Micelle formation and drug release behavior of polypeptide graft copolymer and its mixture with polypeptide block copolymer

Self-association behavior of polypeptide graft copolymer and its mixture with polypeptide block copolymer and drug carrier capability of the formed micelles was examined. The results gained through fluorescence spectroscopy, transmission electron microscopy and nuclear magnetic resonance spectroscopy revealed that both polypeptide graft copolymer and its mixture with polypeptide block copolymer can self-assemble to form polymeric micelles in aqueous media. The molecular structure of the graft copolymer and blending the graft with block copolymer exert marked effects on the critical micelle concentration and the shape of formed micelles. It was found that the hydrophobic inner core of the micelles formed either by graft copolymer or mixture of graft and block copolymers can act as an incorporation site for the hydrophobic drugs. The drug loading content of the graft copolymer micelles tends to be larger when the content of the polypeptide segments in the copolymer increases. The results obtained from the drug-release studies showed that the drug-release rates are dependent on the chemical nature of the graft copolymer, the composition of the graft and block copolymer mixture, and also the pH value of the release media.     

Synthesis and in vitro evaluation of lipoamino acid and carbohydrate-modified enkephalins as potential antinociceptive agents

The principal hindrance to drug uptake into central nervous tissue is the blood–brain barrier (BBB). In addition, potential peptide-based neuropharmaceuticals are rapidly destroyed by intra- and extracellular peptidases. In an attempt to address these biological hurdles, a series of lipo-, glyco- and glycolipo-conjugates of Leu-enkephalin have been synthesised via novel solid-phase strategies, and their in vitro activity assessed using mouse vas deferens (MVD) and guinea pig ileum (GPI) assays. Conjugation of a single lipoamino acid onto the C-terminal of the Leu-enkephalin molecule retains biological activity whilst increasing the molecule's overall lipophilicity. Conjugation of a glucuronic acid analogue in an analogous position, however, increases activity 40-fold when compared to the native peptide and induces a high degree of δ-opioid receptor selectivity.     

Physicochemical characterization and toxicity evaluation of steroid-based surfactants designed for solubilization of poorly soluble drugs

To overcome poor water-solubility of new drug candidates, four innovative surfactants based on naturally-occuring hydrophilic and hydrophobic moities were designed and synthesized: cholesteryl-glutamic acid, cholesteryl-poly[N-2-hydroxyethyl-l-glutamine] (PHEG), ursodeoxycholanyl-PHEG (UDCA-PHEG) and ursodeoxycholanyl-poly-l-glutamic acid (UDCA-PGA). Their self-assembling capacity was evaluated using pyrene fluorescence measurements which allow to determine their critical aggregation concentration (CAC). Size measurements were carried out using dynamic light scattering (DLS). Surfactant cytotoxicity was investigated on human umbilical vein endothelial cells (HUVEC) by determining tetrazolium salt (MTT) activity and lactate dehydrogenase (LDH) release. In addition, surfactant haemolytic activity was assessed using rat red blood cells (RBCs). Finally, the ability of these surfactants to solubilize a model poorly soluble drug was quantified. Surfactant self-assembly, cytotoxicity and solubilization properties were compared to those obtained with polysorbate 80, a model solubilizer. Except for cholesteryl-glutamic acid, surfactants were water-soluble. UDCA-PGA was not able to self-assemble or to increase significantly drug solubility. Results showed that cholesteryl-PHEG and UDCA-PHEG were self-assembling with low CAC values (17 and 120μg/ml) into nano-structures with mean diameters of 13 and 250nm, respectively. Cholesteryl-PHEG was the most efficient surfactant in increasing drug solubility (2mg/ml) but exhibited a similar or higher toxicity than polysorbate 80. UDCA-PHEG did not present any cytotoxicity but was far less efficient to solubilize the drug (0.2mg/ml). These results evidence interesting properties of cholesteryl-PHEG and UDCA-PHEG as novel solubilizers.     

Intercellular Lipid Mediators and GPCR Drug Discovery

G-protein-coupled receptors (GPCR) are the largest superfamily of receptors responsible for signaling between cells and tissues, and because they play important physiological roles in homeostasis, they are major drug targets. New technologies have been developed for the identification of new ligands, new GPCR functions, and for drug discovery purposes. In particular, intercellular lipid mediators, such as, lysophosphatidic acid and sphingosine 1-phosphate have attracted much attention for drug discovery and this has resulted in the development of fingolimod (FTY-720) and AM095. The discovery of new intercellular lipid mediators and their GPCRs are discussed from the perspective of drug development. Lipid GPCRs for lysophospholipids, including lysophosphatidylserine, lysophosphatidylinositol, lysophosphatidylcholine, free fatty acids, fatty acid derivatives, and other lipid mediators are reviewed.     

On the toxicity of kynurenic acid in vivo and in vitro

BackgroundKynurenic acid (KYNA), a tryptophan metabolite is an antagonist of ionotropic glutamate receptors and alpha-7 nicotinic receptor. Moreover, it is an agonist of G-protein receptor GPR35. Its neuroprotective, anticonvulsant, anti-inflammatory and antioxidant activity was documented. KYNA is present in food and herbal medicines. However, the data on effects induced by a long-lasting treatment with KYNA is lacking. The aim of the study was the assessment of toxicity of a prolonged administration of KYNA in rodents. The cytotoxicity of KYNA in vitro was also examined.MethodsAdult mice and rats were used. KYNA was administered in the drinking water in concentrations of 25 or 250 mg/L for 3–21 days. The following cells were cultured in an in vitro study: mouse fibroblast (NIH/3T3), green monkey kidney cells and primary chick embryo cells (CECC). Cell viability was determined with methyl thiazol tetrazolium reduction assay, neutral red uptake assay and lactate dehydrogenase leakage assay.ResultsKYNA affected neither body gain nor body composition. Blood counts were also unaffected. The viability of cells in the culture was lowered at high millimolar concentrations of KYNA. An elevated viability of GMK and CECC cells was detected in the presence of KYNA in micromolar concentrations.ConclusionsThe obtained results showed that a long-term application of KYNA in the drinking water is well-tolerated by rodents. No evidence of a toxic response was recorded. Achieved results indicate that diets containing a high amount of KYNA or enriched with KYNA should not cause any risk to the human health.     

壳聚糖/海藻酸钠自组装微球的制备及释药性能

目的利用壳聚糖(CS)聚阳离子及海藻酸钠(ALG)聚阴离子电解质的性质,在药物微球表面自组装形成多层包覆结构的壳聚糖载药微球,并研究组装层数、温度及盐离子浓度对自组装微球释药性能的影响。方法采用乳化交联法制备CS载四环素(TC)的药物微球,并在其表面交替自组装ALG及CS。利用IR测试技术及电极电位法进行表征。结果CS交联微球未破坏CS及TC的结构,CS与ALG以静电作用相结合。CS交联微球的载药量为40.2%,自组装六层的微球载药量为32%。组装后,药物释放时间延长,初期暴释现象得到极大改善,释药速率随组装层数的增加而下降,温度较高时组装完整,盐离子浓度存在较佳点。结论温度为60℃、盐离子浓度为0.5 mol.L-1、组装层数为四层的微球释药性能较佳。     

我国氨基酸类药物研究进展

氨基酸是一类具有特殊生理功能的小分子化合物,近年来我国氨基酸类药物发展迅速,总结市场和研究情况,今后小肽、氨基酸螯合物以及氨基酸衍生物可能成为氨基酸类原料药发展的新方向;而复方氨基酸制剂则可能偏重于含谷氨酰胺的营养输液、肠内营养制剂和多腔袋营养制剂的开发。     

Doxorubicin-conjugated PLA-PEG-Folate based polymeric micelle for tumor-targeted delivery: Synthesis and in vitro evaluation

Background

Selective delivery of anticancer agents to target areas in the body is desirable to minimize the side effects while maximizing the therapeutic efficacy. Anthracycline antibiotics such as doxorubicin (DOX) are widely used for treatment of a wide variety of solid tumors.This study evaluated the potential of a polymeric micellar formulation of doxorubicin as a nanocarrier system for targeted therapy of a folate-receptor positive human ovarian cancer cell in line.

Results

DOX-conjugated targeting and non-targeting micelles prepared by the dialysis method were about 188 and 182 nm in diameter, respectively and their critical micelle concentration was 9.55 μg/ml. The DOX-conjugated micelles exhibited a potent cytotoxicity against SKOV3 human ovarian cancer cells. Moreover, the targeting micelles showed higher cytotoxicity than that of non-targeting ones (IC₅₀ = 4.65 μg/ml vs 13.51 μg/ml).

Conclusion

The prepared micelle is expected to increase the efficacy of DOX against cancer cells and reduce its side effects.     

与肿瘤有关的氨基酸代谢缺陷及其相关药物的应用

氨基酸代谢是生命活动的基础。在肿瘤组织的恶性转化中,由于肿瘤细胞动力学的改变,导致机体氨基酸代谢缺陷的发生。随着代谢组学的发展,氨基酸及相关衍生药物在肿瘤物诊断及治疗中有着广泛的运用,文章概述了与肿瘤有关的氨基酸代谢缺陷的发生,以及氨基酸衍生药物和氨基酸放射标记物在肿瘤诊断和治疗中的应用,为其在医药领域中进一步地发展提供参考依据。     

A role for lipid rafts in the protection afforded by docosahexaenoic acid against ethanol toxicity in primary rat hepatocytes

Previously, we demonstrated that eicosapentaenoic acid enhanced ethanol-induced oxidative stress and cell death in primary rat hepatocytes via an increase in membrane fluidity and lipid raft clustering. In this context, another n-3 polyunsaturated fatty acid, docosahexaenoic acid (DHA), was tested with a special emphasis on physical and chemical alteration of lipid rafts. Pretreatment of hepatocytes with DHA reduced significantly ethanol-induced oxidative stress and cell death. DHA protection could be related to an alteration of lipid rafts. Indeed, rafts exhibited a marked increase in membrane fluidity and packing defects leading to the exclusion of a raft protein marker, flotillin. Furthermore, DHA strongly inhibited disulfide bridge formation, even in control cells, thus suggesting a disruption of protein–protein interactions inside lipid rafts. This particular spatial organization of lipid rafts due to DHA subsequently prevented the ethanol-induced lipid raft clustering. Such a prevention was then responsible for the inhibition of phospholipase C-γ translocation into rafts, and consequently of both lysosome accumulation and elevation in cellular low-molecular-weight iron content, a prooxidant factor. In total, the present study suggests that DHA supplementation could represent a new preventive approach for patients with alcoholic liver disease based upon modulation of the membrane structures.     

熊果酸脂肪乳注射液的制备及体外评价

目的：制备熊果酸脂肪乳注射液,并对其理化性质进行评价。方法：采用高压均质法制备熊果酸脂肪乳注射液,考察脂肪乳的粒径分布、Zeta电位和微观形态,并初步研究熊果酸脂肪乳的稳定性。结果：熊果酸脂肪乳注射液的平均粒径为（203.6±37.1）nm,Zeta电位为（-36.5±3.6）mV;透射电镜显示脂肪乳注射液粒径均一,呈球状分布。稳定性研究结果表明,熊果酸脂肪乳在25℃/60%条件下放置6个月内稳定。结论：高压均质法制备熊果酸脂肪乳注射液工艺简单易行,有望应用于工业化生产。     

药物载体脂肪乳的研究与应用

综述作为药物载体的脂肪乳的分类、制备方法和工艺、存在的问题和解决方法、应用及发展前景。以甘油三酯和磷脂为主要成分的脂肪乳除可用作肠外营养外,近年来其用作药物载体的研究备受关注。     

13-week repeated dose toxicity study of l-tyrosine in rats by daily oral administration

To evaluate the potential toxicity of l-tyrosine, 4 groups of Crl:CD(SD) rats of both sexes were administered l-tyrosine in water suspension by gavage once daily for 13 weeks at doses of 0 (vehicle), 200, 600 or 2000 mg/kg bw/day. Findings related to l-tyrosine administration were as follows. Edema of the cornified layer at the limiting ridge or forestomach was seen in 600 mg/kg bw/day female group and in both sexes of 2000 mg/kg bw/day group. In the liver, increased weight and hypertrophy of centrilobular hepatocytes were seen in both sexes at 2000 mg/kg bw/day, associated with slight increases in ALT and AST. Regarding the kidney morphology and function, increased hyaline droplets in the proximal tubules and increased urinary protein were seen in the 2000 mg/kg bw/day male group. In addition, increased kidney weight was also observed in both sexes of the 2000 mg/kg bw/day group, although the histological changes attributable to the weight increase remained unclear. As for blood chemistry, increases in triglycerides, total cholesterol, phospholipids, potassium ion, calcium, total protein, and α1 globulin were also seen in both sexes at 2000 mg/kg bw/day. Thus, in this study the no-observed-adverse-effect level (NOAEL) of l-tyrosine was considered to be 600 mg/kg bw/day for males and 200 mg/kg bw/day for females.     

Lipophilic methotrexate conjugates with glucose‐lipoamino acid moieties: Synthesis and in vitro antitumor activity

To obtain methotrexate (MTX) derivatives with a balanced hydrolipophilic character, we synthesized a series of conjugates in which the drug was linked to lipoamino acid (LAA)‐glucose residues (LAAG‐MTX). These conjugates displayed increased solubility in polar media compared with the corresponding LAA‐MTX conjugates previously described. In vitro biological testing of LAAG‐MTX indicated that the introduction of the sugar moiety decreased the biological activity of these MTX conjugates. The tetradecyl derivative 6b , however, was effective in inhibiting the dihydrofolate reductase activity in vitro and showed an inhibitory effect on human lymphoblastoid cell growth. Drug Dev. Res. 52:454–461, 2001. © 2001 Wiley‐Liss, Inc.     

Modulating pH-independent release from coated pellets: Effect of coating composition on solubilization processes and drug release

The aim of the study was to clarify the influences of three coating parameters on the drug release from chlorpheniramine maleate (CPM) pellets, coated with blends of poly(vinyl acetate) (PVAc) and poly(vinyl alcohol)-poly(ethylene glycol) (PVA-PEG) graft copolymer. A central composite design was implemented to investigate the effect of the polymer blend ratio, the film coat thickness and the plasticizer concentration on the drug release. The solubilization inside the pellets was monitored by EPR spectroscopy. The blending ratio of both the polymers and the film thickness were found to have a major influence on the drug release and the solubilization speed, in contrast to the plasticizer concentration. A pH-independent release profile was adjustable via modulating the polymer blend ratio and the coating thickness. A mathematical model was developed, providing a good predictability of the release profile, based on the film coat composition. This model offers the possibility to achieve a defined drug-release profile by selective adaptation of the film coat composition, in view of process times, feasibility or polymer costs.     

高偶合巯基壳聚糖的合成和体外评价

目的:用新合成法合成较高巯基含量的壳聚糖巯基醋酸偶合物,并进一步评价其各项性能.方法:用碳化二亚胺和N-羟基磺酸基丁二酰亚胺的活化,提高巯基醋酸与壳聚糖共价偶合率.用重量法考察本产物的溶胀性能,用离体的猪肠黏膜上皮测定膜黏附性,通过溶菌酶实验考察降解性能,并采用单层Caco-2细胞为模版测定14C-甘露糖醇在壳聚糖巯基醋酸偶合物作用下的渗透性能等.结果:本合成法所得巯基壳聚糖的巯基含量可达7.56%,总黏附功是未经修饰聚合物的16.3倍,偶合物的溶胀行为和未经修饰的聚合物在同一范围内,而且高偶合巯基壳聚糖也可被溶菌酶生物降解.结论:巯基含量的增加可提高壳聚糖巯基醋酸偶合物的多项生物学性能.高偶合巯基壳聚糖是新型给药系统中很有前景的功能性敷料.     

Synthesis and in vitro cytotoxic activity on human anaplastic thyroid cancer cells of lipoamino acid conjugates of gemcitabine

Lipophilic derivatives of the antitumor drug gemcitabine (GEM) with the potential for improving drug loading in lipid‐based colloidal carriers, like liposomes or lipid nanoparticles, are described. GEM free base was conjugated to lipoamino acids bearing an alkyl side chain of different length, by either a carbodiimide‐assisted or an ethylchloroformiate‐assisted coupling reaction, to obtain N⁴‐acyl GEM derivatives. These compounds retained the same in vitro cell growth inhibitory activity of the parent drug against two lines of human anaplastic thyroid cancer cells. Stability studies suggested that the observed activity was due mainly to intact derivatives and not to released GEM. Accordingly, these amphiphilic derivatives can be proposed in a further step for the encapsulation in liposomes or lipid nanocarriers, to achieve as a final goal an improvement of the pharmacokinetics and therapeutic activity of GEM. Drug Dev Res 2010. © 2010 Wiley‐Liss, Inc.     

Comparative behavioural toxicity of domoic acid and kainic acid in neonatal rats

Cumulative behavioural toxicity was measured in groups of male and female rat pups (n=6/sex) at different stages of postnatal development. Dose–response curves (DRCs) for toxicity produced by domoic acid (DOM) were generated using animals on postnatal days (PND) 0, 5, 14, and 22, using a behavioural rating scale. In a subsequent experiment, DRCs for toxicity generated by either DOM or kainic acid were produced in rats at PND 8 and 14 for comparison between the two toxins. DOM was found to be a very potent neurotoxin in newborn rats and the potency of DOM progressively decreased with increasing age (interpolated ED₅₀=0.12, 0.15, 0.30, and 1.06 mg/kg at PND 0, 5, 14, and 22, respectively). In addition, the patterns of behavioural expression were found to differ with age. Comparisons between DOM and kainic acid revealed that DOM was approximately six-fold more potent than kainate at both PND 8 and PND 14 and that both toxins were approximately two-fold less potent in PND 14 rats, compared to PND 8. This implies that the mechanism(s) responsible for reduced potency is/are similar between the two compounds. Consistent with previous reports, however, there were both similarities and differences in the observed patterns of behavioural toxicity produced by the two toxins at both ages.     

水飞蓟宾脂肪乳的制备、性质及体外释药研究

目的：制备水飞蓟宾脂肪乳并对其体外释放进行考察。方法：在单因素试验的基础上,采用正交试验设计优化水飞蓟宾脂肪乳处方。采用高压均质法制备水飞蓟宾脂肪乳。结果：脂肪乳平均粒径为（74.42±14） nm,粒径分布系数（ PDI）为0.106,Zeta电位为（-30.2±2.13）mV,pH为6.48±0.04,包封率（91.45±0.0052）%,体外12 h内,累积释放原药的90%左右。结论：本试验获得了较理想的水飞蓟宾脂肪乳,其体外释放具有显著的缓释作用。     

软骨藻酸对H4细胞氨基酸释放和Ca2+浓度的影响

目的　研究软骨藻酸 (domoicacid)对H4细胞的兴奋性、抑制性氨基酸释放和胞内游离Ca2 ( [Ca2 ]i)浓度的影响。方法　应用高效液相色谱 (HPLC)和荧光分光光度计检测 0、0 0 64、0 64和 6 4μmol/L软骨藻酸作用于细胞 2h后的兴奋性、抑制性氨基酸释放和 [Ca2 ]i浓度。结果　天冬氨酸和谷氨酸 :中、高剂量组均高于对照组 ,差异有显著性(P <0 0 5 ) ;甘氨酸 :仅高剂量组高于对照组 ,差异有显著性 (P <0 0 5 ) ;γ 氨基丁酸 :低、中和高剂量组均低于对照组 ,差异有显著性 (P <0 0 5 ) ;兴奋性氨基酸 /抑制性氨基酸 :低、中和高剂量组均高于对照组 ,差异有显著性 (P <0 0 5 )。胞内[Ca2 ]i :低、中和高剂量组分别为 ( 2 0 8 65± 11 0 1)、( 3 42 3 1± 15 0 8)和 ( 5 81 3 6± 17 2 4)nmol/L ,高于对照组 [( 14 3 2 5±11 97)nmo1/L] ,差异有显著性 (P <0 0 1)。兴奋性氨基酸与 [Ca2 ]i浓度有显著的正相关性 (r =0 93 2 0P <0 0 1)。结论　软骨藻酸能引起H4细胞兴奋性和抑制性氨基酸释放和胞内 [Ca2 ]i变化 ,这种变化可能是软骨藻酸兴奋性神经毒性的重要机制