticosteroids

This report describes the synthesis of 3α, 6β, 11β, 17α, 21-pentahydroxy-5β-pregnane-20-one, 3α, 6β, 11β, 17α, 21-pentahydroxy-5α-pregnane-20-one, 3α, 6α, 11β, 17α, 21-pentahydroxy-5β-pregnane-20-one, 3α, 6α, 11β, 17α, 21-pentahydroxy-5α-pregnane-20-one, 3α, 6β, 17α, 21-tetrahydroxy-5β-pregnane-11, 20-dione, 3α, 6β, 17α, 21-tetrahydroxy-5α-pregnane-11, 20-dione, 3α, 6α, 17α, 21-tetrahydroxy-5β-pregnane-1 1, 20-dione and 3α, 6α, 17α, 21-tetrahydroxy-5α-pregnane-11, 20-dione.The gas chromatographic-mass spectrometric properties of these compounds are given. Proof of structure was accomplished using gas chromatography-mass spectrometry, microchemical reactions, optical rotatory dispersion and nuclear magnetic resonance spectroscopy.     

Microbiological Hydroxylation of Cinerone to Cinerolone

Cinerone [2-(2′-cis-butenyl)-3-methyl-2-cyclopenten-1-one] is hydroxylated to cinerolone [2-(2′-cis-butenyl)-3-methyl-4-hydroxy-2-cyclopenten-1-one] by a number of streptomycetes, bacteria, and fungi. Aspergillus niger ATCC 9,142 and Streptomyces aureofaciens ATCC 10,762 were found to be the most effective hydroxylators. The optical activity of the product was found to range from [α]_D²⁵ 0° to -8.6°, depending on the organism and conditions of culture. Two additional hydroxylated products of cinerone have been isolated and identified as 2-n-butyl-4-hydroxy-3-methyl-2-cyclopenten-1-one and 2-(2′-cis-butenyl-4′-hydroxy)-3-methyl-2-cyclopenten-1-one, respectively.     

Simultaneous determination of stable isotopically labelled l-histidine and urocanic acid in human plasma by stable isotope dilution mass spectrometry

A capillary gas chromatographic—mass spectrometric method for the simultaneous determination of stable isotopically labelled l-histidine (l-[3,3-²H₂,1′,3′-¹⁵N₂]histidine, l-His-[M + 4]) and urocanic acid ([3-²H,1′,3′-¹⁵N₂]urocanic acid, UA-[M + 3]) in human plasma was developed using dl-[2,3,3,5′-²H₄,2′-¹³C,1′,3′-¹⁵N₂]histidine (dl-His-[M + 7]) and [2,3,5′-²H₃,2′-¹³C,1′,3′-¹⁵N₂]urocanic acid (UA-[M + 6]) as internal standards. l-Histidine and urocanic acid were derivatized to ^αN-(trifluoroacetyl)-^imN-(ethoxycarbonyl)-l-histidine n-butyl ester and ^imN-(ethoxycarbonyl)urocanic acid n-butyl ester. Quantification was carried out by selected ion monitoring of the molecular ions of the respective derivatives of l-His-[M + 4], dl-His-[M + 7], UA-[M + 3] and UA-[M + 6]. The sensitivity, specificity, precision and accuracy of the method were demonstrated to be satisfactory for measuring plasma concentrations of l-His-[M + 4] and UA-[M + 3] following administration of trace amounts of l-His-[M + 4] to humans.     

The effect of indomethacin on the biosynthesis and metabolism of prostaglandin F2α in man

Healthy volunteers received 60 μg of [8,10,10-²H₃] PGF_2α by intravenous infusion both before and during a course of treatment with indomethacin (200 mg/day). Excretion of deuterated 5α, 7α-dihydroxy-11-ketotetranor-prostane-1, 16-dioic acid in urine was quantified by GC-MS using a reverse stable isotope dilution procedure. Indomethacin was found to have no detectable effect on the metabolism of the labelled PGF_2α whereas output of the endogenous metabolite was markedly reduced by the effect of the drug on prostaglandin biosynthesis.     

Biosynthetic Origin of Gibberellins A(3) and A(7) in Cell-Free Preparations from Seeds of Marah macrocarpus and Malus domestica

Cell-free preparations from seeds of Marah macrocarpus L. and Malus domestica L. catalyzed the conversion of gibberellin A₉ (GA₉) and 2,3-dehydroGA₉ to GA₇; GA₉ was also metabolized to GA₄ in a branch pathway. The preparation from Marah seeds also metabolized GA₅ to GA₃ in high yield; GA₆ was a minor product and was not metabolized to GA₃. Using substrates stereospecifically labeled with deuterium, it was shown that the metabolism of GA₅ to GA₃ and of 2,3-dehydroGA₉ to GA₇ occurs with the loss of the 1β-hydrogen. In cultures of Gibberella fujikuroi, mutant B1-41a, [1β,2β-²H₂]GA₄, was metabolized to [1,2-²H₂]GA₃ with the loss of the 1α- and 2α-hydrogens. These results provide further evidence that the biosynthetic origin of GA₃ and GA₇ in higher plants is different from that in the fungus Gibberella fujikuroi.     

Urinary excretion of C-20-reduction products of corticosterone

1. A 200 mg. portion of corticosterone was ingested by a healthy man and the urine collected. Part of the urine was treated with the gastric juice of Helix pomatia and extracted with ethyl acetate, and the extract fractionated with Girard T. Paper-chromatographic separation of the non-ketonic fraction in the Bush (1952) system B₅ revealed the presence of two unknown polar components. 2. The unknown compounds did not possess a reducing (blue tetrazolium) or a reducible (potassium borohydride) grouping. Both contained a terminal α-glycollic fragment as shown by the formation of formaldehyde, and of a non-volatile aldehyde on oxidation with sodium bismuthate. 3. Unknown compound (I) had paper-chromatographic mobilities identical with those of 5β-pregnane-3α,11β,20β,21-tetraol. The oxidation product of compound (I) had a retention time (gas–liquid chromatography) on an SE30 column identical with that of 3α,11β-dihydroxy-21-nor-5β-pregnan-20-al. The retention times of various derivatives agreed with those produced in an identical manner on the standard, and accordingly compound (I) is formulated as 5β-pregnane-3α,11β,20ξ,21-tetraol. 4. Unknown compound (II) had a higher R_F than compound (I), and its oxidation product had a longer retention time than that of compound (I). From the group effects observed in paper and gas–liquid chromatography, compound (II) is tentatively formulated as 5α-pregnane-3α,11β,20ξ,21-tetraol. The 5α/5β ratio found was about 2·0.     

Relative stability of α-melanotropin and related analogues to rat brain homogenates

α-Melanotropin (α-MSH) retains less than 1% of its original activity after a 60 min incubation with 10% rat brain homogenate. [Nle⁴, D-Phe⁷]-α-MSH is nonbiodegradable in rat serum (240 min incubation) and still maintains 10% of its original activity in 10% rat brain homogenate (240 min incubation). The related fragment analogue, Ac-[Nle⁴, D-Phe⁷]-α-MSH_4–10-NH₂, retains 50% of its activity after a 240 min incubation in rat brain homogenate, whereas Ac-[Nle⁴, D-Phe⁷]-α-MSH_4–11-NH₂ is totally resistant to inactivation by rat brain homogenate. Both [Nle⁴, D-Phe⁷]-fragments are resistant to degradation by rat serum, but [Nle⁴]-α-MSH, Ac-[Nle⁴]-α-MSH_4–10-NH₂ and Ac-[Nle⁴]-α-MSH_4–11-NH₂ are rapidly inactivated under both conditions. The cyclic melanotropin, [ ]-α-MSH, is inactivated in rat brain homogenate as is the shorter Ac-[ ]-α-MSH_4–10-NH₂ analogue, but neither cyclic melanotropin is inactivated upon incubation in serum from rats. Ac-[ ]-α-MSH_4–10-NH₂ is resistant to inactivation by either rat serum or a brain homogenate. Some of these melanotropin analogues may provide useful probes for the localization and characterization of putative melanotropin receptors in both the central nervous system and peripheral tissues.     

Synthesis of bent titanocene metalloligands with the (diphenylphosphino)tetramethylcyclopentadienyl moiety. X-ray structure of [(η-C5Me4 2)2TiCl2]Mo(CO)4

The reactions of lithium(diphenylphosphino)tetramethylcyclopentadienide with CpTiCl₃ and secondly with TiCl₃ followed by CCl₄ oxidation lead to the formation of two titanocene phosphines: (η⁵-C₅H₅)[η⁵-C₅Me₄P(C₆H₅)₂]TiCl₂ (2) and [η⁵-C₅Me₄P(C₆H₅)₂]₂TiCl₂ (3), respectively. The metalloligand 3 reacts readily with Mo(CO)₄cod, Mo(CO)₅THF and Mo(CO)₆ to give in each case [(η⁵-C₅Me₄ o(CO)₄ (6) as a sole product. The structure of 6 has been determined by X-ray diffraction. Crystal data: P , a = 11.716(1), b = 11.753(2), c = 16.110(2) Å, α = 99.06(1), β = 92.61(1), γ = 104.20(1)°, Z = 2. The molybdenum-titanium distance of 5.194(1) Å rules out any metal-metal interaction. The chlorine substitution reactions by CO in 2 and 3 and by thiolate group (pH₃C-C₆H₄-S) in 16 are reported.     

Stable isotope dilution analysis of human urinary metabolites of 17α-methyltestosterone

A method based on gas chromatography–mass spectrometry–selected-ion monitoring was developed to measure the main metabolites of 17α-methyltestosterone, 17α-methyl-5α-androstan-3α,17β-diol and 17α-methyl-5β-androstan-3α,17β-diol, in human urine. 17α-Methyl-[²H₃]-5α-androstan-3α,17β-diol and 17α-methyl-[²H₃]-5β-androstan-3α,17β-diol were used as internal standards. The methods involved purification using a Sep-Pak C₁₈ cartridge, hydrolysis by β-glucuronidase from Ampullaria and derivatization with N-methyl-N-trimethylsilyl-trifluoroacetamide/dithioerythriol/ammonium iodide. Quantitation was achieved by selected-ion monitoring of the characteristic fragment ions ([(M+H)−2×TMSOH]⁺) of the di-TMS derivatives on the chemical ionization mode. The method provides a specific, sensitive and reliable technique to determine the urine levels of 17α-methyl-5α-androstan-3α,17β-diol and 17α-methyl-5β-androstan-3α,17β-diol, and can be applied to pharmacokinetic studies of 17α-methyltestosterone.     

The structures of bis(1H,5H-S-methylisothiocarbonohydrazidium) di-μ-chloro-octachlorodibismuthate(III) tetrahydrate and tris(1H-S-methylisothiocarbonohydrazidium) esachlorobismuthate(III)

The structures of bis(1H⁺,5H⁺-S-methylisothiocarbonohydrazidium) di-μ-chlorooctachlorodibismuthate(III) tetrahydrate: (C₂H₁₀N₄S)₂(Bi₂Cl₁₀)· 4H₂O (compound [I]) and of tris(1H⁺-S-methylisothiocarbonohydrazidium) esachlorobismuthate(III): (C₂H₉N₄S)₃(BiCl_5.67I_0.33) (compound [II]) were determined from single crystal X-ray diffractometer data. Both compounds crystallize as triclinic (P ); crystals [I] with Z = 1 formula unit in a cell of constants: A = 10.621(3), B = 9.989(5), C = 7.439(3) Å, α = 88.31(2), β = 84.51(2), γ = 68.88(2)°, final R = 0.0427 for 2229 unique reflections with I 2σ(I); crystals [II] with Z = 2 and cell dimensions: A = 14.109(4), B = 12.209(9), C = 8.206(7) Å, α = 103.54(3), β = 104.95(2), γ = 81.96(2)°, final R = 0.0411 for 3637 unique reflections (1 2σ(I)). The structure of [I] is built up of diprotonated organic cations, water molecules and dinuclear centrosymmetric [Bi₂Cl₁₀]⁴⁻ anions held together by N-HCl, N-HO, O-HCl hydrogen bonds and Van der Waals interactions. The [Bi₂Cl₁₀]⁴⁻ complex consists of two edge-sharing octahedra in which three pairs of bonds of similar length are observed (Bi-Cl_av = 2.602(5), 2.712(4), 2.855(5) Å). The structure of [II] consists of monoprotonated cations and [BiCl_5.67I_0.33]³⁻ anions held together by a tridimensional network of hydrogen bonds. Each bismuth atom is octahedrally surrounded by six chlorine atoms, one of which is statistically substituted by a iodine atom.     

Gas chromatographic–mass spectrometric determination of α-ketoisocaproic acid and [H7]α-ketoisocaproic acid in plasma after derivatization with N-phenyl-1,2-phenylenediamine

A method for determination of α-ketoisocaproic acid (KIC) and [4,5,5,5,6,6,6-²H₇]α-ketoisocaproic acid ([²H₇]KIC) in rat plasma was developed using gas chromatography–mass spectrometry-selected ion monitoring (GC–MS-SIM). [5,5,5-²H₃]α-Ketoisocaproic acid ([²H₃]KIC) was used as an analytical internal standard to account for losses associated with the extraction, derivatization and chromatography. The keto acids were extracted by cation-exchange chromatography using BondElut SCX cartridge and derivatized with N-phenyl-1,2-phenylenediamine to form N-phenylquinoxalinone derivatives. Quantitation was performed by SIM of the respective molecular ions at m/z 278, 281 and 285 for the derivatives of KIC, [²H₃]KIC and [²H₇]KIC on the electron impact method. The limit of detection was found to be 70 fmol per injection (S/N=3) and the limit of quantitation for [²H₇]KIC was around 50 nM in rat plasma. Endogenous KIC concentrations in 50 μl of rat plasma were measured with relative intra- and inter-day precision of 4.0% and 3.3%, respectively. The intra- and inter-day precision for [²H₇]KIC spiked to rat plasma in the range of 0.1 to 10 μM gave good reproducibility with relative standard deviation (RSD) of 6.5% and 5.4%, respectively. The intra- and inter-day relative errors (RE) for [²H₇]KIC were less than 6.4% and 3.8%, respectively. The method was applied to determine the plasma concentration of [²H₇]KIC after an intravenous administration of [²H₇]KIC in rat.     

15β-hydroxysteroids (Part IV). Steroids of the human perinatal period: The synthesis of 3α,15β,17α-trihydroxy-5α-pregnan-20-one and its A/B-ring configurational isomers

In recent years several 15β-hydroxysteroids have emerged pathognomonic of adrenal disorders in human neonates of which 3α,15β,17α-trihydroxy-5β-pregnan-20-one (2) was the first to be identified in the urine of newborn infants affected with congenital adrenal hyperplasia. In this investigation we report the synthesis of the three remaining 3ξ,5ξ-isomers, namely 3α,15β,17α-trihydroxy-5α-pregnan-20-one (3), 3β,15β,17α-trihydroxy-5α-pregnan-20-one (7) and 3β,15β,17α-trihydroxy-5β-pregnan-20-one (8) for their definitive identification in pathological conditions in human neonates. 3β,15β-Diacetoxy-17α-hydroxy-5-pregnen-20-one (11), a product of chemical synthesis was converted to the isomeric 3 and 7, while conversion of 15β,17α-dihydroxy-4-pregnen-3,20-dione (4), a product of microbiological transformation, resulted in the preparation of 8. In brief, selective acetate hydrolysis of 11 gave 15β-acetoxy-3β,17α-dihydroxy-5-pregnen-20-one (12) which on catalytic hydrogenation gave 15β-acetoxy-3β,17α-dihydroxy-5α-pregnan-20-one (13) a common intermediate for the synthesis of the 3β(and α),5α-isomers. Hydrolysis of the 15β-acetate gave 7, whereas oxidation with pyridinium chlorochromate gave 15β-acetoxy-17α-hydroxy-5α-pregnan-3,20-dione (14) which on reduction with -Selectride and hydrolysis of the 15β-acetate gave 3. Finally, hydrogenation of 4 gave 15β,17α-dihydroxy-5β-pregnan-3,20-dione (10) which on reduction with -Selectride gave 8.     

Tetrachloroplatinate(II) and hexachloroplatinate(IV) salts of N,N,N′,N′-tetramethylethylenediammonium: significant differences in the conformation of the cation, as shown by infrared and x-ray diffraction data

The structures of (H₂tmen)[PtCl₄], (1), (H₂tmen = N,N,N′,N′-tetramethylethylenediammonium), [triclinic, P ; A = 7.344(3), B = 8.345(3), C = 6.216(2) Å, α = 84.53(3), β = 109.22(3), γ = 69.43(3)°, Z = 1] and (H₂tmen)[PtCl₆], (2), [monoclinic, P2₁/a; A = 14.409(4), B = 12.736(7), C = 8.601(3) Å, β = 99.58(3)°, Z = 4] were determined from diffractometric data by Patterson and Fourier methods and refined by full-matrix least-squares to R = 0.027 and 0.039 for (1) and (2) respectively. In both cases the anions and cations are joined in polymeric chains through hydrogen bonds involving the protonated nitrogens and the co-ordinated chlorine atoms. The square-planar [PtCl₄]²⁻ and octahedral [PtCl₆]²⁻ anions are centrosymmetric; the H₂tmen²⁺ cations are centrosymmetric in (1) with a N-C-C-N dihedral angle of exactly 180°, while in (2) the dihedral angle is 166°. The different symmetry of the organic moieties/C_i and C₁ in (1) and (2) respectively] results in a different i.r. spectrum which is more complex the lower the symmetry; moreover the spectrum of a KBr pellet of (2) changes with time, finally resembling that of (1) and of the (H₂tmen)Cl₂ salt.     

The Involvement of PI3K-Mediated and L-VGCC-Gated Transient Ca2+ Influx in 17β-Estradiol-Mediated Protection of Retinal Cells from H2O2-Induced Apoptosis with Ca2+ Overload

Intracellular calcium concentration ([Ca²⁺]_i) plays an important role in regulating most cellular processes, including apoptosis and survival, but its alterations are different and complicated under diverse conditions. In this study, we focused on the [Ca²⁺]_i and its control mechanisms in process of hydrogen peroxide (H₂O₂)-induced apoptosis of primary cultured Sprague-Dawley (SD) rat retinal cells and 17β-estradiol (βE2) anti-apoptosis. Fluo-3AM was used as a Ca²⁺ indicator to detect [Ca²⁺]_i through fluorescence-activated cell sorting (FACS), cell viability was assayed using MTT assay, and apoptosis was marked by Hoechst 33342 and annexin V/Propidium Iodide staining. Besides, PI3K activity was detected by Western blotting. Results showed: a) 100 μM H₂O₂-induced retinal cell apoptosis occurred at 4 h after H₂O₂ stress and increased in a time-dependent manner, but [Ca²⁺]_i increased earlier at 2 h, sustained to 12 h, and then recovered at 24 h after H₂O₂ stress; b) 10 μM βE2 treatment for 0.5-24 hrs increased cell viability by transiently increasing [Ca²⁺]_i, which appeared only at 0.5 h after βE2 application; c) increased [Ca²⁺]_i under 100 µM H₂O₂ treatment for 2 hrs or 10 µM βE2 treatment for 0.5 hrs was, at least partly, due to extracellular Ca²⁺ stores; d) importantly, the transiently increased [Ca²⁺]_i induced by 10 µM βE2 treatment for 0.5 hrs was mediated by the phosphatidylinositol-3-kinase (PI3K) and gated by the L-type voltage-gated Ca²⁺ channels (L-VGCC), but the increased [Ca²⁺]_i induced by 100 µM H₂O₂ treatment for 2 hrs was not affected; and e) pretreatment with 10 µM βE2 for 0.5 hrs effectively protected retinal cells from apoptosis induced by 100 µM H₂O₂, which was also associated with its transient [Ca²⁺]_i increase through L-VGCC and PI3K pathway. These findings will lead to better understanding of the mechanisms of βE2-mediated retinal protection and to exploration of the novel therapeutic strategies for retina degeneration.     

Bicarbonate-Chloride Exchange in Erythrocyte Suspensions: Stopped-Flow pH Electrode Measurements

A pH-sensitive glass electrode was used in a temperature-controlled stopped-flow rapid reaction apparatus to determine rates of pH equilibration in red cell suspensions. The apparatus requires less than 2 ml of reactants. The electrode is insensitive to pressure and flow variations, and has a response time of < 5 ms. A 20% suspension of washed fresh human erythrocytes in saline at pH 7.7 containing NaHCO₃ and extracellular carbonic anhydrase is mixed with an equal volume of 30 mM phosphate buffer at pH 6.7. Within a few milliseconds after mixing, extracellular HCO₃^- reacts with H⁺ to form CO₂, which enters the red cells and rehydrates to form HCO₃^-, producing an electrochemical potential gradient for HCO₃^- from inside to outside the cells. HCO₃^- then leaves the cells in exchange for Cl^-, and extracellular pH increases as the HCO₃^- flowing out of the cells reacts with H⁺. Flux of HCO₃^- is calculated from the dpH/dt during HCO₃^--Cl^- exchange, and a velocity constant is computed from the flux and the calculated intracellular and extracellular [HCO₃^-]. The activation energy for the exchange process is 18.6 kcal/mol between 5°C and 17°C (transition temperature), and 11.4 kcal/mol from 17°C to 40°C. The activation energies and transition temperature are not significantly altered in the presence of a potent anion exchange inhibitor (SITS), although the fluxes are markedly decreased. These findings suggest that the rate-limiting step in red cell anion exchange changes at 17°C, either because of an alteration in the nature of the transport site or because of a transition in the physical state of membrane lipids affecting protein-lipid interactions.     

Synthesis and characterization of yttrium complexes bearing a bulky arylamido ancillary ligand

Reaction of anhydrous YCl₃ with 1 equiv. of arylamido lithium 2,6-ⁱPr₂C₆H₃NSiⁱPr₃Li in THF gave an anionic mono-arylamido-ligated yttrium dichloride complex {[2,6-ⁱPr₂C₆H₃NSiⁱPr₃]YCl₂(THF)}₂[LiCl(THF)₂] (1). Alkylation of 1 with 4 equiv. of LiCH₂SiMe₃ afforded an anionic arylamido-ligated yttrium tris(alkyl) complex [2,6-ⁱPr₂C₆H₃NSiⁱPr₃]Y(CH₂SiMe₃)₃Li(THF)₂ (2). Both complexes were characterized by NMR, elementary analysis, and X-ray structural determination.