Development and Validation of a HPLC Method Using a Monolithic Column for Quantification of trans-Resveratrol in Lipid Nanoparticles for Intestinal Permeability Studies

Abstract

The development of nanodelivery systems that protect trans-resveratrol is extremely important to preserve its bioactive properties in the development of further applications as nutraceuticals to supplement foods and beverages. In this work, a validated HPLC method was developed for the quantification of trans-resveratrol in lipid nanoparticles for application in studies of in vitro intestinal permeability. The chromatographic separation was achieved in a C18 monolithic column connected to a fluorometric detector (330/374 nm), by isocratic elution consisting of 2% acetic acid/acetonitrile (80:20). Two calibration ranges were established (0.020-0.200 and 0.200-2.00 μmol L(-1)), and low quantification limits (2-6 nmol L(-1), 23-69 pg) were achieved. Stability studies showed that trans-resveratrol is stable for 24 h at 4 °C, and storage at room temperature and freeze-thaw cycles are not recommended. The proposed method was applied to in vitro intestinal permeability studies, in which values between 0.05 ± 0.01 and 1.8 ± 0.3 μmol L(-1) were found.

Nanoscale Delivery of Resveratrol towards Enhancement of Supplements and Nutraceuticals

“…HBSS was used as the control medium, while the last two media mimic the fasted- and fed-state intestinal juices, respectively, because they contain natural surfactants (bile salts and lecithin) that generate micelles that are present during the digestive process [ 44 , 45 ]. The HPLC quantification method was firstly developed and validated for resveratrol in the three different matrices [ 51 ]. The permeation of free resveratrol and resveratrol-loaded lipid nanoparticles across the intestinal barrier is presented as cumulative transport over 4 h in Figure 4 .…”

“…This fact can be related to the micellarization of resveratrol and lipid nanoparticles [ 45 ]. On the other hand, FaSSIF and FeSSIF protection against trans -resveratrol degradation is also reported [ 51 ], which can explain the increased resveratrol absorption when taken with meals. We have performed identical studies to ensure the integrity and stability of the nanoparticles in the different media used, in order to ensure the validity of the intestinal permeability assay.…”

“…Aliquots from the basolateral side were collected at regular intervals of 30 min and treated with 20% acetonitrile/acetic acid (92:8, v/v) to promote the disruption of lipid nanoparticles and the release of resveratrol to be quantified. The quantification was performed by HPLC separation in a C18 monolithic column with isocratic elution in a 2% acetic acid solution/acetonitrile (80:20), followed by fluorimetric detection (330/374 nm), according to a validated method [ 51 ]. TEER values were rechecked at the end to confirm the quality of the monolayer after the experiment.…”

Quantification of Trans-Resveratrol-Loaded Solid Lipid Nanoparticles by a Validated Reverse-Phase HPLC Photodiode Array

“…The literature reports that the LOD and LOQ depend on various factors such as the method (e.g., spectrophotometric, fluorometric, mass spectrometry) and type/quantity of sample. Neves et al developed a methodology using chromatographic separation connected to a fluorometric detector [56]. The objective was to detect the concentration of nanomolars between 1 and 229 nmol/L of tRES loaded in lipid nanoparticles to evaluate in vitro intestinal assays, using Caco-2 cell monolayers.…”