Liying Wang

22 Improved stability and release behaviors of rhGH in PK3/PLGA microspheres with different PK3/PLGA ratios

Objectives We designed a series of formulations of rhGH PK3/PLGA microspheres with different PK3/PLGA ratios to improve the stability and incomplete release of rhGH with the aim of achieving long-acting sustained-release rhGH microspheres. PK3 can undergo acid-catalyzed hydrolysis into low molecular weight hydrophilic compounds and release encapsulated drugs at an accelerated rate in acidic environments. PK3/PLGA microspheres effectively avoided the influence of the acidic environment produced by the degradation of PLGA on rhGH stability and solved the problem of difficult release of denaturated rhGH. The microspheres improved the poor mechanical properties of PK3 microspheres due to low molecular weights. Methods We used a W/O/W double-emulsion technique to prepare rhGH microspheres. Briefly, 100 µL of 40 mg/mL rhGH was dissolved in 2 mL of acetone/methylene chloride solution (0.8:1.2) containing PK3/PLGA blends and homogenized at 6000 rpm for 120 s in an ice bath. The primary emulsion was injected into an aqueous solution containing 0.5% (w/v) PVA and 3% (w/v) NaCl and homogenized at 6000 rpm for 120 s. Microspheres were obtained by evaporating the organic solvents and centrifuging at 5000 rpm. We investigated the effects of different PK3/PLGA ratios (10:0, 8:2, 6:4, 5:5, 4:6, 2:8, 0:10) and the buffer pH value (pH 4.5, pH 7.4) on the in vitro release of rhGH microspheres. We studied the structural stability of rhGH in release medium and inside the microspheres. Results Native PAGE results revealed the microspheres displaying enhanced structural stability of rhGH with a neutral microenvironment compared with PLGA microspheres. With an increase in PK3 ratios, the microspheres showed reduced incomplete release and increased total release of rhGH. When the PK3/PLGA ratios were above PK3-PLGA mass ratio 5:5, the mixing microspheres had a low balling rate and irregular form. Conclusions The optimal ratio of PK3/PLGA was 3:7, producing significant rhGH stability and an in vitro release profile with less burst release and extended sustained release.

37 Optimization of supercritical carbon dioxide extraction conditions of Semen cassiae volatile oil using response surface methodology and antimicrobial activity detection

Objectives The objective of this project was to study the process of extracting Semen cassia. The volatile components of Semen cassia were analyzed using supercritical CO2 extraction (SCDE). Based on a single factor experiment, response surface methodology was used to investigate the extraction of volatile oil. According to the fitted curves under different conditions, the effects of pressure, temperature and time on extraction and interactions between various factors were determined. Antimicrobial activity was also measured. Methods A Box-Behnken central composite design method was used based on single factor experiments. The influence of extraction temperature, extraction time and extraction pressure on extraction yield was studied. The response surface method was employed to analyze the results of experiments. The disk diffusion method was used to detect the antimicrobial activity of Semen cassiae volatile oil. Results The results indicated that the optimum extraction conditions were as follows: extraction temperature 51°C, extraction time 3.22 hours and extraction pressure 25 MPa. Extraction yield reached 2.34%. The volatile oil of Semen cassiae extracted from the tested strains showed antimicrobial activity, with MIC values ranging from 2.5 to 5 mg/mL.Abstract 37 Figure 1 Responsive surfaces and contours Conclusions SCDE is a stable and efficient process. Semen cassiae volatile oil had antimicrobial activity which could provide a theoretical basis for application of this oil.

16 Discrimination of Brazilian green propolis and Chinese propolis based on high-performance liquid chromatographic fingerprints and multivariate statistical analysis

Objectives The determination of chemical components is usually used in the quality control of propolis. However, chemical components from different types of propolis are similar. The objective of this investigation was to establish a method based on a specific chemical fingerprint profile and a multivariate mixed model statistical analysis which could easily distinguish propolis of different origins and promote the quality control of propolis. Methods A novel approach using high performance liquid chromatography (HPLC) coupled with multivariate statistical analysis was established for profiling and distinguishing Chinese and Brazilian green propolis. A batch of 22 propolis samples was analyzed, and the datasets on retention time, peak area and sample codes were subjected to mixed multivariate statistical analysis consisting of principal component analysis (PCA) and a self-organization mapping net (SOM). Results The fingerprints were profiled. PCA score plots showed Chinese and Brazilian green propolis clearly classified into two groups. The visualized SOM results showed data from the two groups projected to the adjacent neurons clearly separated from each other. Artepillin C, which contributed greatly to the differentiation, was screened out and identified as the reference compound. Artepillin C is the characteristic component in Brazilian propolis which can be used as chemical marker to distinguish propolis of different origins. Conclusions In this study, fingerprints coupled with multivariate statistical analysis have been successfully applied to distinguish Chinese from Brazilian green propolis. The research identified a chemical marker, and thus helps to investigate and promote the quality control of propolis.

46 Sandwich-type ethylcellulose films for controlled release of anti-restenosis drugs

Objectives Restenosis is a response of the vessel wall to balloon-induced injury and is characterized primarily by elastic recoil of the vessel wall and a series of pathological processes including thrombus, inflammation and vascular smooth muscle cell (VSMC) proliferation. To treat restenosis, appropriate drug delivery vehicles are needed which can release therapeutic agents targeting different symptoms into blood vessels in a controlled manner. The main objective of the present study was to prepare sandwich-type ethyl cellulose films with high performance for efficient drug loading and controlled drug release for restenosis treatment. Methods Sandwich-type ethyl cellulose films loaded with probucol for treating coronary artery disease, or aspirin as an antithrombotic drug, were prepared by casting three individual layers in sequence using an ethyl cellulose/toluene solution. On a glass plate, the first ethyl cellulose layer (bottom layer) was cast without drugs, on to which the middle layer containing probucol or aspirin was then cast. After solvent evaporation at room temperature, a third top layer was cast on to the middle layer. The obtained drug-loading films were further dried at room temperature under vacuum. Results The sandwich-type ethyl cellulose films exhibited a drug loading content (DLC) of 12.1 ± 0.9% and a drug loading efficiency (DLE) of 73.5 ± 3.6% for aspirin, and a DLC of 11.0 ± 0.8% and a DLE of 69.3 ± 3.4% for probucol. Under physiological conditions (37°C, pH 7.4), the release half-life of aspirin from the films was 2.7 ± 0.2 hours, while that of probucol was 7.1 ± 0.6 days. The two drugs showed totally different release behaviors, which can be employed in combination to treat restenosis. Conclusion Sandwich-type ethylcellulose films loaded with probucol or aspirin were successfully prepared and showed ability to release the two drugs in different ways: rapid release of aspirin to treat thrombus and inflammation typical of early-stage restenosis, and sustained release of probucol for inhibition of VSMC proliferation frequently seen in the later stage of restenosis. These drug-loaded ethylcellulose films provide new insight into restenosis therapy.

44 Core-shell nanospheres for pH-responsive release of anticancer drugs and near-infrared imaging

Objectives Nanoscaled drug carriers with pH-responsiveness have attracted extensive interest in view of the acidic environment in cancerous cells. Rapid response to pH changes plays a key role in efficient intracellular drug release. In addition, real-time tracking of drug carriers is important for understanding distribution and targeted accumulation of the drug carriers. This work aims at developing silver selenide quantum dots (Ag2Se QDs)@carboxymethyl chitosan (CMCS) core-shell nanospheres with encapsulated paclitaxel (PTX) for cancer therapy and bioimaging. Methods Oleic acid-capping Ag2 Se QDs were synthesized by a one-pot strategy, washed with ethanol, and obtained by centrifugation. The as-synthesized Ag2Se QDs were reacted with N-hydroxysuccinimide and conjugated with CMCS at the amino sites. In an aqueous solution of PTX, the hydrophobic oleoyl groups tended to aggregate locally and entrap PTX by hydrophobic interaction, spontaneously producing Ag2Se QDs (PTX)@CMCS nanospheres. Results By conjugating the oleic acid-capping Ag2 Se QDs with pH-sensitive CMCS at a degree of substitution (DS) of 13%, biocompatible core-shell nanospheres loaded with PTX were successfully prepared, which had an average size of 36.3 ± 0.2 nm. The drug loading content (DLC) and drug loading efficiency (DLE) for the PTX was 5.01 ± 0.8% and 52.4 ± 3.2%, respectively. The PTX release half-life was 4.1 hours under conditions resembling the intracellular environment of cancerous cells (37°C, pH 5.0). Conclusions Core-shell structured Ag2Se QDs (PTX)@CMCS nanospheres capable of releasing PTX in an acidic environment and emitting NIR fluorescence under NIR laser excitation were synthesized and characterized. The hydrophobic oleoyl groups entrapped PTX via hydrophobic interaction and the oleoyl-CMCS chains were extended at lowered pH to release the otherwise encaged drug. In addition, the encapsulated Ag2Se QDs can emit bright NIR fluorescence for bioimaging by which nanosphere distribution in a patient can be monitored. This study provides a new approach for developing nanocomposite drug carriers for cancer therapy.

1 An investigation of micro-hydroxyapatite spheres loaded with indomethacin for targeted drug delivery

Objectives A major obstacle to be overcome for oral drug delivery is that drug absorption/release must be avoided before the drug reaches the target site. In an attempt to overcome this challenge, we selected synthetic hydroxyapatite (HA) as a drug carrier for the hydrophobic drug indomethacin (IDM) used in local treatment of the colon, exploring the loaded drug and in vitro release characteristics of IDM/HA. Methods An easy one-step hydrothermal method was employed to prepare micro-hydroxyapatite (HA) spheres. Infrared spectroscopy, X-ray diffraction and scanning electron microscopy further confirmed the composition, structure and morphology of the obtained sample. In vitro release of the anti-inflammatory drug (IDM) was performed successively under simulated conditions. The loading and release profiles of the drug were analysed by UV-spectrophotometry. Results The use of ethanol enhances solubilization, and the drug loading rate is about 62.21% at 37° C for 12 hours when the mass ratio of IDM/HA is 2:1. The in vitro release of IDM/HA is dependent on solution pH. There is almost no release of IDM (only 0.159%) at pH 1.0, poor release (about 10.55%) at pH 6.8, and a cumulative release rate of up to 84.95% at pH 7.8. These results suggest that HA particles can be used as a pH responsive vehicle for delivering drugs. Conclusions The as-synthesized micro-hydroxyapatite spheres may act as a promising drug delivery system due to their good biocompatibility, pH sensitivity and high drug loading/release efficiency. The IDM/HA drug delivery system satisfies the basic demand of oral site-specific delivery. Acknowledgments This work was financially supported by a college students’ innovative training project (Grant No. 4041906168).