Preparation, characterization and uptake by primary cultured rat hepatocytes of liposomes surface-modified with glycyrrhetinic acid.

3-succinyl-30-stearyl glycyrrhetinic acid (Suc-GLAOSt) was synthesized as a targeting molecule, and incorporated in ordinary to liposomes (LP) to prepare a liposome surface-modified with glycyrrhetinic acid (LP-GLA), which could bind to the hepatocyte through the specific binding site of glycyrrhetinic acid (GLA) on the surface of rat cellular membrane. The maximal molar ratio of Suc-GLAOSt to total lipids in LP-GLA was 1:10. Calcein loaded liposome (Cal-LP) and calcein loaded LP-GLA (Cal-LP-GLA) were prepared by an ethanol injection method. The average diameter of Cal-LP and Cal-LP-GLA was 65 nm +/- 16 nm and 68 nm +/- 21 nm, respectively. The characteristics of cellular uptake of the two types of liposome were investigated through cellular uptake and competitive inhibition experiments. The uptake of Cal-LP-GLA by rat hepatocytes was markedly higher (3.3-fold) than that of Cal-LP (P < 0.01). The uptake of Cal-LP-GLA was inhibited, but the uptake of Cal-LP was not influenced by adding extraneous GLA. LP-GLA may be internalized by hepatocytes via the specific binding site, and can be used as a novel and promising carrier for targeting drug delivery to hepatocytes.

[Transdermal and lymph targeting transfersomes of vincristine].

Vincristine (VCR) is mainly used to treat acute lymphocytic leukemia, Hodgkin and non-Hodgkin lymphoma in clinic with definite therapeutic effect. But the obvious neurotoxicity and local stimulation of which limit its clinic use. In order to increase the lymph targeting to enhance the curative effect and to lower the adverse reaction of VCR, the VCR loaded transfersomes (VCR-T) were prepared with dry-film and ultrasonic dispersing methods, and the corresponding pharmaceutical properties, pharmacokinetical characteristics and the targeting ability were studied. The average particle size of VCR-T prepared was 63 nm with an entrapment ratio of 59%. The in vitro transdermal research with modified Franz cell showed that VCR-T permeated through the skin in accordance with polynomial equation, and with an accumulation permeation percentage of 67.4% up to 12 h. An HPLC method was utilized to determine the pharmacokinetics and tissue distribution of VCR. Compared with the iv injection of VCR solution, the retention time of VCR in blood was extended by 12 times with VCR-T, and the targeting index in rat lymph was increased by 2.75 times. As a result, transfersomes could penetrate the skin and enter into the systemic circulation carrying VCR with good lymph targeting ability, which makes it probably a new lymphtic targeting drug delivery system.

[Comparison of preparing two polylactide nanoparticles loaded lipophilic anti-cancer herb drug by nanoprecipitation method].

This paper introduced an experimental study of polylacticacid (PLA) nanoparticles of lipophilic anti-cancer herb drug using a precipitation method. Cucurbitacins (Cu) and Curcuminoids (Cur) were selected to be model drugs. They had similar solubility but their incorporation effects were significantly different: the average drug entrapment ratio, the average drug loading and the average drug recovery were 38.53%, 2.21% and 27.02% respectively; while those of Cur-PLA-NP were 94.36%, 14.35% and 91.23% respectively. To analyse the reason, drug incorporation process was investigated. By measuring solvent evaporation rate, ratio of drug PLA precipitates, drug distribution in system and entrapping ratio at different time of preparation, we found the difference of precipitation velocity of drug was the main reason. We also concluded that not all lipophilic drug can be well entrapped into PLA nanoparticle by nanoprecipitation method. The drug incorporation depended on the interations among drug, PLA and organic solvents, in addition to the solubility of the drug.

[Comparison of two preparation methods applied in tanshinone II(A)-loaded PLGA nanoparticles].

OBJECTIVE: To optimize formulation of tanshinone II(A)-loaded PLGA nanoparticles and compare the difference of two methods in preparation and quality of nanoparticles. METHOD: The two methods were nanoprecipitation method and emulsion-evaporation method. Single factor experiments and central composite design and response surface method were used to optimize the formulation of nanoparticles. The nanoparticles were characterized at size, morphology, entrapment efficiency, drug loading, drug recovery rate, crystallinity and drug release in vitro. RESULT: The mean diameters were 225 nm and 183 nm, the entrapment efficiency were 95.49% and 87.99%, the drug loading were 2.03% and 0.16%, and the drug recovery rates were 38.42% and 17.59% respectively for nanoprecipitation method and emulsion-evaporation method. CONCLUSION: Nanoprecipitation method was better than emulsion-evaporation method for preparation of tanshinone II(A)-loaded PLGA nanoparticles.

[Preparation of liposome entrapped vincristine sulfate and mitoxantrone chlorhydric acid].

OBJECTIVE: To investigate the prepation of the liposome carried with vincristine sulfate (VCR) and mitoxantrone chlorhydric acid (MTO) and to evaluate the quality of the liposome. METHOD: The liposome carried with VCR and MIT was prepared by pH-gradients method and reverse evaporation technique. HPLC was employed to determine VCR and MIT entrapping efficiency of liposomal. Laser particle analyzer was applied to determine the size and zeta potential of the liposomes carried with VCR and MTO. RESULT: The mean diameter of the liposome carried with MIT and VCR was 72.22 nm, with the entrapping rate of 95.77% for VCR and 99.53% for MTO. The liposome had perfect shape. CONCLUSION: The liposomes with high entrapping rate and small particle size had been prepared by pH-gradient method and reverse evaporation technique.

[Study on drug release in vitro and rat intestinal absorption of resveratrol nanoliposomes].

OBJECTIVE: To study the release feature of Res-nanoliposomes in vitro and clarify the difference in absorption of Res-nanoliposomes from varied intestinal segments and the absorptive mechanism in vivo. METHOD: Dialytic method was used to determine resveratrol release rate of Res-nanoliposomes in vitro. An in situ rat perfusion method was used to investigate the intestinal absorption of Res-nanoliposomes. RESULT: Resveratrol release from nanoliposomes in vitro fitted the log-normal distribution equation and had a property of sustained release. Compared with other intestinal segments, significantly high percentage of Res-nanoliposomes was absorbed in ileum (P < 0.001). The absorption rate constants (ka) of Res-nanoliposomes in intestine were not significantly different. CONCLUSION: Res-nanoliposomes could sustain to release drug in vitro. The absorption was a first-order process with the passive diffusion mechanism. The Res-nanoliposomes could promote the absorption of Res in rat small intestine.

[Release profile of compound liposomes entrapped with vincristine sulfate and mitoxantrone chlorhydric acid in vitro and their distribution in mice].

AIM: To study on the release profile in vitro and biodistribution in mice of the compound liposomes carried with vincristine sulfate (VCR) and mitoxantrone chlorhydric acid (MTO). METHODS: The release behaviors of the VCR and MTO from compound liposomes were studied in vitro. HPLC was developed for the determination of the contents of VCR and MTO in tissues in mice. RESULTS: The release time of VCR from compound liposome was 24 h and that from free drug (in control solution) was 6 h. The release of MTO from compound liposome was 0.05% after 288 h and release time of MTO from free drug (in control solution) was 12 h. The liposomes and free drugs were injected intravenously at same dose to mice. The elimination half-life time (T 1/2) in plasma of liposomal and free VCR were 0.16 h and 0.14 h, and the AUCs (0 - 48 h) of them were 2.69 (ug x g(-1)) x h and 1.58 (ug x g(-1)) x h, respectively. The elimination half-life times (T 1/2) in plasma of liposomal and free MTO were 21.6 h and 0.05 h and the AUCs (0 - 48 h) of them were 17.06 (ug x g(-1)) x h and 0.42 (ug x g(-1)) x h, respectively. CONCLUSION: The compound liposome with high entrapping efficiency and small particle size could be prepared by pH-gradients method and reverse evaporation technique. Two drugs were sustained-released from the compound liposome. Mice tail intravenous injection of compound liposomes showed that compound liposome prolonged the retention time and improved the concentration of MTO and VCR in the blood circulation system compared to control. In the mean time, compound liposome reduced the concentration of the MTO and VCR in heart, lung, kidney etc. These observations indicated that compound liposome could improve anticancer activity and reduce side effect.

[Preparation of mitoxantrone liposomes and study on its in vivo distribution in rats after transdermal delivery].

OBJECTIVE: To investigate in vivo distribution of the mitoxantrone(MIT) liposomes in SD rats after transdermal delivery. METHODS: The mitoxantrone liposomes were prepared by thin-film method. Particle size, particle distribution and delta-potential of colloid solution were obtained on laser scatterometer; the encapsulation efficiency was measured by membrane diffusion technique. The time courses of mitoxantrone concentration in vivo after the transdermal delivery of mitoxantrone liposomes were measured by HPLC assays and compared with the injection of mitoxantrone solution. RESULTS: The mean diameter of the MIT liposomes was 50.98 nm, with the entrapping efficiency of 100%. The liposomes had perfect shape. A significantly higher amount of mitoxantrone was delivered in cutis after transdermal delivery of mitoxantrone liposomes, compared with the injection of mitoxantrone solution; meanwhile, a significantly lower amount of mitoxantrone was delivered in plasma and other tissues after transdermal delivery of mitoxantrone liposomes, compared with the injection of mitoxantrone solution. CONCLUSION: Transdermal delivery of mitoxantrone liposomes could be a potential therapy for cutaneous malignant melanoma.

[Study on preparation of konjac glucomannan-hydroxypropyl methyl cellulose compression coated tablets for colonic delivery and in vitro release].

OBJECTIVE: Prepare konjac glucomannan-hydroxypropyl methyl cellulose (HPMC) compression coated tablets and study the effects of the formulation, technics and in vitro dissolution condition on drug release behavior to elevate the colon-specific effects of preparation. METHOD: Berberine hydrochloride core tablets were prepared by wet granulation technique and konjac glucomannan-HPMC mixture as the coating layer were used with compression coated technique. The effects of the formulation and technics on drug release behavior were investigated by dissolution test. The erosion of coat layer during dissolution test was investigated. RESULT: Drug almost not released in dissolution medium stimulating gastric and intestinal condition, and released completely by coating layer erosion and rupture by enzyme in stimulating colonic condition. Drug release decreased with decreasing the ratio of konjac glucomannan-HPMC and increasing coat weight (P < 0.05), compression force was not found to be a significant factor on drug release. Drug release increased with increasing the concentration of beta-mannase in dissolution medium (P < 0.05), rotation speed has no effect on drug release. The release of drug was correlative with erosion of coat layer. The mechanism of drug release were diffusion and erosion. CONCLUSION: The konjac glucomannan-HPMC compression coated tablets was a promising delivery system for drugs to be delivered to the colon.

[Studies on characteristics of absorption and separation of traditional Chinese medicine compound prescription by macroporous resin].

OBJECTIVE: Study the characteristics of absorption and separation of traditional Chinese medicine compound prescription using macroporous resin. METHOD: Study the techniquecs and characteristics of absorption and separation of a sample by macroporous resin, which is composed of coptis root, rhubarb and common anemarrhena rhizome, containing alkaloid, anthraquinone and saponin. RESULT: It is proved by qualitative and quantitative researches studies that after absorbed and separated by optimized technics process, most prime effective components or section fractions in traditional Chinese medicine compound prescription can be reserved maintained. CONCLUSION: If the techniquecs of separation is properly designed, the same kind of macropore resin can absorbd and separate various effective components or section in traditional Chinese medicine compound prescription which have with different chemical structures efficiently.

[Studies on purification of the extract of fructus Gardeniae for injection by macroreticular resins].

OBJECTIVE: To purify the extract of fructus gardeniae for injection by macroreticular resins in purification process of traditional Chinese medicine (TCM) Injections. METHOD: Using fructus gardeniae as sample, on base of obtaining the extract by employing macroreticular resin, quality evaluation and rationality of purification methods had been studied by the quantitative analysis of active ingredients and the characteristics of micromeritics, safety and stability of the extract. RESULT: The experiment showed the extract of fructus gardeniae for injection had been produced successfully by macroreticular resin. CONCLUSION: Using macroreticular resins is a promising purification way of TCM injections, whereas a more consummate method of quality evaluation must be established to ensure safety, efficiency and stability of the preparation in the process.

[The preparation of the polylacticacid nanoparticles of cucurbitacin and their drug loading].

This paper introduced an experimental study of the preparation of polylacticacid (PLA) nanoparticles of cucurbitacin (CuC) using a precipitation method. The residual acetone, ratio of CuC PLA precipitates, and the relationships between the ratios of two precipitates and drug incorporation rates were measured. It appeared that the nanoparticles with 60% of PLA incorporated with 5.5% of CuC were formed when acetone was injected into the aqueous phase. As the acetone gradually evaporated, drug incorporation/encapsulation continued, with most of CuC (about 70%) formed new crystalline cores and suspended in the form of microcrystals in the medium, resulting a suspension containing both nanoparticles and microcrystals. We also concluded that this system may not necessarily be suitable for all lipophilic drugs to be prepared to PLA nanoparticles with good incorporation rate. The drug incorporation depended on the interactions among drug, PLA, and organic solvents, in addition to the solubility of the drug.

[Advances in the study of vincristine: an anticancer ingredient from Catharanthus roseus].

Vincristine is a dimer-indo-alkaloid which is extracted from the leaves of Catharanthus roseus. It is effective to treat acute lymphocytic cell leukemia, Hodgkin disease and non-Hodgkin disease clinically. But the severe side effects, such as neurotoxic and tissue damage, limit its application. In this paper, we summarize physical, chemical, pharmacological and pharmacokinetical properties of VCR and advances in decreasing its side effects. In clinic, association with other medication is adopted. In pharmaceutics, people adopt some new methods and technology such as conjugation with the antibody, encapsulation in liposomes or controlled release films.

[Preparation of liposomes surface-modified with glycyrrhetinic acid targeting to hepatocytes].

OBJECTIVE: To study the preparation of liposomes surface-modified with glycyrrhetinic acid targeting to hepatocytes. METHOD: 3-succinic-30-stearyl glycyrrhetinic acid(Suc-GAOSt), one of the amphiphilic glycyrrhetinic acid derivatives, was synthesized as targeting molecules, liposomes surface-modified with glycyrrhetinic acid has been produced with ethanol injection method. RESULT: Targeting molecules can be mixed into the liposomal membrane. It was confirmed that the targeting molecules is 9% of the total lipids at the most in the liposomes. CONCLUSION: Liposomes surface-modified with glycyrrhetinic acid was successfully prepared, which is considered to be a potential approach targeting to hepatocytes.

[Study on quality evaluation methods of pretreatment for polystyrene-type macroporous absorbing resins].

OBJECTIVE: To establish a GC method using wide bore open tubular columns f or controlling pretreatment for polystyrene-type macroporous absorbing resins and its eligible standard. METHOD: A model macroporous absorbing resins made in our lab was eluted by anhydrous alcohol. The residual solvents in elution (xylene, styrene, diethylbenzene, divinybenzene, decane) were assayed by GC. When the residual solvents can not be detected, the pretreatment is eligible. Compare this method with the methods referring in the literatures. RESULT: When using this method to control the pretreatment, the resins need to be elute 11 times with anhydrous alcocol. When using "adding several times water in the alcohol elution will not be turbid" to control the pretreatment, the resins need to be elute 3 times with anhydrous alcocol. When using "the elution have no UV absorption between the wavelength 200-400 nm" to control the pretreatment, the resins need to be elute over 20 times with anhydrous alcocol. CONCLUSION: The method is simple and feasible.

Development of PLGA nanoparticles simultaneously loaded with vincristine and verapamil for treatment of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the malignant tumors with poor chemo-sensitivity to vincristine sulfate (VCR) due to multi-drug resistance (MDR). Combinations of encapsulated VCR and verapamil hydrochloride (VRP, a chemo-sensitizer) might be a potential strategy to improve HCC therapeutic efficacy of VCR. PLGA nanoparticles (PLGANPs) simultaneously loaded with VCR and VRP (CVn) were prepared. The entrapment efficiencies of VCR and VRP were 70.92 ± 3.78% and 85.78 ± 3.23%, respectively (n = 3). The HCC therapeutic activity of CVn was assessed using MTT assay. In BEL7402 and BEL7402/5-FU human hepatocarcinoma cell lines, CVn had the same antitumor effect as one free drug/another agent-loaded PLGANPs (C + Vn or Cn + V) combination and coadministration of two single-agent-loaded PLGANPs (Cn + Vn), which was slightly higher than that of the free VCR/VRP combination (C - V). CVn might cause lower normal tissue drug toxicity by the enhanced permeation and retention effect in vivo. Additionally, CVn might cause fewer drug-drug interaction and be the most potential formulation to simultaneously deliver VCR and VRP to the target cell in vivo than the other three nanoparticle formulations (C + Vn, Cn + V, and Cn + Vn). Therefore, we speculate that CVn might be the most effective preparation in the treatment of drug-resistant human HCC in vivo.

Reversion of multidrug resistance by co-encapsulation of vincristine and verapamil in PLGA nanoparticles.

Multidrug resistant (MDR) cancer may be treated using combinations of encapsulated cytotoxic drugs and chemosensitizers. To optimize the effectiveness of this combinational approach, poly(d,l-lactide-co-glycolide acid) (PLGA) nanoparticles formulations capable of delivering a cytotoxic drug, vincristine, a chemosensitizer, verapamil, or their combination were prepared via combining O/W emulsion solvent evaporation and salting-out method. Moreover, this work evaluated a number of approaches for the administration of chemosensitizer-cytotoxic drug combinations in a systematic fashion. The results showed that the administration sequence of anticancer drug and chemosensitizer was critical for maximal therapeutic efficacy and the simultaneous administration of vincristine and verapamil could achieve the highest reversal efficacy. In addition, PLGA nanoparticles (PLGANPs) showed moderate MDR reversal activity on MCF-7/ADR cells resistant to vincristine. The dual-agent loaded PLGA nanoparticles system resulted in the similar cytotoxicity to one free drug/another agent loaded PLGANPs combination and co-administration of two single-agent loaded PLGANPs, which was slightly higher than that of the free vincristine/verapamil combination. Co-encapsulation of anticancer drug and chemosensitizer might cause lower normal tissue drug toxicity and fewer drug-drug interactions. Therefore, we speculate that PLGANPs simultaneously loaded with anticancer drug and chemosensitizer might be the most potential formulation in the treatment of drug resistant cancers in vivo.