| Vitexin（VT）is a naturally occurring flavonoid that is widely distributed in nature.VT possesses various pharmacological activities including cardiovascular protection,anticancer,antioxidant,anti-inflammatory and analgesic,however,the disadvantages of VT involve infusibility in water,short half-life,and low bioavailability.Therefore,how to increase the water solubility of VT and improve the pharmacokinetic defects of VT is extraordinarily critical for the clinical application of VT.Objective:The topic initially improves the solubility of VT by nanosizing VT into micronized powder and thereafter establishing an albumin nanoparticle drug delivery system to encapsulate VT,hoping to improve the aqueous solubility of VT and increase its bioavailability.Methods:Firstly,DMSO was used as the main solvent and water as an antisolvent to prepare VT micronized powder by the antisolvent recrystallization method.The physical and chemical properties of the VT micro powders were analyzed by laser particle sizer,Scanning Electron Microscope（SEM）,Fourier transform infrared spectroscopy（FT-IR）,X-ray diffraction（XRD）,and differential scanning calorimetry（DSC）.Subsequently,a method for the determination of VT content in accordance with the relevant requirements of the 2020 version of the Pharmacopoeia of the People’s Republic of China was established.VT-BSA NPs were prepared by the desolventization-cross-linking curing method.Optimal preparation conditions were screened out by single-factor examination and orthogonal test sieve.Characterization of VT-BSA NPs with respect to particle size,Zeta potential,encapsulation efficiency（EE%）,morphological appearance,placement stability,and in vitro release.In the following,the freeze-drying method was used to prepare VT-BSA NPs lyophilized powder,and the appearance,dispersibility,and particle size were selected as evaluation indexes to screen the best lyophilization technology.The properties of VT-BSA NPs lyophilized powder including particle size,Zeta potential,EE%,appearance and morphology,and placement stability were further characterized.Eventually,the blood concentrations of VT API,and VT-BSA NPs in rats after a single intravenous administration were determined by HPLC-MS/MS analysis,the values were fitted to the results using DAS2.0 pharmacokinetic analysis software,and the differences in pharmacokinetic parameters between the groups were compared.Results:1 Preparation and characterization of VT micro powderThe VT micro powder prepared by the antisolvent recrystallization method was nearly spherical with regular morphology,its average particle size was（187.13±7.15）nm with uniform size distribution and its PDI was（0.168±0.008）.The FT-IR spectra of VT micro-powder and VT raw powder are almost identical,indicating that no chemical changes occur in micronized treated VT.The XRD values revealed that the position of diffraction peaks did not change after the micro-pulverization treatment of VT,and only the vibrational intensity was reduced,indicating that micro-pulverization treatment would not change the state of existence of VT,but its crystallinity was reduced.The DSC analysis showed that the melting point of VT decreased from 265.95°C to252.55°C after micro-pulverization treatment,indicating that the crystallinity of VT decreased after micro-pulverization treatment,and VT changed from stable crystallization to metastable crystallization.2 Construction and characterization of VT-BSA NPs drug delivery systemThe VT content determination method is specific and conforms to the relevant requirements of the 2020 version of the Pharmacopoeia of the People’s Republic of China,and the ultrafiltration centrifugation method is effective in separating the free VT from VT-BSA NPs and is suitable for the measurement of the encapsulation efficiency of VT-BSA NPs.The VT-BSA NPs were prepared under optimal conditions with a subspherical appearance and non-adhesive surface,with average particle size,PDI,Zeta potential,and EE%were（124.33±0.47）nm,（0.184±0.012）,（-48.83±2.20）m V,and（83.43±0.39）%,respectively.In addition,the nano-delivery system was able to maintain stability for 15 d at 4°C without significant drug leakage.The in vitro release results indicated that VT-BSA NPs could significantly prolong the administration time of VT with a slow release mechanism.3 Preparation and characterization of lyophilized powder of VT-BSA NPsThe average particle size of VT-BSA lyophilized powder was（200.77±3.86）nm,which was slightly larger than before lyophilization,the PDI was（0.100±0.054）,the Zeta potential was（-41.63±0.40）m V,and the EE%was（80.69±0.17）%.The results of TEM revealed that the VT-BSANPs after lyophilization and re-solubilization were found to be nearly spherical in appearance to the freshly prepared VT-BSA NPs,indicating that the lyophilization protectant played a positive protection effect on VT-BSA NPs.The results of stability determination demonstrated that the prepared VT-BSA NPs lyophilized powder could maintain stability within 60 d when stored at 4°C and room temperature.4 Pharmacokinetic parameters research of VT-BSA NPsThe results of in vivo pharmacokinetic studies showed a significant increase in AUC（0→t）,AUC（0→∞）,MRT（0→t）,and MRT（0→∞）in the VT-BSA NPs group compared to the VT solution group（P<0.001）.The VT solution group was difficult to detect its blood concentration after 2 h.The VT-BSA NPs group was still measurable at 8 h.The half-life of the VT solution group was 0.61 h,while that of the VT-BSA NPs group was 3.09h,a 5.07-fold improvement.These results indicated that the VT-BSA NPs circulated longer in vivo compared to VT injection and that the bioavailability of VT was improved.Conclusions:In this study,VT-BSA NPs,which can improve the disadvantages of poor water solubility,short half-life and low bioavailability of VT,were successfully prepared as a nanoformulation with good application potential. |
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