Apocarotenoid Bixin Research Articles

First Vesicular Self-assembly of an Apocarotenoid Bixin in Aqueous Liquids and its Antibacterial Activity.

Bixin1 is the major constituent of the reddish carotenoids present in the seed-coat of Bixa orellana. The use of the extract of the seed-coat of Bixa orellana in food, cosmetics and garments is well known. The nano-sized long 24C chain molecule has nine conjugated double bonds having extended conjugation with the '-COOH' and '-COOMe' groups present at the two ends of the molecule. Herein, we report the first self-assembly of bixin in several aqueous liquids. The molecule undergoes spontaneous self-assembly in several liquids yielding vesicular self-assembly. Characterizations of the self-assemblies of bixinwere carried out by various microscopic techniques, X-ray diffraction and FTIR studies. The critical vesicular concentrations (CVCs) of the compound carried out in DMSO-water in three different solvent ratios as 2: 1 (v/v), 1: 1 (v/v) and 1: 4 (v/v) were determined to be 100 mM, 90 mM and 60 mM respectively indicating lower CVC values at higher proportion of water. Utilization of the vesicular self-assemblies of bixin have been demonstrated in the entrapment and release of fluorophores including the anticancer drugs doxorubicin and curcumin. Self-assembled bixin and curcumin loaded self-assembled bixin showed significant antibacterial activity with both Gram positive as well as Gram negative bacteria.

Isolation and biological activities of naturally occurring apocarotenoid bixin

The naturally occurring apo-carotenoid bixin is obtained from annatto (Bixa orellana). It is unique among the natural carotenoids through its water solubility and easily extractability from the seed coat of Bixa orellana. Bixin is (2E,4E,6E,8E,10E,12E,14E,16Z, 18E)-20- methoxy-4,8,13,17-tetramethyl-20-oxicosa 2,4,6,8,10,12,14,16,18-nonaenoic acid, an apo-carotenoid which is a terpenoid compound derived from the oxidative cleavage of carotenoids. Several biological activities of bixin has been reported earlier such as selective antimyeloma effects of cis-bixin, antinociceptive and anti-Inflammatory effects, antioxidant action against cis-platine induced chromosome in rats, hypoglycemic effect in alloxan-induced diabetic rats, allodynia, anxious and depressive-like behaviors, targeting NRF2 for improved skin barrier function and photoprotection, antiatherogenic effect in hypercholesterolemic rabbits, triggering apoptosis of cancer cells, immunomodulatory role in dogs, sensitization of human melanoma cells to dacarbazine-induced apoptosis, sensitization of human lung cancer and cervical cancer cell to cisplatin in vitro, anti-SARS-CoV-2 activity, antibacterial activity of cis-bixin conjugated silver nanoparticles, protective effect on carbon tetrachloride-induced hepatotoxicity in rats, bixin/crocetin nanoparticles on quality and shelf life improvement of cheddar cheese etc. In this review, the biological properties of bixin has been discussed. A practical method for the isolation of bixin 1 from the seed coat of Bixa orellana commonly known as “Sinduri” (in Bengali) and its characterization has also been reported.

Bixin as a new class of biosorbent for Hg2+ removal from aqueous solutions

In this study, the apocarotenoid bixin was investigated as a potential biosorbent for Hg2+ in aqueous solutions. Bixin was used to form a thin film on an inert support using the drop-coating technique. This film was characterised using ATR-FTIR, SEM, and TGA-DTG techniques. Parameters that influence adsorption, such as pH, temperature, NaCl concentration, salt type, bixin mass, and supports for the bixin film, were evaluated. Using 0.112 mg of a bixin covering on a cellulose filter, the sorbent material was able to remove 75 % of Hg2+ from a 2.0 mL, 0.3 mg g−1, pH 6.00, 25 °C solution after 180 min of contact. Kinetics studies were carried out between 5 and 1440 min, and the pseudo-first-order adsorption model was applied. Equilibrium isotherm data were analysed, and the best fit was for the non-linear Freundlich model, with an R2 of 0.9868, RMSE of 0.37, and maximum sorption capacity of 85.38 mg g−1 obtained by the Langmuir model. The bixin film was applied in spiked environmental samples (lake and mining effluent), and it achieved 86 % Hg2+ removal. The results show that the bixin film is a suitable alternative for efficiently removing mercury from various aqueous systems. This introduces a new class of biosorbent based on apocarotenoids, which are sustainable, low cost, nontoxic, and easily obtainable.

Activation of NRF2 by topical apocarotenoid treatment mitigates radiation-induced dermatitis

Radiation therapy is a frontline treatment option for cancer patients; however, the effects of radiotherapy on non-tumor tissue (e.g. radiation-induced dermatitis) often worsen patient quality of life. Previous studies have implicated the importance of redox balance in preventing dermatitis, specifically in reference to modulation of the nuclear factor (erythroid-derived 2)-like 2 (NRF2) signaling pathway. Due to the cytoprotective functions of transcriptional target genes of NRF2, we investigated how modulation of NRF2 expression could affect DNA damage, oxidative stress, and cell viability in response to radiotherapy. Specifically, it was noted that NRF2 knockdown sensitized human skin keratinocytes to ionizing radiation; likewise, genetic ablation of NRF2 in vivo increased radiosensitivity of murine epidermis. Oppositely, pharmacological induction of NRF2 via the apocarotenoid bixin lowered markers of DNA damage and oxidative stress, while preserving viability in irradiated keratinocytes. Mechanistic studies indicated that topical pretreatment using bixin as an NRF2 activator antagonized initial DNA damage by raising cellular glutathione levels. Additionally, topical application of bixin prevented radiation-induced dermatitis, epidermal thickening, and oxidative stress in the skin of SKH1 mice. Overall, these data indicate that NRF2 is critical for mitigating the harmful skin toxicities associated with ionizing radiation, and that topical upregulation of NRF2 via bixin could prevent radiation-induced dermatitis.

5-Azacytidine downregulates the SABATH methyltransferase genes and augments bixin content in Bixa orellana L. leaves

Bixa orellana L. seeds are an important source of the liposoluble apocarotenoid bixin and the natural hydrosoluble dye norbixin. These natural dyes are used for food, pharmaceutical, and cosmetic applications. The participation of methyltransferases has already been demonstrated in the bixin biosynthesis pathway, and the alteration of these genes expression can lead to changes in the production of bixin. Treatment with demethylating agents, such as 5-azacytidine (5-AzaC), was previously related to an increased production of secondary compounds and serves for the study of secondary metabolism pathways in plants. Here, we evaluated for the first time the effect of 5-AzaC on the expression of SABATH methyltransferase genes as well as some bixin pathway candidate genes, global DNA methylation, and bixin, chlorophylls, and total carotenoid content in the leaves of B. orellana plants cultured in vitro. Exposing shoot apexes to 5-AzaC for 48 h affected global genome methylation but did not alter chlorophylls and total carotenoid content in the leaves of B. orellana. Importantly, plants submitted to this demethylating agent exhibited downregulation of the SABATH and CCD4 families of genes, upregulation of the ALDH3H1 gene, and increased bixin content in the leaves, suggesting that this methyltransferase inhibitor can selectively regulate the biosynthesis and production of bixin in B. orellana by an unknown mechanism. Moreover, here we demonstrate the possibility to use 5-AzaC to increase bixin contents in B. orellana plants. The methyltransferase inhibitor, 5-Azacytidine, can selectively regulate the biosynthesis and production of bixin in annatto plant leaves, by upregulating BoALDH3H1 gene and downregulating BoSABATH and BoCCD4 gene families.

Carotenoids as Novel Therapeutic Molecules Against Neurodegenerative Disorders: Chemistry and Molecular Docking Analysis.

Alzheimer’s disease (AD) is the most devastating neurodegenerative disorder that affects the aging population worldwide. Endogenous and exogenous factors are involved in triggering this complex and multifactorial disease, whose hallmark is Amyloid-β (Aβ), formed by cleavage of amyloid precursor protein by β- and γ-secretase. While there is no definitive cure for AD to date, many neuroprotective natural products, such as polyphenol and carotenoid compounds, have shown promising preventive activity, as well as helping in slowing down disease progression. In this article, we focus on the chemistry as well as structure of carotenoid compounds and their neuroprotective activity against Aβ aggregation using molecular docking analysis. In addition to examining the most prevalent anti-amyloidogenic carotenoid lutein, we studied cryptocapsin, astaxanthin, fucoxanthin, and the apocarotenoid bixin. Our computational structure-based drug design analysis and molecular docking simulation revealed important interactions between carotenoids and Aβ via hydrogen bonding and van der Waals interactions, and shows that carotenoids are powerful anti-amyloidogenic molecules with a potential role in preventing AD, especially since most of them can cross the blood-brain barrier and are considered nutraceutical compounds. Our studies thus illuminate mechanistic insights on how carotenoids inhibit Aβ aggregation. The potential role of carotenoids as novel therapeutic molecules in treating AD and other neurodegenerative disorders are discussed.

Expression and Interaction Analysis among Saffron ALDHs and Crocetin Dialdehyde.

In saffron, the cleavage of zeaxanthin by means of CCD2 generates crocetin dialdehyde, which is then converted by an unknown aldehyde dehydrogenase to crocetin. A proteome from saffron stigma was released recently and, based on the expression pattern and correlation analyses, five aldehyde dehydrogenases (ALDHs) were suggested as possible candidates to generate crocetin from crocetin dialdehydes. We selected four of the suggested ALDHs and analyzed their expression in different tissues, determined their activity over crocetin dialdehyde, and performed structure modeling and docking calculation to find their specificity. All the ALDHs were able to convert crocetin dialdehyde to crocetin, but two of them were stigma tissue-specific. Structure modeling and docking analyses revealed that, in all cases, there was a high coverage of residues in the models. All of them showed a very close conformation, indicated by the low root-mean-square deviation (RMSD) values of backbone atoms, which indicate a high similarity among them. However, low affinity between the enzymes and the crocetin dialdehyde were observed. Phylogenetic analysis and binding affinities calculations, including some ALDHs from Gardenia jasmonoides, Crocus sieberi, and Buddleja species that accumulate crocetin and Bixa orellana synthetizing the apocarotenoid bixin selected on their expression pattern matching with the accumulation of either crocins or bixin, pointed out that family 2 C4 members might be involved in the conversion of crocetin dialdehyde to crocetin with high specificity.

Topical Bixin Confers NRF2-Dependent Protection Against Photodamage and Hair Graying in Mouse Skin.

Environmental exposure to solar ultraviolet (UV) radiation causes acute photodamage, premature aging, and skin cancer, attributable to UV-induced genotoxic, oxidative, and inflammatory stress. The transcription factor NRF2 [nuclear factor erythroid 2 (E2)-related factor 2] is the master regulator of the cellular antioxidant response protecting skin against various environmental stressors including UV radiation and electrophilic pollutants. NRF2 in epidermal keratinocytes can be activated using natural chemopreventive compounds such as the apocarotenoid bixin, an FDA-approved food additive and cosmetic ingredient from the seeds of the achiote tree (Bixa orellana). Here, we tested the feasibility of topical use of bixin for NRF2-dependent skin photoprotection in two genetically modified mouse models [SKH1 and C57BL/6J (Nrf2+/+ versus Nrf2-/-)]. First, we observed that a bixin formulation optimized for topical NRF2 activation suppresses acute UV-induced photodamage in Nrf2+/+ but not Nrf2-/- SKH1 mice, a photoprotective effect indicated by reduced epidermal hyperproliferation and oxidative DNA damage. Secondly, it was demonstrated that topical bixin suppresses PUVA (psoralen + UVA)-induced hair graying in Nrf2+/+ but not Nrf2-/- C57BL/6J mice. Collectively, this research provides the first in vivo evidence that topical application of bixin can protect against UV-induced photodamage and PUVA-induced loss of hair pigmentation through NRF2 activation. Topical NRF2 activation using bixin may represent a novel strategy for human skin photoprotection, potentially complementing conventional sunscreen-based approaches.

Targeting NRF2 for Improved Skin Barrier Function and Photoprotection: Focus on the Achiote-Derived Apocarotenoid Bixin.

The transcription factor NRF2 (nuclear factor-E2-related factor 2) orchestrates major cellular defense mechanisms including phase-II detoxification, inflammatory signaling, DNA repair, and antioxidant response. Recent studies strongly suggest a protective role of NRF2-mediated gene expression in the suppression of cutaneous photodamage induced by solar UV (ultraviolet) radiation. The apocarotenoid bixin, a Food and Drug Administration (FDA)-approved natural food colorant (referred to as ‘annatto’) originates from the seeds of the achiote tree native to tropical America, consumed by humans since ancient times. Use of achiote preparations for skin protection against environmental insult and for enhanced wound healing has long been documented. We have recently reported that (i) bixin is a potent canonical activator of the NRF2-dependent cytoprotective response in human skin keratinocytes; that (ii) systemic administration of bixin activates NRF2 with protective effects against solar UV-induced skin damage; and that (iii) bixin-induced suppression of photodamage is observable in Nrf2+/+ but not in Nrf2−/− SKH-1 mice confirming the NRF2-dependence of bixin-induced antioxidant and anti-inflammatory effects. In addition, bixin displays molecular activities as sacrificial antioxidant, excited state quencher, PPAR (peroxisome proliferator-activated receptor) α/γ agonist, and TLR (Toll-like receptor) 4/NFκB (nuclear factor kappa-light-chain-enhancer of activated B cells) antagonist, all of which might be relevant to the enhancement of skin barrier function and environmental stress protection. Potential skin photoprotection and photochemoprevention benefits provided by topical application or dietary consumption of this ethno-pharmacologically validated phytochemical originating from the Americas deserves further preclinical and clinical examination.

The Apocarotenoid Bixin Protects Mouse Skin against UV-induced Damage in an NRF2-dependent Manner

Skin barrier function and integrity can be impaired as a result of exposure to environmental stressors, including solar ultra violet (UV) radiation. UV induces oxidative stress, inflammation, premature ageing, and cancer. Therefore, there is a need to find effective and safe strategies to protect the skin from these deleterious effects. An effective strategy to prevent UV damage involves activation of the transcription factor nuclear factor-E2-related factor 2 (NRF2) to induce cellular stress responses and damage repair. We recently characterized the apocarotenoid bixin, and FDA-approved natural food colorant from the seeds of the achiote tree (Bixa orellana), as a safe and potent NRF2 activator. In this study, we tested the feasibility of enhancing NRF2-dependent skin protection using a topical bixin formulation in a model for acute UV-induced skin damage and UV-induced hair graying. Nrf2+/+ and Nrf2-/- SKH-I hairless mice (acute skin damage) or C57BL/6J mice (hair graying) were treated twice with a topical bixin formulation 48 h and 24 h before exposure to UV. Skin tissues were harvested at 24 h (acute skin damage) or 2 weeks (hair graying) post-exposure and analyzed by immunoblotting and immunohistochemistry. We observed that bixin activated cutaneous expression of NRF2 and its target genes, and limited photodamage in Nrf2+/+ but not Nrf2-/- mice. Moreover, bixin reduced inflammation, oxidative damage, and apoptosis in the epidermis of Nrf2+/+ SKH-I mice. Consistently, bixin prevented UV-induced hair graying in Nrf2+/+ C57BL/6J mice. These results suggest that bixin-based interventions may be useful for NRF2-dependent skin protection against environmental insults.

The achiote-derived apocarotenoid bixin protects skin against environmental insults through NRF2-dependent activation of the epidermal antioxidant response

The achiote-derived apocarotenoid bixin protects skin against environmental insults through NRF2-dependent activation of the epidermal antioxidant response

Systemic administration of the apocarotenoid bixin protects skin against solar UV-induced damage through activation of NRF2

Exposure to solar ultraviolet (UV) radiation is a causative factor in skin photodamage and carcinogenesis, and an urgent need exists for improved molecular photoprotective strategies different from (or synergistic with) photon absorption. Recent studies suggest a photoprotective role of cutaneous gene expression orchestrated by the transcription factor NRF2 (nuclear factor-E2-related factor 2). Here we have explored the molecular mechanism underlying carotenoid-based systemic skin photoprotection in SKH-1 mice and provide genetic evidence that photoprotection achieved by the FDA-approved apocarotenoid and food additive bixin depends on NRF2 activation. Bixin activates NRF2 through the critical Cys-151 sensor residue in KEAP1, orchestrating a broad cytoprotective response in cultured human keratinocytes as revealed by antioxidant gene expression array analysis. Following dose optimization studies for cutaneous NRF2 activation by systemic administration of bixin, feasibility of bixin-based suppression of acute cutaneous photodamage from solar UV exposure was investigated in Nrf2+/+ versus Nrf2−/− SKH-1 mice. Systemic administration of bixin suppressed skin photodamage, attenuating epidermal oxidative DNA damage and inflammatory responses in Nrf2+/+ but not in Nrf2−/− mice, confirming the NRF2-dependence of bixin-based cytoprotection. Taken together, these data demonstrate feasibility of achieving NRF2-dependent cutaneous photoprotection by systemic administration of the apocarotenoid bixin, a natural food additive consumed worldwide.

Control of Carotenoid Gene Expression in Bixa orellana L. Leaves Treated with Norflurazon

Bixa orellana (annatto or lipstick tree) is a perennial tropical plant that stores and produces considerable quantities of the apocarotenoid bixin, a culturally and economically important pigment used worldwide. However, the mechanisms underlying the gene regulation and pigment accumulation of bixin and carotenoids in annatto remain unknown. Bixin is present in the different tissues of the plant, although this pigment is primarily accumulated in the seed coat. Thus, the leaves are useful organs for understanding carotenoid and bixin production, thereby facilitating the study of this pigment, which would otherwise be difficult in ligneous adult plants. To study the regulation of bixin synthesis and to determine which genes are important regulatory molecules at the transcription level, the herbicide norflurazon (NF) was used to block carotenoid synthesis and bixin concentrations in B. orellana leaves. The genes activated in the early stages of the carotenoid pathway are involved in lycopene production (dxs, psy and pds), and those induced in the later stages of the carotenoid pathway, such as β and e-lycopene cyclases and boccd1, were differentially expressed compared with the control. The expression of some genes was more susceptible to certain concentrations of NF, potentially reflecting the roles of these genes in carotenoid synthesis in B. orellana. These results suggest that apocarotenoids, such as bixin, are synthesized using alternative precursors through the actions of genes that have not yet been identified.

Analysis of Binding Interaction between the Natural Apocarotenoid Bixin and Human Serum Albumin by Circular Dichroism and Fluorescence Spectroscopy

Bixin is an important, pharmacologically active dietary cis-carotenoid, but its interaction with potential macromolecular targets is completely unexplored. This work was aimed to study the binding of bixin to human serum albumin (HSA), the most abundant protein in blood plasma. Circular dichroism (CD) spectroscopy in combination with UV/VIS absorption spectroscopy and fluorescence quenching techniques were applied. Appearance of induced CD bands in the UV- and VIS-absorption spectral regions indicated the formation of non-covalent carotenoid-albumin complexes. Shape and spectral position of the extrinsic Cotton effects suggested the binding of a single bixin molecule to HSA in chiral conformation. Scatchard and non-linear regression analyses of CD titration data resulted in similar values for the association constant (Ka = 6.6 and 4.6x10(5) M(-1), resp.) and for the number of binding sites (n = 1). The binding interaction was independently confirmed by fluorescence-quenching experiment from which the binding parameters were also calculated. CD Displacement measurements performed with marker ligands established that the main drug binding sites of HSA are not involved in binding of bixin. Palmitic acid decreased the amplitude of the induced CD bands suggesting a common albumin binding site for bixin and long-chain fatty acids. The above data indicate that HSA plays a significant role in the plasma transportation of bixin and related dietary carboxylic acid carotenoids.