Folate Moiety Research Articles

Folate-Functionalized Amphiphilic Chitosan Polymeric Micelles Containing Andrographolide Analogue (3A.1) for Colorectal Cancer

A site-specific drug delivery system of anticancer agents has been delveloped to enhance the therapeutic efficacy and reduce toxicity to the normal tissue. Semi-synthetic andrographolide analogue 3A.1 (19-tert-butyldiphenylsilyl-8,17-epoxy andrographolide) is one of the potential natural anticancer compounds against many types of cancer including colorectal cancer cells. However, the clinical applications of this compound are limited because of low water solubility and lack of suitable delivery carriers. This study aimed to increase the aqueous solubility and improve the anticancer efficacy of 3A.1 via active targeting approaches. In this study, 3A.1 was loaded into the polymeric micelles self-assembled from N-naphthyl-N,O-succinyl chitosan (NSC). The micelles were conjugated with folate moiety (Fol-NSC) for targeting to the cancer cells. All of the 3A.1-loaded micelles were prepared by dropping method, and the physicochemical properties (size, charge, morphology, encapsulating efficiency, loading capacity), in vitro release behavior and in vitro anticancer activities against HT29 colorectal cancer cells were investigated. The 3A.1-loaded micelles were successfully formulated by dropping method using NSC or Fol-NSC. The micelles loaded with 40% initial 3A.1 showed the maximum encapsulating efficiency and loading capacity. The micelles were in the nanometer range, having a negative surface charge and a spherical structure. The colon site-specific release of the 3A.1 from the 3A.1-loaded micelles was obtained. The release of 3A.1 from the Fol-NSC micelles was slower than that from the NSC micelles. Moreover, the Fol-NSC micelles exhibited superior anticancer efficacy than that of the NSC micelles and free 3A.1. In conclusions, the 3A.1-loaded Fol-NSC micelles developed in the present study had suitable physicochemical properties. These nanocarriers may be a potential delivery system for targeted delivery of the 3A.1 to colorectal cancer cells.

The Synthesis of the Folate Acid Quantum Dots and Its Endocytosis in the Hepatoma Cells Mediated by the Folate Receptor

Objectives: To synthesize folate acid conjugated quantum dots (QDs) by a simple and green method and explore endocytosis of the QD-folic acid conjugates (FA-QDs) into hepG2 cells. Methods: CdS QDs were prepared using stearic acid as synthetic medium, and were conjugated with mercaptoacetic acid (MAA) to be water-soluble. Then folic acid moiety was coupled to the surface of the QDs using EDC (1-ethyl-3-(3-dimethylamino propyl)carbodiimide) as a coupling agent. The CdS QDs and FA-QDs were characterized by X-ray spectroscopy (XRD), transmission electron microscopy (TEM), spectro fluorimetry and infrared spectroscopy. Endocytosis of the FA-QDs into hepG2 cells was observed by confocal microscopy and TEM. Results: TEM shows that the CdS QDs and FA-QDs were spherical in shape and had a diameter of 10 nm. Fluorescence spectrum of the QDs also shows their narrow size distribution. Confocal microscopic and TEM results show that FA-QDs were successfully internalized into the hepG2 cells. In addition, confocal results show that when the cells were first be saturated with excess free FA, the endocytosis of the FA-QDs became less, and CdS QDs that were not conjugated with FA were not able to be internalized into the hepG2 cells. Conclusions: The FA-QDs can be synthesized by a simple and green method and be used to successfully target tumor cells: hepG2 cell lines, which was probably mediated by the FA receptor.

Flower-like curcumin-loaded folic acid-conjugated ZnO-MPA- βcyclodextrin nanostructures enhanced anticancer activity and cellular uptake of curcumin in breast cancer cells.

Non-spherical structures are beneficial to advance drug delivery effectiveness compared with common spherical ones, due to increased drug loading capability, improved bonding to a vascular wall, enhanced cellular uptake efficacy and prolonged circulation times. In this study, flower-like Zinc oxide-βcyclodextrin (βCD) nanostructures functionalized by 3-mercaptopropionic acid (MPA) as a non-spherical delivery system was successfully synthesized for aqueous delivery of curcumin (CUR) to enhance its targeting, bioavailability, and release profile. Terminal carboxyl functional groups were used for the conjugation of folic acid (FA) with the aim of active targeting to folate overexpressing breast cancer cells. The in vitro experimental study and mathematical modeling of CUR release revealed a sustained release with Fickian diffusion as the major release mechanism. MTT, colony formation and Annexin-V FITC/PI assays showed the superior anticancer effect of the system compared to free CUR against breast cancer cell line MDA-MB-231 by promoting the apoptotic respond with no cytotoxic effect on HEK293 normal cells. The efficacy of targeting strategy with FA moieties was demonstrated using the augmented cellular uptake of the FA-conjugated system on overexpressed folate receptor alpha (FRα) cells (MDA-MB-468 breast cancer cell line). Furthermore, loading of CUR to the delivery systems significantly lowered the MIC values (2.5 to 5-fold) against S. aureus and E. coli the infections of which are serious problems in cancer patients. According to the results of this study, the system can serve as a promising non-spherical delivery vehicle for enhancing bioavailability and targeting of hydrophobic anticancer agents in the future.

Folic-Acid-Adorned PEGylated Graphene Oxide Interferes with the Cell Migration of Triple Negative Breast Cancer Cell Line, MDAMB-231 by Targeting miR-21/PTEN Axis through NFκB.

Triple negative breast cancer (TNBC), characterized by its aggressive and highly metastatic nature, is difficult to cure by the currently available therapies. In our investigation, folic-acid-adorned PEGylated graphene oxide (FA-PEG-GO) was synthesized by modifying graphene oxide (GO) with folic acid-PEG conjugate (FA-PEG-NH2) by EDC/NHS coupling reaction. FA-PEG-GO exhibited an exceptional potential to attenuate cell migration of TNBC cell line ,MDAMB-231 as compared to GO because of the adorned folic acid moiety, which rendered better targeting. FA-PEG-GO inhibited cell migration by actin depolymerization and perturbing lamellipodia formation. The immunocytochemistry and western blot data unraveled the fact that FA-PEG-GO inhibited cell migration by targeting miR-21 by restricting the nuclear translocation of NFκB. The downregulation of miR-21 resulted in the elevation of PTEN expression which sequentially downregulated pFAK resulting in inhibition of cell migration. Moreover, upregulation of PTEN in FA-PEG-GO treated cells led to the decrease in expression of the downstream regulators including pAkt(Ser473) and pERK1/2, which contributed to the retardation of cell migration. Interestingly, the overexpression of NFκB-p65 by the transfection of NFκB-p65 expression plasmid in TNBC cells reversed the inhibitory effect of FA-PEG-GO on the nuclear translocation of NFκB-p65 which stabilized miR-21 expression and successively downregulated PTEN expression in FA-PEG-GO treated cells. Furthermore, miR-21 overexpression by transfection of miR-21 mimic in turn downregulated PTEN expression and sequentially restored the expression of pFAK even upon FA-PEG-GO treatment. miR-21 overexpression also compensated the inhibitory effect of FA-PEG-GO on pAkt(Ser473) and pERK1/2 which was evident from their significant expression in FA-PEG-GO-treated cells. The studies on chick embryo model ratified the ex ovo antimigratory efficacy of FA-PEG-GO. Altogether, our study unveiled the enormous potential of FA-PEG-GO to attenuate migration of TNBC cell line, MDAMB-231 by targeting the miR-21/PTEN axis through NFκB and thereby providing insights on cancer treatment.

X-ray radiation-induced and targeted photodynamic therapy with folic acid-conjugated biodegradable nanoconstructs.

IntroductionThe depth limitation of conventional photodynamic therapy (PDT) with visible electromagnetic radiation represents a challenge for the treatment of deep-seated tumors.Materials and methodsTo overcome this issue, we developed an X-ray-induced PDT system where poly(lactide-co-glycolide) (PLGA) polymeric nanoparticles (NPs) incorporating a photosensitizer (PS), verteporfin (VP), were triggered by 6 MeV X-ray radiation to generate cytotoxic singlet oxygen. The X-ray radiation used in this study allows this system to breakthrough the PDT depth barrier due to excellent penetration of 6 MeV X-ray radiation through biological tissue. In addition, the conjugation of our NPs with folic acid moieties enables specific targeting of HCT116 cancer cells that overexpress the folate receptors. We carried out physiochemical characterization of PLGA NPs, such as size distribution, zeta potential, morphology and in vitro release of VP. Cellular uptake activity and cell-killing effect of these NPs were also evaluated.Results and discussionOur results indicate that our nanoconstructs triggered by 6 MeV X-ray radiation yield enhanced PDT efficacy compared with the radiation alone. We attributed the X-ray-induced singlet oxygen generation from the PS, VP, to photoexcitation by Cherenkov radiation and/or reactive oxygen species generation facilitated by energetic secondary electrons produced in the tissue.ConclusionThe cytotoxic effect caused by VP offers the possibility of enhancing the radiation therapy commonly prescribed for the treatment of cancer by simultaneous PDT.

Tumor pH-Responsive Release of Drug-Conjugated Micelles from Fiber Fragments for Intratumoral Chemotherapy.

The tumor accumulation of micelles is essential to enhance the cellular uptake and extend the release of chemotherapeutic agents. In the previous study camptothecin (CPT)-conjugated micelles (MCPT) were constructed with disulfide linkages and folate moieties for reduction-sensitive release and cell-selective uptake. This study proposes a strategy to integrate the promicelle polymers (PMCPT) into fiber fragments for intratumoral injection, realizing acid-liable release of PMCPT in response to acidic tumor microenvironment and spontaneous self-assembly into MCPT. Acid-liable 2-propionic-3-methylmaleic anhydride (CDM)-linked poly(ethylene glycol) initiates the ring-opening polymerization of dl-lactide as the fiber matrix. There is no apparent burst release of MCPT from fiber fragments and around 80% of accumulated releases after incubation in pH 6.5 buffers for 40 days. Compared to MCPT freshly prepared via solvent evaporation, the micelles released from fiber fragments reveal similar profiles, such as folate-mediated cellular uptake and glutathione-sensitive drug release. Taking advantage of the aggregation-induced emission (AIE) effect of tetraphenylethylene (TPE) derivatives, TPE-conjugated micelles (MTPE) have been successfully been used to track the self-assembly into micelles after release from fibers and subsequent cell internalization into cytosol. The self-assembly induced fluorescence light-up was also detected after intratumoral injection of fiber fragments. Compared with CPT-loaded fiber fragments and intratumoral or intravenous injection of free MCPT, the sustained release from fiber fragments and high accumulation of micelles in tumors result in significantly higher inhibition of tumor growths, prolongation of animal survival, and induction of tumor cell apoptosis. Thus, the integration of double targeting and double stimuli responsiveness into fragmented fibers provides a feasible strategy to realize the sustained micelle release from fibers and promote the therapeutic efficacy.

Identification of the Uncertainty Structure to Estimate the Acoustic Release of Chemotherapeutics From Polymeric Micelles.

This paper estimates the acoustic drug release from micelles after accurately identifying the underlying statistical noise characteristics in experimental data. The drug release is measured as a change in fluorescence as ultrasound is applied. First, the noise structure affecting the process dynamics and the measurement process is identified in terms of statistical covariance of the aforementioned quantities. Then, the identified covariance magnitudes are utilized to estimate the dynamics of drug release. The performance of different filters is investigated. The identified a priori knowledge is used to implement an optimal Kalman filter, a multi-hypothesis Kalman filter, and a variant of the full information estimator (moving horizon estimator) to the problem at hand. The proposed algorithms are initially deployed in a simulation environment, and then the experimental data sets are fed into the algorithms to validate their performance. Experiments span a number of ultrasonic power densities for both non-targeted and targeted polymeric micelles (the targeting being accomplished using the folate moiety). The results suggest that the proposed algorithm, the optimal Kalman filter, performs better than the other two in all tests performed.

Folic Acid-Targeted Etoposide Cubosomes for Theranostic Application of Cancer Cell Imaging and Therapy.

BackgroundThe aim of this study was to develop a novel Poloxamer-based drug delivery system featuring a tumor-targeting folate moiety, which was expected to provide better targeting properties and therapeutic effects compared with the traditional cubosomes (Cubs).Material/MethodsBoth folate-modified Cubs containing etoposide (ETP-Cubs-FA) and normal cubic nanoparticles loaded with etoposide (ETP-Cubs) were prepared through the fragmentation of bulk gels under the homogenization condition of 1500 bar, and a mean particle size of around 180 nm was obtained with a narrow size distribution. The cubosomes were further characterized by differential scanning calorimetry (DSC) and Polarized light microscopy (PLM). The release of ETP in vitro from these nanoparticles was found to be 82.5% at 36 h, showing a sustained release property compared with the free drug administration.ResultsFolate-modified cubosomes exhibited best anti-proliferative activity followed by normal cubosomes and the free drug. A further cell uptake study of Rhodamine B-loaded Cubs-FA (Rh-B-Cubs-FA) showed a marked increase of cellular accumulation compared with free Rh-B and Rh-B-loaded Cubs (Rh-B-Cubs). In vivo Rh-B-based tumor imaging demonstrated that Cubs-FA specifically targeted the tumor tissue.ConclusionsThe folate-modified cubosomes containing ETP may be a promising drug candidate for antitumor treatment.

Synthesis of Lipophilic Core-Shell Fe3O4@SiO2@Au Nanoparticles and Polymeric Entrapment into Nanomicelles: A Novel Nanosystem for in Vivo Active Targeting and Magnetic Resonance-Photoacoustic Dual Imaging.

In this work, iron/silica/gold core-shell nanoparticles (Fe3O4@SiO2@Au NPs) characterized by magnetic and optical properties have been synthesized to obtain a promising theranostic platform. To improve their biocompatibility, the obtained multilayer nanoparticles have been entrapped in polymeric micelles, decorated with folic acid moieties, and tested in vivo for photoacoustic and magnetic resonance imaging detection of ovarian cancer.

Theranostic Nanoparticles Loaded with Imaging Probes and Rubrocurcumin for Combined Cancer Therapy by Folate Receptor Targeting.

The combination of different therapeutic modalities is a promising option to combat the recurrence of tumors. In this study, polylactic and polyglycolic acid nanoparticles were used for the simultaneous delivery of a boron-curcumin complex (RbCur) and an amphiphilic gadolinium complex into tumor cells with the aim of performing boron and gadolinium neutron capture therapy (NCT) in conjunction with the additional antiproliferative effects of curcumin. Furthermore, the use of Gd complexes allows magnetic resonance imaging (MRI) assessment of the amount of B and Gd internalized by tumor cells. Poly(lactic-co-glycolic acid) (PLGA) nanoparticles were targeted to ovarian cancer (IGROV-1) cells through folate receptors, by including in the formulation a PEGylated phospholipid functionalized with the folate moiety. NCT was performed on IGROV-1 cells internalizing 6.4 and 78.6 μg g-1 of 10 B and 157 Gd, respectively. The synergic action of neutron treatment and curcumin cytotoxicity was shown to result in a significant therapeutic improvement.

Targeting selenium nanoparticles combined with baicalin to treat HBV-infected liver cancer

Herein, we demonstrate baicalin-loaded selenium nanoparticles with a liver-targeting folic acid moiety to treat HBV-infected liver cancer.

Synergistic dual-modified liposome improves targeting and therapeutic efficacy of bone metastasis from breast cancer

Breast cancer frequently metastasizes to bone, where it leads to poor clinical prognosis. Due to the peculiarity of the bone microstructure, the uptake of drugs often happens at non-targeted sites, which produces a similar cytotoxicity in both cancerous and healthy cells. In this study, a rational strategy was implemented to take advantage of a combination of both an octapeptide with eight repeating sequences of aspartate (Asp8) and folate to create a more selective and efficient drug delivery system to target cancer cells in bone tissue. Asp8 and folate were conjugated to the distal ends of DSPE-PEG2000-maleimide and DSPE-PEG2000-amine to create DSPE-PEG2000-Asp8 and DSPE-PEG2000-Folate, respectively, which were incorporated onto the surface of a doxorubicin (DOX)-loaded liposomes (A/F-LS). Asp8, similar to the hydroxyapatite-binding domains of osteopontin and osteocalcin, has been used as bone-targeting moieties for exclusive delivery of drugs to bone. The folate moiety binds selectively to folate receptor-positive tumors. The dual-targeting effects were evaluated by both in vitro and in vivo experiments. By taking advantages of dual-targeting drug delivery, the dual-modified liposomal drug system could relieve pain and improve survival. Inspired by its enhanced therapeutic efficacy and low toxicity, DOX-loaded A/F-LS could serve as an effective drug system for targeted therapy of bone metastases.

Acid-Responsive Polymeric Doxorubicin Prodrug Nanoparticles Encapsulating a Near-Infrared Dye for Combined Photothermal-Chemotherapy

Combination therapy with high spatial and temporal resolution is highly promising for efficient medical treatment of cancer. In this study, doxorubicin (DOX) conjugated amphiphilic block copolymer with a terminal folic acid moiety was prepared, which could self-assemble into nanoparticles by encapsulating organic near-infrared (NIR) absorbing dye IR825 for combined photothermal-chemotherapy. The resulting PDOX/IR825 nanoparticles showed excellent colloidal stability and monodispersity in aqueous solution. Specifically, the conjugated DOX could be released quickly in weak acidic environment for chemotherapy due to the cleavage of acid-labile hydrazone bond. Meanwhile, the encapsulated dye could convert the NIR light energy into heat with high efficiency, which makes the self-assembled nanoparticles an effective platform for photothermal therapy. Confocal microscopy observations and flow cytometry analysis confirmed that the PDOX/IR825 nanoparticles could be efficiently endocytosed by HeLa cells and deliver...

Targeting the Folate Receptor: Effects of Conjugating Folic Acid to DOX Loaded Polymeric Micelles.

In this paper, we report on a potential cancer drug delivery system that utilizes the ligand targeting of the folate receptor. Our drug delivery system consists of Pluronic-P105 micelles, targeted with folic acid moieties. A melanoma folate positive (FR+) (B16-F10), and a fibroblast folate negative (FR-) (NIH-3T3) cell lines are used to compare the cellular accumulation of a chemotherapeutic drug (Doxorubicin) when the delivery is mediated by folated Pluronic P105 micelles. In order to obtain a proper comparison, we corrected for the quenching of Doxorubicin by folic acid molecules and illustrated the significant effect of quenching on the analysis of similar systems. Results show an 80% increase in the accumulation of the antineoplastic agent in the FR+ cell line, when compared to the FR- cell line, thus providing evidence that the efficacy of Pluronic micelles, as drug delivery vehicles, can be enhanced via folic acid targeting.

Unmodified and positively charged gold nanoparticles for sensitive colorimetric detection of folate receptor via terminal protection of small molecule-linked ssDNA

The detection of protein/small molecule interactions plays important roles in drug discovery and protein/metabolite interactions in biology. In this work, by coupling the terminal protection of small molecule-linked ssDNA strategy with the unmodified and positively charged gold nanoparticle ((+)AuNP) nanoprobes, we have developed a sensitive and simple colorimetric sensor for the detection of folate receptor, a highly expressed protein in many kinds of malignant tumors. The target folate receptor binds the folate moieties of the folate-linked ssDNA through high affinity interactions and protects the protein-bound ssDNA from digestion by exonuclease I. The protected ssDNA thus adsorbs the ((+)AuNP) through electrostatic interactions, leading to a red-to-blue color change of the sensing solution for sensitive colorimetric detection of folate receptor at the sub-nanomolar level. Besides, this colorimetric sensor shows high selectivity toward folate receptor against other control proteins. The developed sensor avoids the modification/conjugation of the AuNP nanoprobes and the involvement of any expensive instruments for signal transduction in protein detection. Featured with these obvious advantages, the colorimetric sensor strategy demonstrated herein can be easily expanded for sensitive and convenient detection of various protein/small molecule interactions.

Targeted drug delivery system for ribonuclease A

Ribonuclease A is a very accessible protein that has been tested as a therapeutic for cancer in vitro and in vivo due to its ability to hinder protein production and induce cell death. However, the results in vivo were only positive when high amounts of enzyme were injected directly in the tumors. To overcome this limitation we are testing the use of a drug delivery system consisting of nanoparticles made out of the protein itself coated with poly(lactic‐co‐glycolic acid)‐poly(ethylene glycol)‐folate conjugates (PLGA‐PEG‐FA), in which the folate moiety confers selectivity towards cells overexpressing folic acid receptors on their membrane. Using a solvent displacement nanoprecipitation method we obtained nanoparticles constituted of ribonuclease A. Nanoparticles obtained under different conditions were analyzed by scanning electron microscope and dynamic light scattering, confirming spherical shape and an average size of 600 nm. The FA‐PEG‐PLGA copolymer to coat the nanoparticles was synthesized by conjugating the free alcohol group in PLGA with the carboxyl group in the folate‐conjugated‐PEG. The structure of the product was confirmed by 1H‐NMR. Experiments for coating optimization are in process and the efficacy of the system will be tested in HeLa cells as well as normal cells to confirm the selectivity. This drug delivery system does not have the limitation of a maximum loading capacity, and has been proven by us to efficiently deliver other proteins to cancer cells. Therefore, it is expected to selectively deliver higher amounts of ribonuclease A to cancer cells than previously reported systems.Support or Funding InformationSupported by the MSRC (Molecular Science Research Center)

Silica@zirconia@poly(malic acid) nanoparticles: promising nanocarriers for theranostic applications.

Silica@zirconia@poly(malic acid) nanocarriers of 110 nm mean diameter were designed, synthesized and characterized for the targeted delivery of diagnostic and therapeutic 99mTc to folate-overexpressing tumors. An important achievement was that a multifunctional l-(-)-malic-acid-based copolymer was formed in situ at the surface of the inorganic cores in a single synthetic step incorporating l-(-)-malic acid, β-cyclodextrin rings, folic acid moieties, and polyethylene glycol chains. Morphological and in-depth structural analysis of the particles proved their core@shell structure. Stability experiments in aqueous media evidenced that stable suspensions can be obtained from the lyophilized powder in 10 mM phosphate buffer at pH 7.4. During 14-day degradation experiments, the nanoparticles were found to be slowly dissolving (including inorganic core) in saline and also in total cell medium. An in vitro toxicity assay on hepatocytes showed a concentration-dependent decrease of cell viability down to 63 ± 1% at the highest applied concentration (0.5 mg ml-1). Proof of concept experiments of technetium-99m radiolabelling and in vivo labelling stability are presented.

Multi-Functionalized Carbon Nano-onions as Imaging Probes for Cancer Cells.

Carbon-based nanomaterials have attracted much interest during the last decade for biomedical applications. Multimodal imaging probes based on carbon nano-onions (CNOs) have emerged as a platform for bioimaging because of their cell-penetration properties and minimal systemic toxicity. Here, we describe the covalent functionalization of CNOs with fluorescein and folic acid moieties for both imaging and targeting cancer cells. The modified CNOs display high brightness and photostability in aqueous solutions and their selective and rapid uptake in two different cancer cell lines without significant cytotoxicity was demonstrated. The localization of the functionalized CNOs in late-endosomes cell compartments was revealed by a correlative approach with confocal and transmission electron microscopy. Understanding the biological response of functionalized CNOs with the capability to target cancer cells and localize the nanoparticles in the cellular environment, will pave the way for the development of a new generation of imaging probes for future biomedical studies.

An investigation of folic acid-protein association sites and the effect of this association on folic acid self-assembly.

The contribution of folic acid (FA)-tryptophan interactions to FA-protein association was investigated in the context of using FA as a drug carrier in protein delivery systems. Bovine serum albumin (BSA) and indolicidin were used as model proteins in the study. The FA-BSA complex was characterized by using the Bradford reagent to identify the impact of FA-BSA association on BSA-dye reagent interactions. UV-visible spectroscopic analysis of the FA-BSA mixture showed that the absorbance maximum of BSA-dye reagent occurred at 595nm, even after the association of FA with BSA. This confirms that protonated amino acid groups of the protein are not involved in FA-BSA association. Moreover, molecular dynamics (MD) simulation confirmed the presence of an associative interaction between aromatic moieties in FA and tryptophan moieties in the indolicidin molecule, which disrupted FA self-assembly. An X-ray diffraction (XRD) study showed that there was limited disruption of FA self-assembly after the addition of BSA or tryptophan. This suggests that FA and BSA are compatible and associate with each other. Graphical Abstract Mechanism of folic acid and protein association.

Dual nutraceutical nanohybrids of folic acid and calcium containing layered double hydroxides

Dual nutraceutical nanohybrids consisting of organic nutrient, folic acid (FA), and mineral nutrient, calcium, were prepared based on layered double hydroxide (LDH) structure. Among various hybridization methods such as coprecipitation, ion exchange, solid phase reaction and exfoliation-reassembly, it was found that exfoliation-reassembly was the most effective in terms of intercalation of FA moiety between Ca-containing LDH layers. X-ray diffraction patterns and infrared spectra indicated that FA molecules were well stabilized in the interlayer space of LDHs through electrostatic interaction. From the atomic force and scanning electron microscopic studies, particle thickness of LDH was determined to be varied with tens, a few and again tens of nanometers in pristine, exfoliated and reassembled state, respectively, while preserving particle diameter. The result confirmed layer-by-layer hybrid structure of FA and LDHs was obtained by exfoliation-reassembly. Solid UV–vis spectra showed 2-dimensional molecular arrangement of FA moiety in hybrid, exhibiting slight red shift in n→π* and π→π* transition. The chemical formulae of FA intercalated Ca-containing LDH were determined to Ca1.30Al(OH)4.6FA0.74·3.33H2O and Ca1.53Fe(OH)5.06FA2.24·9.94H2O by inductively coupled plasma-atomic emission spectroscopy, high performance liquid chromatography and thermogravimetry, showing high nutraceutical content of FA and Ca.