Crocin-1 laden thermosensitive chitosan-based hydrogel with smart anti-inflammatory performance for severe full-thickness burn wound therapeutics.

Burns are the fourth most common type of civilian trauma worldwide, and the management of severe irregular scald wounds remains a significant challenge. Herein, crocin-1 laden hydroxybutyl chitosan (CRO-HBC) thermosensitive hydrogel with smart anti-inflammatory performance was developed for accelerating full-thickness burn healing. The injectable and shape adaptability of the CRO-HBC gel make it a promising candidate for effectively filling scald wounds with irregular shapes, while simultaneously providing protection against external pathogens. The CRO-HBC gel network formed by hydrophobic interactions exhibited an initial burst release of crocin-1, followed by a gradual and sustained release over time. The excessive release of ROS and pro-inflammatory cytokines should be effectively regulated in the early stage of wound healing. The controlled release of crocin-1 from the CRO-HBC gel adequately addresses this requirement for wound healing. The CRO-HBC hydrogel also exhibited an excellent biocompatibility, an appropriate biodegradability, keratinocyte migration facilitation properties, and a reactive oxygen species scavenging capability. The composite CRO-HBC hydrogel intelligently mitigated inflammatory responses, promoted angiogenesis, and exhibited a commendable efficacy for tissue regeneration in a full-thickness scalding model. Overall, this innovative temperature-sensitive CRO-HBC injectable hydrogel dressing with smart anti-inflammatory performance has enormous potential for managing severe scald wounds.

Phytochemical Characterization of Two New Olive Oil Genotypes Growing in Southern Tunisia.

This research can be considered as the first complete survey for the valorization of new olive genotypes cultivated in the South-East of Tunisia as well as their oils. The study aimed to characterize the phytochemical composition of virgin olive oil produced from two olive cultivars, namely Nourgou and Gousalani. The pomological characterization of fruits, the quality criteria and the phytochemical profile were quantified. Additionally, antioxidant activity was evaluated using Ferric reducing antioxidant power (FRAP) and Oxygen radical absorbance capacity (ORAC) tests to also obtain a bioactive characterization of these monovarietal olive oils. The obtained results revealed that the analyzed olive oils samples can be classified into Extra Virgin category (EVOO) according to the regulated physicochemical characteristics. Our findings showed a significant variability in the chemical parameters of the analyzed EVOO likely associated with the genetic potential, mainly for chlorophylls contents (1.37-1.64 mg/kg), in carotenoids pigments (3.97-10.86 mg/kg), in α-tocopherol (175.59-186.87 mg/kg), in sterols (1036.4-1931.4 mg/kg) in oleic acid (65.33-68.73%), in palmitic acid (C16:0) (13.32-17.48%), in linoleic acid (C18:2) (11.06-13.47%). Additionally, the HPLC-MS/MS analysis showed that the two EVOOs analyzed contained appreciable amounts of total polyphenols, ranging from 348.03 up to 516.16 mg/kg, in Nourgou and Gousalani oils, respectively. Regarding the individual phenolic compounds, the EVOO samples were mainly characterized by phenolic alcohols, phenolic acids, secoiridoids, verbascoside, flavonoids and phenolic aldehydes. The prevalent simple phenolics detected were secoiridoids with the dominance of the oleuropein aglycone in Gousalani oil. In addition, findings from in vitro antioxidant assays (FRAP and ORAC) revealed that the two studied oils possessed a powerful antiradical activity and a good reducing power capacity. In conclusion, these new EVOOs exhibited a superior quality compared to other Tunisian varieties, considering their antiradical activity and reducing power capacity.

Content of Primary and Secondary Carotenoids in the Cells of Cryotolerant Microalgae Chloromonas reticulata.

Snow (cryotolerant) algae often form red (pink) spots in mountain ecosystems on snowfields around the world, but little is known about their physiology and chemical composition. Content and composition of pigments in the cells of the cryotolerant green microalgae Chloromonas reticulata have been studied. Analysis of carotenoids content in the green (vegetative) cells grown under laboratory conditions and in the red resting cells collected from the snow surface in the Subpolar Urals was carried out. Carotenoids such as neoxanthin, violaxanthin, anteraxanthin, zeaxanthin, lutein, and ß-carotene were detected. Among the carotenoids, the ketocarotenoid astaxanthin with high biological activity was also found. It was established that cultivation of the algae at low positive temperature (6°C) and moderate illumination (250 µmol quanta/(m2⋅s) contributed to accumulation of all identified carotenoids, including extraplastidic astaxanthin. In addition to the pigments, fatty acids accumulated in the algae cells. The data obtained allow us to consider the studied microalgae as a potentially promising species for production of carotenoids.

Microbial bacterioruberin: The new C50 carotenoid player in food industries.

The demand for natural products has significantly increased, driving interest in carotenoids as bioactive compounds for both human and animal consumption. Carotenoids, natural pigments with several biological properties, like antioxidant and antimicrobial, are increasingly preferred over synthetic colorants by the consumers (chemophobia). The global carotenoid market is projected to reach US$ 2.45 billion by 2034, driven by consumer preferences for natural ingredients and regulatory restrictions on synthetic products. Among carotenoids, bacterioruberin (BR), a C50 carotenoid naturally found in microbial hyperhalophilic archaea and in moderate halophilic archaea, stands out for its exceptional antioxidant capabilities, surpassing even well-known carotenoids like astaxanthin. BR's and its derivatives unique structure, with 13 conjugated double bonds and four -OH groups, contributes to its potent antioxidant activity and potential applications in food, feed, supplements, pharmaceuticals, and cosmeceuticals. This review explores BR's chemical and biological properties, upstream and downstream technologies, analytical techniques, market applications, and prospects in the colorants industry. While BR is not intended to replace existing carotenoids, its inclusion enriches the range of natural products available to meet the rising demand for natural alternatives. Furthermore, BR's promising antioxidant capacity positions it as a key player in the future carotenoid market, offering diverse industries a natural and potent alternative for several applications.

Ionic Liquids toward Enhanced Carotenoid Extraction from Bacterial Biomass.

Carotenoids are high added-value products primarily known for their intense coloration and high antioxidant activity. They can be extracted from a variety of natural sources, such as plants, animals, microalgae, yeasts, and bacteria. Gordonia alkanivorans strain 1B is a bacterium recognized as a hyper-pigment producer. However, due to its adaptations to its natural habitat, hydrocarbon-contaminated soils, strain 1B is resistant to different organic solvents, making carotenoid extraction through conventional methods more laborious and inefficient. Ionic liquids (ILs) have been abundantly shown to increase carotenoid extraction in plants, microalgae, and yeast; however, there is limited information regarding bacterial carotenoid extraction, especially for the Gordonia genus. Therefore, the main goal of this study was to evaluate the potential of ILs to mediate bacterial carotenoid extraction and develop a method to achieve higher yields with fewer pre-processing steps. In this context, an initial screening was performed with biomass of strain 1B and nineteen different ILs in various conditions, revealing that tributyl(ethyl)phosphonium diethyl phosphate (IL#18), combined with ethyl acetate (EAc) as a co-solvent, presented the highest level of carotenoid extraction. Afterward, to better understand the process and optimize the extraction results, two experimental designs were performed, varying the amounts of IL#18 and EAc used. These allowed the establishment of 50 µL of IL#18 with 1125 µL of EAc, for 400 µL of biomass (cell suspension with about 36 g/L), as the ideal conditions to achieve maximal carotenoid extraction. Compared to the conventional extraction method using DMSO, this novel procedure eliminates the need for biomass drying, reduces extraction temperatures from 50 °C to 22 ± 2 °C, and increases carotenoid extraction by 264%, allowing a near-complete recovery of carotenoids contained in the biomass. These results highlight the great potential of ILs for bacterial carotenoid extraction, increasing the process efficiency, while potentially reducing energy consumption, related costs, and emissions.

Isolation of Bioactive Compounds (Carotenoids, Tocopherols, and Tocotrienols) from Calendula Officinalis L., and Their Interaction with Proteins and Oils in Nanoemulsion Formulation.

Calendula officinalis L. has numerous health-promoting properties due to the presence of a large number of lipophilic compounds. Their effective delivery to the body requires the use of an appropriate technique such as emulsification. So, the main purpose of this study was to understand how the profile of lipophilic compounds from pot marigold and the pro-health potential are shaped by different types of protein, oil, and drying techniques in o/w nanoemulsion. To obtain this, the profiles of carotenoid compounds and tocols were measured. Additionally, antioxidant potential and the ability to inhibit α-amylase and α-glucosidase were measured. Pea protein emulsion exhibited a higher final content of carotenoid compounds (23.72-39.74 mg/100 g), whereas those with whey protein had stronger α-amylase inhibition (487.70 mg/mL). The predominant compounds in the studied nanoemulsions were ß-carotene (between 19% and 40%), followed by α-tocopherol/γ-tocopherol. The type of proteins shaped the health-promoting properties and determined the content of health-promoting compounds.

Valorization of tomato processing by-products: Predictive modeling and optimization for ultrasound-assisted lycopene extraction.

Lycopene is a carotenoid highly valuable to the food, pharmaceutical, dye, and cosmetic industries, present in ripe tomatoes and other fruits with a distinctive red color. The main source of lycopene is tomato crops. This bioactive component can be successfully isolated from tomato processing waste, commonly called tomato pomace, mostly made from tomato skins, seeds, and some residual tomato tissue. The main investigative focus in this work was the application of green engineering principles in each stage of the optimized ultrasound-assisted extraction (UAE) of enzymatically treated tomato skins to obtain functional extracts rich in lycopene. The experimental plan was designed to determine the influence of studied operating parameters: enzymatic reaction time (60, 120, and 180 min), extraction time (0, 5, 10, 15, 30, 60, and 120 min), and temperature (25, 35 and 45 ℃) on lycopene yield. Process optimization was performed based on the yield of lycopene [1018, 1067, and 1120 mg/kg] achieved at optimal operating conditions. An artificial neural network (ANN) model was developed and trained for predictive modeling of the closed extraction system, with operating parameters used as input neurons and experimentally obtained values for lycopene content defined as the output neural layer. Applied ANN architecture provided a high correlation of experimental output with ANN-generated data (R=0.99914) with a model deviation error for the entire data set of RMSE=5.3 mg/kg. The k-Nearest Neighbor algorithm was introduced to predict lycopene yield using experimental key features: operating temperature, extraction time, and time of enzymatic treatment, split into training and testing sets with an 85/15 ratio. The model interpretation was conducted through the SHAP (SHapley Additive exPlanations) methodology.

Chitosan-coated manganese ferrite nanoparticles enhanced Rhodotorula toruloides carotenoid production.

The present study aims to analyze the interaction between Rhodotorula toruloides and magnetic nanoparticles and evaluate their effect on carotenoid production. The manganese ferrite nanoparticles were synthesized without chitosan (MnFe2O4) and chitosan coating (MnFe2O4-CS) by the co-precipitation method assisted by hydrothermal treatment. XRD (X-ray diffraction), Magnetometry, Dynamic Light Scattering (DLS) and FTIR (Fourier-Transform Infrared Spectroscopy), are used to characterize the magnetic nanoparticles. The crystallite size of MnFe2O4 was 16 nm for MnFe2O4 and 20 nm for MnFe2O4-CS. The magnetic saturation of MnFe2O4-CS was lower (39.6 ± 0.6 emu/g) than the same MnFe2O4 nanoparticles (42.7 ± 0.3 emu/g), which was attributed to the chitosan fraction presence. The MnFe2O4-CS FTIR spectra revealed the presence of the characteristic chitosan bands. DLS demonstrated that the average hydrodynamic diameters were 344 nm for MnFe2O4 and 167 nm for MnFe2O4-CS. A kinetic study of cell immobilization performed with their precipitation with a magnet demonstrated that interaction between magnetic nanoparticles and R. toruloides was characterized by an equilibrium time of 2 h. The adsorption isotherm models (Langmuir and Freundlich) were fitted to the experimental values. The trypan blue assay was used for cell viability assessment. The carotenoid production increased to 256.2 ± 6.1 µg/g dry mass at 2.0 mg/mL MnFe2O4-CS. The use of MnFe2O4-CS to stimulate carotenoid yeast production and the magnetic separation of biomass are promising nanobiotechnological alternatives. Magnetic cell immobilization is a perspective technique for obtaining cell metabolites.

A Comparative Analysis on Impact of Extraction Methods on Carotenoids Composition, Antioxidants, Antidiabetes, and Antiobesity Properties in Seagrass Enhalus acoroides: In Silico and In Vitro Study.

Enhalus acoroides, a tropical seagrass, is known for its significant contribution to marine ecosystems and its potential health benefits due to bioactive compounds. This study aims to compare the carotenoid levels in E. acoroides using green extraction via ultrasound-assisted extraction (UAE) and microwave-assisted extraction (MAE) and to evaluate the biological properties of these extracts against oxidative stress, diabetes, and obesity through in silico and in vitro analyses. E. acoroides samples were collected from Manado City, Indonesia, and subjected to UAE and MAE. The extracts were analyzed using UHPLC-ESI-MS/MS to identify carotenoids, including ß-carotene, lutein, lycopene, ß-cryptoxanthin, and zeaxanthin. In silico analysis was conducted to predict the compounds' bioactivity, toxicity, and drug-likeness using WAY2DRUG PASS and molecular docking with CB-Dock2. The compounds C3, C4, and C7 demonstrated notable interactions, with key metabolic proteins and microRNAs, further validating their potential therapeutic benefits. In vitro assays evaluated antioxidant activities using DPPH and FRAP assays, antidiabetic properties through α-glucosidase and α-amylase inhibition, and antiobesity effects via lipase inhibition and MTT assay with 3T3-L1 cells. Results indicated that both UAE and MAE extracts exhibited significant antioxidant, antidiabetic, and antiobesity activities. MAE extracts showed higher carotenoid content and greater biological activity compared to UAE extracts. These findings suggest that E. acoroides, mainly when extracted using MAE, has promising potential as a source of natural bioactive compounds for developing marine-based antioxidant, antidiabetic, and antiobesity agents. This study supplements existing literature by providing insights into the efficient extraction methods and the therapeutic potential of E. acoroides carotenoids.

Towards more sustainable cooking practices to increase the bioaccessibility of colourless and provitamin A carotenoids in cooked carrots.

The effect of different cooking methods (boiling, baking, steaming and microwaving) on the colour and texture of carrots, as well as on the bioaccessibility of carotenoids, was investigated in order to identify the more "sustainable cooking" methods. Cooking resulted in statistically significant increases in total carotenoid bioaccessibility, both with intensity and duration of treatments. In particular, significant increases in carotenoid bioaccessible content (CBC) were observed, ranging from 6.03-fold (microwave) to 8.90-fold (baking) for the most intense cooking conditions tested. Although the relative concentration of the colourless carotenoids (phytoene and phytofluene) in raw carrots is lower than that of provitamins A α- and ß-carotene, the bioaccessible content of the colourless ones is much higher. From an energy consumption standpoint and considering samples with the same tenderness, the highest CBC values per kWh decreased in the order microwaving > baking > water cooking > steaming. Our findings are important to help combat vitamin A deficiency since increases of up to ∼40-fold and ∼70-fold in the CBCs of the vitamin A precursors α- and ß-carotene, respectively, were observed. These results provide a basis for defining "sustainable cooking" as "cooking practices that optimize intensity, duration and other parameters leading to a more efficient use of energy to maximize the bioavailability of nutrients and other beneficial food components (such as bioactives) while ensuring food appeal and safety".

Preparation of two amphiphilic dendritic small molecule gelators based on poly (aryl ether) modified silica-based chromatographic stationary phases and molecular shape recognition for shape-restricted isomers.

Geometric isomers tend to have similar polarities and differ only in molecular shape. Vigorously developing new stationary phases to meet the requirements for the separation of isomers that have similar physicochemical properties is still an urgent topic in separation science. Poly (arylene ether)-based dendrimers are known for their multifunctional branched peripheral structures and high self-assembly properties. In this paper, two amphiphilic dendritic organic small molecule gelling agents based on poly (aryl ether), PAE-ANT and PAE-PA, were prepared and conjugated to the silica surface. SiO2@PAE-ANT and SiO2@PAE-PA were used as HPLC stationary phases for the separation of non-polar shape-restricted isomers. Both stationary phases have very high molecular shape selectivity for isomers such as polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), tocopherols and carotenoids. Separation of cis-trans geometric isomers such as diethylstilbestrol and polar compounds such as monosubstituted benzenes and anilines can also be achieved. These two columns offer more flexible selectivity and higher separation performance than commercial C18 and phenyl columns. There is a difference in molecular shape selectivity between the two stationary phases for the same analyte test probes. SiO2@PAE-ANT showed slightly better linear selectivity for non-polar shape-restricted isomers compared to SiO2@PAE-PA with Janus-type PAE-PA bonding phase. This separation behavior may be attributed to the ordered spatial structure formed by the gel factor on the surface of the stationary phase and the combined effect of multiple weak interaction centers (hydrophobic, hydrophilic, hydrogen bonding and π-π interactions). It was also possible to separate nucleoside and nucleobase strongly polar compounds well in the HILIC mode, suggesting that hydrophilic groups in PAE-ANT and PAE-PA are involved in the interactions, reflecting their amphiphilic nature. The results show that the ordered gelation of dendritic organic small molecule gelators on the SiO2 surface, along with multiple carbonyl-π, π-π and other interactions, play a crucial role in the separating shape-restricted isomers. The integrated and ordered functional groups serve as the primary driving force behind the exceptionally high molecular shape selectivity of SiO2@PAE-ANT and SiO2@PAE-PA phases. Alterations in the structure of dendritic organic small molecule gelators can impact both molecular orientation and recognition ability, while changes in the type of functional groups influences the separation mechanism of shape-restricted isomers.

Detecting bioactive compound contents in Dancong tea using VNIR-SWIR hyperspectral imaging and KRR model with a refined feature wavelength method.

Hyperspectral imaging (HSI) provides opportunity for non-destructively detecting bioactive compounds contents of tea leaves and high detection accuracy require extracting effective features from the complex hyperspectral data. In this paper, we proposed a feature wavelength refinement method called interval band selecting-competitive adaptive reweighted sampling-fusing (IBS-CARS-Fusing) to extract feature wavelengths from visible-near-infrared (VNIR) and short-wave-near-infrared (SWIR) hyperspectral images. Combined with the proposed IBS-CARS-Fusing method, a kernel ridge regression (KRR) model was established to predict the contents of bioactive compounds including chlorophyll a, chlorophyll b, carotenoids, tea polyphenols, and amino acids in Dancong tea. It was revealed that the IBS-CARS-Fusing method can improve Rp2 of KRR model for these bioactive compounds by 4.77%, 4.60%, 6.74%, 15.52%, and 13.10%, respectively, and Rp2 of the model reached high values of 0.9500, 0.9481, 0.8946, 0.8882, and 0.8622. Additionally, a leaf compound mass per area thermal map was used to visualize the spatial distribution of the compounds.

Apo-12'-capsorubinal exhibits anti-inflammatory effects and activates nuclear factor erythroid 2-related factor 2 in RAW264.7 macrophages.

Apocarotenoids have short carbon chain structures cleaved at a polyene-conjugated double bond. They can be biosynthesized in plants and microorganisms. Animals ingest carotenoids through food and then metabolize them into apocarotenoids. Although several apocarotenoids have been identified in the body, their precise health functions are still poorly understood. This study investigated the anti-inflammatory activities of apo-12'-capsorubinal in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. It was confirmed that apo-12'-capsorubinal was not cytotoxic to the macrophages at the concentrations tested. Apo-12'-capsorubinal treatment led to a marked downregulation of interleukin (IL)-6 protein and Il6 mRNA levels. This apocarotenoid exhibited more potent inhibitory effects than its parent carotenoids, capsanthin and capsorubin. Furthermore, apo-12'-capsorubinal, but not its parent carotenoids, promoted the nuclear accumulation of nuclear factor erythroid 2-related factor 2 (Nrf2) and upregulated the expression of Nrf2-target genes, such as heme oxygenase 1 (HO-1) and NAD(P)H:quinone oxidoreductase 1 (NQO-1), in a dose-dependent manner. Furthermore, a comparison using apo-12'-zeaxanthinal and 7,8-dihydro-8-oxo-apo-12'-zeaxanthinal revealed that the α, ß-unsaturated carbonyl group on the polyene linear chain mediated the enhanced nuclear Nrf2 translocation, HO-1 expression, and inhibition of IL-6 production. In contrast, apo-12'-mytiloxanthinal, which harbored a hydroxyl group at C-8 of apo-12'-capsorubinal, did not exhibit any of these activities. These results indicated that the ß carbon of the α, ß-unsaturated carbonyl group in the linear part of the polyene chain is crucial to the Nrf2-activating and anti-inflammatory effects of apo-12'-capsorubinal. This study will advance our knowledge of the physiological significance of xanthophyll-derived apocarotenoids and their potential use as nutraceuticals and pharmaceuticals.

Multiple Mechanisms of Haematococcus pluvialis-Derived Carotenoids to Inhibit Glycidyl Ester Formation in Rice Oil and a Chemical Model at High Temperatures.

The common presence of glycidyl esters (GEs) in refined vegetable oils has been a concern for food safety. The present study aimed to investigate the inhibitory effects of three carotenoids derived from Haematococcus pluvialis microalga on GE formation in both rice oil and a chemical model during heating. The addition of astaxanthin (AS), lutein (LU), and ß-carotene (CA) at 0.6 mg/g in rice oil can reduce GE formation by 65.0%, 57.1%, and 57.5%, respectively, which are significantly higher than those achieved by common antioxidants such as l-ascorbyl palmitate (39.0%), α-tocopherol (18.5%), tert-butyl hydroquinone (42.7%), and quercetin (26.2%). UPLC-Q-TOF-MS/MS analysis showed that two new compounds, that is, propylene glycol monoester and diester of palmitic acid, were formed in the CA-added chemical model, which provided direct experimental evidence for the inhibition of antioxidants including AS, LU, and CA against GE formation not only by indirect antioxidative action but also by direct radical reactions to competitively prevent the formation of cyclic acyloxonium intermediates. Furthermore, it was interestingly found that only AS could react with the GEs. The adduct of AS with GEs, astaxanthin-3-O-propanetriol esters, was preliminarily identified using Q-TOF-MS/MS in the heated AS-GE model, suggesting that reacting with GEs might represent another distinct mechanism of AS to eliminate GEs.

Evaluation of the Shelf Life of Myristica-fragrans Powder-Flavored Oils Obtained through the Application of Two Processes: Infusion and Co-Pressing Technology.

This work aimed to evaluate the impact of enrichment processing on the quality parameters, bioactivity and sensorial aspects of Myristica fragrans (mace)-flavored olive oil storage for one year. The mace powder was added to extra virgin olive oil through two different processes: immediately after crushing the olives by mixing mace (1% weight/weight (w/w)) with the olive paste (MAVOO-M) and by adding mace to extra virgin olive oil (C) (2% w/w) (MAVOO-I). A multi-analytical approach was applied to measure the main qualitative indexes, such as the free acidity, peroxide value and ultraviolet parameters. The total phenolic and carotenoid contents (TPC and TCC, respectively) and α-tocopherol were also evaluated, as well as the sensory attributes. The radical scavenging potential was estimated by using two different in vitro tests, namely, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH). A significant increase in the free acidity parameter was found in all the flavored oils, and particularly in the MAVOO-M (1.27% oleic acid); at the same time, this oil was the sample with the lowest peroxide value (i.e., 9.68 meqO2/kg) after 360 days of storage. At the end of the storage, an increase in L* values was found in both the MAVOO-M and -I vs. the C (43.88 and 43.02, respectively, vs. 42.62). The TCC was strongly influenced by the addition of mace, especially when the infusion process was used. In fact, after one year of storage, the TCC in the MAVOO-I resulted in ~34.7% more than the MAVOO-M. A promising DPPH radical scavenging activity was observed independently by the applied aromatization process, with IC50 values of 19.77 and 17.80 µg/mL for the MAVOO-M and MAVOO-I, respectively. However, this activity decreased during storage, and a similar trend was observed using the ABTS test. In conclusion the infusion as enrichment methodology led to more promising results in terms of functionality compared with the co-mixing one.

Dynamic and Energetic Aspects of Carotenoids In-and-Around Model Lipid Membranes Revealed in Molecular Modelling.

In contrast to plants, humans are unable to synthesise carotenoids and have to obtain them from diet. Carotenoids fulfil several crucial biological functions in the organism; however, due to poor solubility in water, their bioavailability from plant-based food is low. The processes of carotenoid absorption and availability in the human body have been intensively studied. The recent experimental findings concerning these processes are briefly presented in the introductory part of this review, together with a summary of such topics as carotenoid carriers, body transport and tissue delivery, to finally report on molecular-level studies of carotenoid binding by membrane receptors. The main message of the review is contained in the section describing computational investigations of carotenoid intercalation and dynamic behaviour in lipid bilayers. The relevance of these computational studies lies in showing the direct link between the microscopic behaviour of molecules and the characteristics of their macroscopic ensembles. Furthermore, studying the interactions between carotenoids and lipid bilayers, and certainly proteins, on the molecular- and atomic-level using computational methods facilitates the interpretation and explanation of their macroscopic properties and, hopefully, helps to better understand the biological functions of carotenoids.

Comparative profiling of Chlorella vulgaris cells, extracts, and intact chloroplasts using electron transfer matrix-assisted laser desorption/ionization mass spectrometry (ET-MALDI-MS).

The intricate composition of microalgal pigments plays a crucial role in various biological processes, from photosynthesis to biomarker identification. Traditional pigment analysis methods involve complex extraction techniques, posing challenges in maintaining analyte integrity. In this study, we employ Electron Transfer Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry (ET-MALDI-MS) to compare the pigmentary profiles of Chlorella vulgaris intact cells, chloroplasts, and solvent extracts. We aim to obtain comprehensive extracts rich in polar and non-polar compounds using ultrasound-assisted and supercritical fluid extraction methods. Additionally, intact chloroplasts are isolated using a lysis buffer and sucrose density gradient centrifugation. Our ET-MALDI-MS analysis reveals distinct compositional differences, highlighting the impact of extraction protocols on microalgal pigment identification. We observe prominent signals corresponding to radical cations of key pigments, including chlorophylls and carotenoids, which are crucial for C. vulgaris identification. Furthermore, ET-MALDI-MS facilitates the identification of specific lipids within chloroplast membranes and other organelles. This study underscores the rapid and precise nature of ET-MALDI-MS in microalgal biomarker analysis, providing valuable insights into phytoplankton dynamics, trophic levels, and environmental processes. C. vulgaris emerges as a promising model for studying pigment composition and membrane lipid diversity, enhancing our understanding of microalgal ecosystems.

In vitro dioxygenase activity characterization using headspace stir bar sorptive extraction (HSSE).

An analytical approach employing headspace sorptive extraction coupled with gas chromatography-mass spectrometry (HSSE-GC-MS) has been successfully developed for the determination of apocarotenoid volatiles arising from the enzymatic activity of carotenoid cleavage enzymes (CCDs) in Escherichia coli. The GjCCD4a enzyme derived from gardenia, known for its cleavage specificity at 7,8 and 7',8' double bonds across diverse carotenoid substrates, was utilized as a reference enzyme, using ß-carotene as the substrate for the enzymatic activity assays. Optimal headspace conditions for analysis were established following a 5 hours induction period of the recombinant GjCCD4a protein within E. coli cells, engineered to produce ß-carotene. The analytical method demonstrated linearity, with correlation coefficient (R2 > 0.95) in calibration, while achieving detection and quantification limits conducive to the accurate determination of ß-cyclocitral. Notably, this methodological framework significantly reduced both the handling complexity and sample processing time in comparison to conventional liquid chromatography methods employed for the detection of cleavage products and determination of CCD activities. The proposed HSSE-GC-MS approach not only enhances the efficiency of apocarotenoid analysis but also provides a sensitive means for unraveling the intricate enzymatic processes associated with CCD-mediated carotenoid cleavage in a bacterial model system.

Unveiling the critical role of K+ for xanthorhodopsin expression in E. coli.

Xanthorhodopsin (XR), a retinal-binding 7-transmembrane protein isolated from the eubacterium Salinibacter ruber, utilizes two chromophores (retinal and salinixanthin (SAL)) as an outward proton pump and energy-donating carotenoid. However, research on XR has been impeded owing to limitations in achieving heterogeneous expression of stable forms and high production levels of both wild-type and mutants. We successfully expressed wild-type and mutant XRs in Escherichia coli in the presence of K+. Achieving XR expression requires significant K+ and a low inducer concentration. In particular, we highlight the significance of Ser-159 in helix E located near Gly-156 (a carotenoid-binding position) as a critical site for XR expression. Our findings indicate that replacing Ser-159 with a smaller amino acid, alanine, can enhance XR expression in a manner comparable to K+, implying that Ser-159 poses a steric hindrance for pigment formation in XR. In the presence of K+, the proton pumping and photocycle of the wild-type and mutants were characterized and compared; the wild-type result suggests similar properties to the first reported XR isolation from the S. ruber membrane fraction. We propose that the K+ gradient across the cell membrane of S. ruber serves to uphold the membrane potential of the organism and plays a role in the expression of proteins, such as XR, as demonstrated in our study. Our findings deepen the understanding of adaptive protein expression, particularly in halophilic organisms. We highlight salt selection as a promising strategy for improving protein yield and functionality.

Borneol and lactoferrin dual-modified crocetin-loaded nanoliposomes enhance neuroprotection in HT22 cells and brain targeting in mice.

Crocetin (CCT), a natural bioactive compound extracted and purified from the traditional Chinese medicinal herb saffron, has been shown to play a role in neurodegenerative diseases, particularly depression. However, due to challenges with solubility, targeting, and bioavailability, formulation development and clinical use of CCT are severely limited. In this study, we used the emulsification-reverse volatilization method to prepare CCT-loaded nanoliposomes (CN). We further developed a borneol (Bor) and lactoferrin (Lf) dual-modified CCT-loaded nanoliposome (BLCN) for brain-targeted delivery of CCT. The results of transmission electron microscope (TEM) and particle size analysis indicated that the size of BLCN (∼140 nm) was suitable for transcellular transport across olfactory axons (∼200 nm), potentially paving a direct path to the brain. Studies on lipid solubility, micropolarity, and hydrophobicity showed that BLCN had a relatively high Lf grafting rate (81.11 ± 1.33 %) and CCT entrapment efficiency (83.60 ± 1.04 %) compared to other liposomes, likely due to Bor improving the lipid solubility of Lf, and the combination promoting the orderly arrangement of liposome membrane molecules. Microplate reader and fluorescence microscopy analysis showed that BLCN efficiently promoted the endocytosis of fluorescent coumarin 6 into HT22 cells with a maximal fluorescence intensity of (13.48 ± 0.80 %), which was significantly higher than that of CCT (5.73 ± 1.17 %) and CN (12.13 ± 1.01 %). BLCN also exhibited sustained function, remaining effective for more than 12 h after reaching a peak at 1 h in cells, while CN showed a significant decrease after 4 h. The uptake mechanisms of BLCN in HT22 cells mainly involve energy-dependent, caveolae-mediated, and microtubule-mediated endocytosis, as well as micropinocytosis. Furthermore, BLCN displayed a significant neuroprotective effect on HT22 cells in glutamate-, corticosterone-, and H2O2-induced models. Tissue fluorescence image analysis of mice showed that BLCN exhibited substantial retention of fluorescent DiR in the brain after nasal administration for 12 h. These findings suggest that CCT has the potential for cellular uptake, neuroprotection, and targeted delivery to the brain following intranasal administration when encapsulated in Bor and Lf dual-modified nanoliposomes.