Transdermal Delivery Of Paeonol Using Microemulsion

Paeonol is a representative bioactive compound in root bark of Paeonia suffruticosa Andr. and the herb of Pycnostelma Paniculatum(Bunge)K . Schu. It has the pharmacologic activity of sedative-hypnotic, antipyretic-analgesic, anti- inflammatory, anti-allergic, antibacterial and so on. Paeonol is as the main active ingredients of Chinese herbal compound in the clinical application, but also is administrated in the form of chinese monomer, lasting dosage form is paeonol injections, paeonol tablets and paeonol Ointments. It is used to treat rheumatology, stomach pain and other pains, eczema, atopic dermatitis, has good effect. When paeonol is used for transdermal delivery, it overcomes the disadvantages of traditional Hormone-type drugs when it is used to treat eczema and atopic dermatitis that is big side effects, moreover it has the advantage of low toxic side effects, easy to carry and sustained effect. But it has the disadvantage of low water-soluble, low melting point , volatile, bad stability, which increased the difficulty of selecting transdermal delivery dosage form.As a prospected carrier of transdermal delivery, microemulsion provides a new method for solve the problem which is the trandermal delivery of Water-insoluble drugs. Microemulsion has been used in the study of many transdermal preparations such as lidocaine, indomethacin, triptolide. As the carrier of transdermal delivery, microemulsion has the advantage that is increasing the solubility of lipophilic and hydrophilic drugs, improving the transdermal rate of drugs, increasing the stability ,extending the active time of drugs, maintaining the constant and active plasma concentration. So microemulsion is used in transdermal delivery, that has been the focus of pharmaceutics.In this study, paeonol is prepared for transdermal delivery system, that may be a promising vehicle for the transdermal delivery of paeonol. Though preparing the pseudo-tertiary phase diagrams and the transdermal test, screen the formulation and preparation process, and then prepare the microemulsion-based gel according the optimal formulation, evaluate the transdermal ability of the various systems. Meantime study the appearance, morphology, size distribution and stability of microemulsion. With the LC-MS technique, determine the time-course of paeonol concentrations using the in vivo microdialysis sampling in the skin and blood, and compare the pharmacokinetic parameters.Part one Preliminary studies A sensitive high-performance liquid chromatography assay was established to measure paeonol in vitro. Column: Diamond C18 column (4.6mm×250mm,5μm),Mobile phase: methanol-water (70∶30), Flow rate : 1.0ml/min, Detective wavelength : 273nm. Meanwhile study the apparent oil-water distribution coefficient and the balanced solubility of paeonol.Part two The preparation of paeonol microemulsionThough studing the solubility of paeonol in different oils and the viscosity of oils, IPM is selected as the oil. Though preparing the pseudo-tertiary phase diagrams, screen the variety and component ratio of surfactants and co-surfactants. Though the transdermal test in vitro, screen the optimal formulation, the optimal formulation is IPM-lecithin-APG-1,2- propylene glycol-water(0.58∶0.3∶0.6∶0.45∶3).though optimizing drug loading and preparation process, determine the largest drug loading of paeonol microemulsion is 1%, and preparation process is dropwise technique.Part three Evaluation and stability of paeonol microemulsionA sensitive high-performance liquid chromatography assay was established to measure the content of paeonol microemulsion, the results show the quality of microemulsion meets the requirement. And evaluate its physical and chemical properties such as appearance, morphology, size distribution. The stability is preliminarily studied, the results show the appearance, size distribution and content of microemulsion have no significant change.Part four Preparation of microemulsion-based gel and comparison of transdermal delivery in vitroEstablish the formulation and preparation method of microemulsion-basd gel, and study its physical and chemical properties. Compare the paeonol microemulsion, microemulsion-based gel and paeonol saturated solution in transdermal test in vitro. The steady-state permeation rate of paeonol microemulsion, microemulsion-based gel and paeonol saturated solution are separately 47.846μg·cm^-2·h^-1, 103.760μg·cm^-2·h^-1 and 70.401μg·cm^-2·h^-1, the 12h cumulative permeated amount are 657.179μg·cm^-2, 1266.484μg·cm^-2 and 881.217μg·cm^-2. So the permeated effect of paeonol microemulsion and microemulsion-based gel is better than paeonol saturated solution significantly. Part five Research of pharmacokinetics on transdermal delivery of paeonol using microemulsionWith the LC-MS technique, determine the time-course of paeonol concentrations using the in vivo microdialysis sampling in the skin and blood. The conditions of HPLC technique: column is XTerraR MS C18 column(2.1mm×150 mm,5μm); mobile phase is methanol-water (85∶15); flow rate is 0.3ml/min; column temperature is 40℃; injection volume is 5μL; running time is 3min. The conditions of MS techniques: nebulizer pressure 40psi, drying gas temperature 350oC, flow-rate of drying gas 8 L/min, capillary voltage was 4000v, collision energy is 20 and fragmentor voltage 105eV. MRM of the ion was carried in negative mode and the quanlity ion at m/ z 122 was measured for paeonol. The linear range of paeonol was 5.25-2100ng/mL, intra-day RSD and inter-day RSD are less than 7%.Recovery by loss and by gain is used to study the in vitro recovery of paeonol, recovery by loss is used to study the in vivo recovery of paeonol. In microdialysis in vitro, the recovery detected by gain method is the same as that by loss (retrodialysis) method. And the recovery is independent of the drug concentration surrounding the probe. The microdialysis system of skin is in a mean in vitro recovery of 64.06%±5.04%, the microdialysis system of blood is in a mean in vitro recovery of 30.38%±5.37%. In vivo recovery of paeonol in skin is 69.71%±4.80% and in blood is 51.63%±7.28%. It was suggested that microdialysis sampling could be used for the pharmacokinetic study of paeonol and loss (retrodialysis) method could be used for the determination of recovery of paeonol.The pharmaceutical parameters of unbound paeonol in skin after paeonol microemulsion administrated are shown as AUC_0-660=(334917±10064.2)ng·min/mL, AUC_{0–∞}=(348321.3±9459.8)ng·min/mL, C_max=(3212.5±166.5)ng/mL, Tmax=(60±0.0)min. The pharmaceutical parameters of unbound paeonol in blood after paeonol microemulsion administrated are shown as AUC_0-660=(34368.7±2122.8)ng·min/mL, AUC_{0–∞}=(47809.7±1604.1)ng·min/mL, C_max=(208.6±15.1)ng/mL, T_max=(76.7±5.8)min.The pharmaceutical parameters of unbound paeonol in skin after paeonol microemulsion-based gel administrated are shown as AUC_0-660=(958176.0±49107.1)ng·min/mL, AUC_{0–∞}=(1074505.0±25745.8)ng·min/mL, C_max=(3825.1±826.5)ng/mL, T_max=(240.0±113.1)min. The pharmaceutical parameters of unbound paeonol in blood after paeonol microemulsion-based gel administrated are shown as AUC_0-660=(56142.4±12832.4)ng·min/mL, AUC_{0–∞}=(74385.6±17172.8)ng·min/mL, C_max=(211.6±21.5)ng/mL, T_max=(130.0±14.1)min.The pharmaceutical parameters of unbound paeonol in skin after paeonol ointment administrated are shown as AUC_0-660= ( 84513.5±23977.3 ) ng·min/mL,AUC_{0–∞}=(90674.8±25416.0)ng·min/mL, C_max=(485.1±355.7)ng/mL, T_max=(60.0±30.5)min. The pharmaceutical parameters of unbound paeonol in blood after paeonol ointment administrated are shown as AUC_0-660=( 67200.8±40771.9 ) ng·min/mL, AUC_{0–∞}=(78282.6±36704.6)ng·min/mL, C_max=(285.6±222.6)ng/mL, T_max=(40.0±23.1)min.So we know when microemulsion is administrated, big concentration gradient can rapidly form at both side of the skin, the largest concentration could reach at 60min, the premeated rate can be improved greatly, and the lag time is shortened. But when microemulsion-based gel is administrated, its AUC is more than that of microemulsion during 11h. At a certain extent, the paeonol microemulsion-based gel solves the problem of microemulsion that is low viscosity and loss of dosages, moreover improves the bioavailability.This research provides experimental evidence for the development of transdermal delivery of paeonol using microemulsion, may be provide a new preparation for the clinical treatment of skin Allergic diseases.