Solubilization of a mammalian β-adrenergic receptor

Summary Binding sites for iodohydroxybenzylpindolol with characteristics of a ₂-adrenergic receptor have been identified in a crude membrane fraction from guinea-pig lung. The binding sites could be solubilized by treatment of the membrane fraction with digitonin. Upon solubilization receptors retain their ₂-adrenergic type as indicated by the rank order of potencies of agonists in binding-inhibition experiments. The solubilized receptors demonstrate a marked increase in affinity for agonists compared with particulate receptors whereas antagonist affinity remains unchanged. Solubilized receptors are insensitive to divalent cations (Mg²⁺, Mn²⁺, Ca²⁺) which increase the potency of agonists for particulate receptors. The effects of Mg²⁺ can be reversed by Gpp(NH)p in particulate preparations; Gpp(NH)p is ineffective for the solubilized preparation. These experiments establish that -adrenergic receptors can be solubilized even from crude mammalian membrane preparations. They also show that the mammalian -adrenergic receptor in situ is under constraints with respect to agonist affinity and is modulated by divalent cations and guanyl nucleotides in the intact membrane.with technical assistance of L. Braun and C. KonradThis report is part of a dissertation to be presented by J. K. to the Fachbereich Humanmedizin, Justus Liebig-Universität Giessen, in partial fulfillment of the requirements for a Doctor of Medical Science degree     

微米黄连的溶出性研究

目的：研究黄连3种微粒的溶出性。方法：选用3因素(粒径、加水量、溶出时间)、3水平正交设计试验，用苦味酸离子对萃取法测定黄连中的季铵碱溶出量。结果：粒径6.4～57.0 μm的微粉，5 min，45倍水量溶出性能最佳，粒径＜6.4 μm的微粉溶出性较差。结论：粒径6.4～57.0 μm的黄连微粉直接应用于临床具有重要意义。     

胆盐/磷脂混合胶束对疏水性天然药物增溶性能的研究

目的:研究胆盐/磷脂混合胶束(BS/PC-MM)体系对疏水性天然药物的增溶能力及影响因素。方法:选择不同纯度的大豆磷脂(SPC)、蛋黄磷脂(EPC)与高纯度的胆酸盐制备胆盐/磷脂混合胶束(BS/PC-MM)。以透光率方法测定胆酸盐对磷脂的溶解能力,以20(S)人参皂苷Rg3、Rh2、黄芪甲苷、20(S)原人参二醇、原人参三醇和呋喃二烯为疏水性模型药物,考察了SDC/SPC-MM对药物的增溶特性。结果:高纯度的大豆磷脂和蛋黄磷脂适合制备成澄明的BS/PC-MM溶液。胆酸盐对磷脂的增溶能力为,脱氧胆酸钠>胆酸钠,胆酸盐对蛋黄磷脂的溶解度增加略高于大豆磷脂。20(S)原人参二醇、原人参三醇和呋喃二烯与SPC的亲和性较高。而20(S)人参皂苷Rh2、Rg3和黄芪甲苷与SDC的亲和性强,SDC/SPC-MM的脂质总质量增加,疏水性药物的溶解度提高。结论:胆盐/磷脂混合胶束(BS/PC-MM)对疏水性药物具有较好的溶解度,增溶能力受药物的理化性质和混合胶束的组成影响,可成为疏水性药物的非肠道给药新型载体。     

Solubilization of liposomes by weak electrolyte drugs II. Cefotaxime

The solubilization of dimyristoylphosphatidylcholine (DMPC) liposomes by the weak electrolyte drug, cefotaxime (CFX), has been studied as a function of pH, DMPC, temperature, presence of cholesterol (CHOL), and method of liposome preparation. At 7.5mMCFX the lag time for solubilization increased, the rate of solubilization decreased, and the minimum turbidity reached increased as a function of DMPC at pH 1.0 and 40C. Solubilization was most pronounced at pHs below the pKa but inhibited at least one pH unit above the pKa. The critical mole ratio of unionized CFX:DMPC, Rec, for solubilization was estimated to be 0.12. Reducing the temperature slowed the rate of solubilization as did the addition of CHOL. Encapsulation of CFX in liposomes did not significantly reduce CFX degradation, k1=0.048h-1 at 40C and a complex of DMPC and a degradation product of CFX precipitated as rectangular crystals. As a result, an increase in the turbidity of solubilized systems was observed from about 20h to 48h depending on the conditions. Liposomes in the gel state or with at least 20% CHOL did not undergo an apparent reversal of solubilization. It is concluded that the inclusion of weak electrolyte drugs existing predominantly as the unionized species in liquid crystalline state liposomes may undergo a slow solubilization process not necessarily recognized during characterization.     

Beta-cyclodextrin as a suitable solubilizing agent for in situ absorption study of poorly water-soluble drugs

To evaluate the intestinal permeability of poorly water-soluble compounds, it is of importance to completely dissolve them in a medium and to avoid precipitation during experiments. This study was undertaken to find an agent possessing a high-solubilizing capacity and exhibiting minimal modulating impact on membrane integrity and absorption systems such as passive diffusion and carrier-mediated permeation. Phenytoin dissolution was compared in the presence of seven solubilizing agents at concentrations of 1, 2, or 5% using a centrifugation method. The capacity to dissolve phenytoin was great in β-cyclodextrin (β-CD) and hydroxypropyl β-cyclodextrin, followed by Tween 80. Those of methanol, dimethyl sulfoxide, dimethyl acetoamide, and polyethylene glycol 400 were much lower than expected. One percent β-CD did not alter the absorption of fluorescein isothiocyanate-dextran 4000 or the release of protein and lactate dehydrogenase into in situ loop contents, suggesting that 1% β-CD had no significant impact on the integrity of the intestinal membrane. One percent β-CD also did not alter the absorption of caffeine, ceftibuten, or rhodamine 123 from in situ jejunal loops, indicating no interference with passive diffusion and active transports mediated by a peptide transporter and P-glycoprotein. In conclusion, 1% β-CD is a suitable solubilizing agent for evaluating in situ intestinal absorption of poorly water-soluble compounds.     

A polymeric micellar carrier for the solubilization of biphenyl dimethyl dicarboxylate

A polymeric micelle drug delivery system was developed to enhance the solubility of poorly-water soluble drug, biphenyl dimethyl dicarboxylate, DDB. The block copolymers consisting of poly(D,L-lactide) (PLA) as the hydrophobic segment and methoxy poly(ethylene glycol) (mPEG) as the hydrophilic segment were synthesized and characterized by NMR, DSC and MALDI-TOF mass spectroscopy. The size of the polymeric micelles measured by dynamic light scattering showed a narrow monodisperse size distribution with the average diameter less than 50 nm. The MW of mPEG-PLA, 3000 (MW of mPEG, 2 K; MW of PLA, 1 K), and the presence of hydrophilic and hydrophobic segments on the polymeric micelles were confirmed by MALDI-TOF mass spectroscopy and NMR, respectively. Polymeric micelle solutions of DDB were prepared by three different methods, i.e. the matrix method, emulsion method and dialy-sis method. In the matrix method, DDB solubility was reached to 13.29 mg/mL. The mPEG-PLA 2K-1 K micelle system was compared with the poloxamer 407 micelle system for their critical micelle concentration, micelle size, solubilizing capacity, stability in dilution and physical state. DDB loaded-polymeric micelles prepared by the matrix method showed a significantly increased aqueous solubility (>5000 fold over intrinsic solubility) and were found to be superior to the poloxamer 407 micelles as a drug carrier.     

Design of fenofibrate microemulsion for improved bioavailability

The objective of the present study was to formulate a microemulsion system for oral administration to improve the solubility and bioavailability of fenofibrate. Various formulations were prepared using different ratios of oils, surfactants and co-surfactants (S&CoS). Pseudo-ternary phase diagrams were constructed to evaluate the microemulsification existence area. The formulations were characterized by solubility of the drug in the vehicles, mean droplet size, and drug content. The stability was also investigated by store for 3 months under 4 °C, 25 °C and 40 °C and diluted 100 times for 3 days. The optimal formulation consists of 25% Capryol 90, 27.75% Cremophore EL, 9.25% Transcutol P and 38% water (w/w), with a maximum solubility of fenofibrate up to ∼40.96 mg/mL. The microemulsion was physicochemical stable and mean droplet size was about 32.5-41.7 nm. The pharmacokinetic study was performed in dogs and compared with Lipanthy^® capsule. The result showed that microemulsion has significantly increased the C_max and AUC compared to that of Lipanthy^® capsule (p < 0.05). The oral bioavailability of fenofibrate microemulsions (FEN-MEs) in ME-3 and ME-4 were 1.63 and 1.30-fold higher than that of the capsule. Our results indicated that the microemulsions could be used as an effective formulation for enhancing the oral bioavailability of fenofibrate.     

The competition of (−)-[3H]nicotine binding by the enantiomers of nicotine,nornicotine and anatoxin-a in membranes and solubilized preparations of different brain regions of rat

Summary In order to characterize the properties of nicotinic acetylcholine receptor (nAChR) subtypes in the CNS, the enantiomers of nicotine, nornicotine and anatoxin-a were studied for their ability to displace (–)-[³H]nicotine binding to membranes and solubilized preparations of different brain regions of rats. In hippocampal membranes, (–)-[³H]nicotine binding was stereo-selectively displaced from two sites by (+)- and (–)-nicotine, as well as by (+)- and (–)-anatoxin-a. (–)-Nicotine displayed a larger proportion of high affinity binding sites than did (+)-nicotine, while the proportions of high and low affinity binding sites for (+)-anatoxin-a was the same as that for (–)-anatoxin-a. In cerebellar membranes, the (–)-[³H]nicotine binding was stereoselectively displaced from a single binding site by nicotine and anatoxin-a with Ki values that did not correspond with their K_H and K_L values observed in hippocampus. The (–)-[³H]-nicotine binding was displaced from a single site by both (+)- and (–)-nornicotine with similar K_i values in both hippocampal and cerebellar membranes. In Triton X-100 solubilized preparations, the (–)-[³H]nicotine binding was displaced from a single site by all of the drugs tested and the Ki values for each individual drug were similar in the cortex, hippocampus and cerebellum. These results provided further evidence for pharmacological heterogeneity of membrane bound nAChRs and clearly indicated that detergent solubilization changed the binding properties of nAChRs in rat brain.Correspondence to X. Zhang at the above address     

聚氧乙烯蓖麻油C-125及吐温80对维生素D2的增溶与稳定性的研究

维生素D₂的表观饱和溶解度与增溶剂的浓度呈直线关系,聚氧乙烯蓖麻油C-125的增溶效能,约相当吐温80的4倍多。紫外吸收光谱法推测在聚氧乙烯蓖麻油C-125胶团中维生素D₂分子更近于聚氧乙烯基的栅层部位,这可能是使增溶效能不同的1个重要原因。用初均速法测定了维生素D₂的稳定性,在聚氧乙烯蓖麻油C-125溶液中t_0.8^20°为1294.9 h,在吐温80中为153.5 h,反应活化能测得分别为14.2及9.3 kcal/mol,因而使维生素D₂的稳定性呈现差异。     

Cyclothiazide decreases [HAMPA binding to rat brain membranes: evidence that AMPA receptor desensitization increases agonist affinity

The effects of cyclothiazide, a drug which blocks AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor desensitization, were tested on binding of [³HAMPA to rat brain membranes. Cyclothiazide reduced [³HAMPA binding by lowering the apparent affinity of the AMPA receptor. The magnitude of the decrease was temperature dependent and greater for membrane-bound than for solubilized receptors. These data provide evidence that desensitization increases the affinity of the AMPA receptor for agonists and indicate that a significant percentage of AMPA receptors in conventional equilibrium binding assays are in a desensitized state.