Enhancement In Solubility And Absorption Of Isosteviol Using Cyclodextrin Metal-Organic Framworks

Objective:Isosteviol（STV）has a wide range of biological activities and good pharmacological effects,like anti-cardiovascular disease,hypoglycemic,antifungal,antitumor,antiviral,and also used for pulmonary fibrosis and cancer inhibition.However,STV is insoluble in water,which is the rate-limiting step for drug absorption,affecting bioavailability.Additionally,and it is not suitable for injection,which limits its application to a certain extent.Improving the solubility of the drug is the basis for improving the dissolution and bioavailability of the formulation.Therefore,this study used Cyclodextrin-Metal Organic Frameworks（CD-MOF,refers toγ-CD-MOF）as a carrier to load STV molecules into the nanostructure of superhydrophilic CD-MOF),which had advantages over conventionalγ-CD encapsulation.CD-MOF not only had a high drug loading efficiency,but also had a simple preparation process.Using CD-MOF as supramolecular carrier,the solubility of STV in different media was significantly improved and the oral bioavailability was also improved,laying the foundation for the application of STV which was still in the early stage of exploration.Methods:1.Study of STV-loaded by CD-MOF（STV@CD-MOF）:Firstly,high performance liquid chromatography（HPLC）and high performance liquid chromatography-mass spectrometry（HPLC-MS）methods were established for determinating the content and solubility of STV.Solubilization effects of STV@CD-MOF(n _STV:n _CD=0.5:1,n _STV:n _CD=1:1)and STV@CD in different p H media were investigated with the optimal conditions for loading STV.The in vitro release profiles from different groups were compared simultaneously to determine the dissolution characteristics.2.Study of loading mechanism:Molecular simulation was used to carry out the molecular docking of STV and CD-MOF,and molecular dynamics was used to calculate the possible distribution of STV in CD-MOF.In addition,the field scanning emission electron microscope（SEM）and powder X-ray diffraction（PXRD）were conducted to characterize the morphology and crystallinity of the samples respectively.The Synchrotron radiation-Fourier transform infrared spectroscopy（SR-FTIR）data were obtained using a spectrometer（Nicolet 6700）at the Shanghai Synchrotron Radiation Facility（SSRF）with a range of 400-4000 cm^–1.The nitrogen adsorption-desorption isotherm was used to verify the successful encapsulation of STV into CD-MOF,molecular simulations was conducted to explore the possible encapsulation mechanism between STV and CD-MOF and Differential scanning calorimeter（DSC）was employed under nitrogen purge.3.Pharmacokinetics study of STV@CD-MOF in vivo:To establish a high performance liquid chromatography（HPLC）and high performance liquid chromatography-tandem spectrometry（HPLC-MS）for the determination of the content and solubility of STV.24 male rats were orally administered by STV,STV@CD,STV@CD-MOF（0.5:1）and STV@CD-MOF（1:1）at the same dose of 4 mg/kg.Blood was collected from the orbits at indicated time point to determine the plasma concentration of STV in rats.The pharmacokinetics among STV@CD-MOF（1:1）,STV@CD-MOF（0.5:1）,STV@CD inclusion and STV were compared.Results:1.STV@CD-MOF greatly improved the solubility of STV:The solubility of STV@CD-MOF in p H 1.0,4.5,6.8 and water after optimizing drug loading conditions were 3.4,2.6,3.9 and 24.9 times higher than that of STV@CD inclusion compound respectively.Compared with STV,the solubility in water was increased by nearly154,000 folds,and its solubility at p H 1.0,4.5 and 6.8 was increased by 130,000,120,000 and 549,000 folds,respectively.2.STV@CD-MOF optimized STV releasesignificantly:It can be seen from the release curve that the solubility of STV in the four media was poor,and showed the solubility was p H-dependent.The release effect of STV@CD-MOF was obviously better than that of STV.It was almost completely released within 5 min and had no p H dependence,while the release degree of STV@CD inclusion compound reached 70–80%,which highlighted the advantages of CD-MOF carrier delivery.In the STV@CD-MOF（1:1）sample,STV:CD（0.5:1）was considered to be a CD inclusion,and the other STV:CD（0.5:1）was considered to be a nanocluster mechanism.The cumulation release of STV in p H 6.8 and water was 70-80%,and the cumulation release of STV in p H 1.0 and 4.5 was only half of that in p H 6.8 and water.It was considered that when the p H of STV molecules was 1.0 or 4.5,the presence of nanoclusters may affect its further release.3.The drug loading mechanism of STV@CD-MOF was determined:Molecular simulations showed that the drug molecules were more likely to stay in the bi-CDwhen the molar ratio of STV:CD was 0.5:1.If the STV molecules increased to a molar ratio of STV:CD=1:1,three STV molecules preferentially existed in the large cavity of MOF in the form of nanoclusters with a molar ratio of about 0.5:1 for each six CD units.In STV@CD-MOF,CD-MOF was soluble in water,and there were STV@CD and STV nanoclusters in the solution system.The apparent solubility of STV was significantly increased when STV@CD and nanoclusters released once STV@CD-MOF was dissolved into water,which also increased rate of encapsulation of CD by virtue of increasing contact area.In the STV@CD complex,STV was encapsulated by CD with the molar ratio STV:CD being 1:1.In conclusion,it was considered that cyclodextrin inclusionand ionization synergistically enhanced the solubility of the drug in water,while the CD-MOF markedly augmented the rate of the CD inclusion process described above.4.STV@CD-MOF significantly increased the oral bioavailability of STV:The bioavailability of STV@CD and STV@CD-MOF（0.5:1）in rats were almost the same comparing the AUC after oral administration.The bioavailability of STV@CD-MOF（1:1）group(AUC_（0-24h）=307.25±114.60μg/m L·min)was 8.67-fold higher than that of STV（p<0.001）and about 1.32-fold and 1.27-fold higher than STV@CD and STV@CD-MOF（0.5:1）,indicating that STV@CD-MOF（1:1）was better than CD inclusion in absorption.Conclusions:In order to improve the solubility and bioavailability of STV,STV was loaded into the nanostructure of supramolecular carrier CD-MOF,and STV@CD-MOF（1:1）had obvious advantages over the traditionalγ-CD inclusion.CD-MOF not only had a high drug loading efficiency but also a simple preparation process.Uing CD-MOF as supramolecular carrier,the solubility of STV was significantly increased in different media and released in a non-p H-dependent manner.Notably,STV@CD-MOF（1:1）showed better bioavailability compared to STV@CD.The successful loading of STV and the formation of nanoclusters in CD-MOF contributed to the betterment of solubility and bioavailability,and the drug loading mechanism was explored using molecular simulations and confirmed by in vivo pharmacokinetics in animals.As a new carrier,CD-MOF provides new ideas to the formulation of insoluble drugs.