| Natural fatty acids such as oleic acid(OA)and linoleic acid,present in various plant oils,are commonly used absorption enhancers.Compared with chemical penetration enhancers,they possess lower toxicity and less skin irritation.Punicic acid(PA),the major constituent of pomegranate seed oil(PSO),occupies more than 60% of total fatty acids in PSO and exists in the principal form of glycerides.PA,linoleic acid and oleic acid belong to C18 acids,containing three,two and one double bond,respectively.For this reason,it is hypothesized that PA may have activity in promoting absorption.Due to the poor stability of PA,PA was ethylated into punicic acid ethyl ester(PAEE)to improve its stability in this study.Grape polyphenols have diverse biological activities,such as antioxidant,anti-inflammatory,antibacterial,and anti-cancer activities.Though they are the principal components responsible for the pharmacological effects of grapes,they have low oral bioavailability.In this study,catechin,quercetin and resveratrol were selected as representative drugs of grape polyphenols,respectively.PSO,PA,and PAEE were combined with three types of grape polyphenols to study their possible absorption-promoting effects,using OA as a control.Nanoemulsions can significantly increase the solubility of poorly soluble drugs and improve their oral bioavailability.Thus,PSO,PA and PAEE were employed as oil phases to prepare grape polyphenol nanoemulsions.The absorption promoting effect of nanoemulsions and oils were compared in the study.The optimization of the enrichment of PA from PSO based on low-temperature solvent crystallization through response surface methodology was performed in this work.PA content of the optimal process was 950.00 mg/g.The purity was 95.39%.The yield was 19.04%.PAEE was synthesized from PA under catalyst by hydrochloric acid.The structure was confirmed by UV,IR,NMR and GC-MS.The refractive index,dielectric constant(ε)and r HLB values of PSO,PA,PAEE and OA were measured,respectively.The antioxidant activity of the oils were determined by measuring the scavenging rates against ABTS and DPPH free radicals.It found that the antioxidant activity of PSO was significantly stronger than that of PA and PAEE(p<0.05).The antibacterial activities of PSO,PA and PAEE against three different bacteria and fungi were determined by disc diffusion method.The results showed that PA had the strongest antibacterial activity,followed by PAEE,and PSO,respectively.Absorption enhancers may change the physiochemical properties of drugs by interacting with drugs.For this consideration,molecular docking technology was conducted to examine the interaction between PA,PAEE,OA and grape polyphenols.The results illustrated that OA and PA interacted with the three grape polyphenols through hydrogen bonds and hydrophobic bonds,and PAEE through hydrophobic bonds.The strength of action was quercetin>resveratrol>catechin.The oils and nanoemulsions were combined with grape polyphenols,respectively,to investigate the influence on the solubility and lipid/water partition coefficient(P)of the drugs.The results demonstrated that PA series(PAs)and OA had almost no effect on the solubility of catechin.However,the four oils can significantly increase the solubility of quercetin and resveratrol(p<0.05).Compared with using oils alone,nanoemulsions had more obvious effect in enhancing the solubility of the components with poor water solubility,like quercetin and resveratrol(p<0.05).Catechin has low P value due to its high polarity.The introduction of four oils can significantly raise the P value of catechin(p<0.05).On the contrary,quercetin and resveratrol have high P value due to their poor water solubility.The addition of PAs and OA can significantly reduce the P values of the two(p<0.05).There was no significant difference among the four oils(p>0.05)in adjusting P value of grape polyphenols.Compared with the use of oils alone,nanoemulsions had more obvious effect on the modulation of the P values of the three grape polyphenols(p<0.05).Apart from the influence on the solubility and P value of the drugs,absorption enhancers may also change the permeability of the small intestinal membrane.The influence of oils and nanoemulsions on the permeability of the small intestine membrane was assessed.The results revealed that the four oils can significantly enhance the permeability of the small intestinal membrane(p<0.05)at proper concentrations.The effect of PAs was significantly stronger than that of OA(p<0.05).There was no obvious difference among the PAs(p>0.05).Compared to oils,nanoemulsions did not show stronger capacity in enhancing permeation.Only OA and PAEE nanoemulsions can significantly improve the permeability of the small intestine membrane(p<0.05),while the other two nanoemulsions have no obvious penetration enhancing effects.Finally,the four oils and their nanoemulsions were combined with the three grape polyphenols to compare the absorption promotion strength using an in vitro mouse small intestine model.The results indicated that the three PAs had obvious effects in enhancing the absorption of the three grape polyphenols(p<0.05).The strength of the three forms of PA was significantly higher than that of OA(p<0.05).There was no obvious difference among the three(p>0.05).Compared to the oils,nanoemulsions did not exhibit stronger effect in increasing transmembrane rate of catechin,but significantly improved the intestinal absorption of quercetin and resveratrol(p<0.05).Moreover,nanoemulsions presented sustained-release characteristics.Multivariate linear fitting was performed using the cumulative permeation rate versus the solubility,P value of polyphenols,and the intestinal membrane permeability,which showed that the effect of oils and nanoemulsions in enhancing the absorption of grape polyphenols was related to the increase of solubility,adjustment of the P value of grape polyphenols,and the improvement of small intestinal permeation.This study discussed the mechanisms of the effect of fatty acids in enhancing the intestinal absorption of grape polyphenols from the viewpoints of the change of drug physiochemical properties and the augment of small intestinal permeability.It provided a new option for improving the oral bioavailability of grape polyphenol and laid a theoretical foundation for the application of PAs fatty acid esters as a new type of absorption enhancer. |
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