Cell Wall Research Articles

Novel triazole-nucleoside hybrids as potential antimicrobial agents: Design, synthesis, and molecular docking

Despite decoding many important genes in the formation of microbes, particularly pathogenic and mutant ones that lead to resistance to a wide range of antibiotics, this opens the door to discovering new antimicrobial compounds, whether synthetic or natural in origin. A new set of nucleoside-triazole hybrids was created to provide a more effective topoisomerase II for microbial therapy. All freshly synthesized compounds' structures were confirmed using spectral data and elemental analysis. The newly synthesized compounds were docked to the area of topoisomerase II's active site; compounds 3, 7, 15, 18, and 19 exhibited distinct hydrogen bonding formation and pi-pi interactions. The results of testing every compound against six different microbial strains were expressed as the width of the inhibitory zone.Except for Aspergillus Niger, most of the synthetic compounds inhibit all bacteria. Compounds 15 and 19 had the lowest MIC values (1.95, 3.9 µg/mL against B. subtills and 1.95, 3.9 µg/mL against S. aureus). Nucleoside 15 had MBC values between 3.9 and B. subtills. The in-vitro topoisomerase II enzyme assay revealed that compounds 3, 15, 18, and 19 had IC50 values of 0.17, 0.05, 0.13, and 0.08 µmol/mL, respectively. A transmission electron microscope (TEM) was utilized to assess morphological changes on the bacterial surface following treatment with nucleoside 15. It was discovered that nucleoside 15 induced substantial cell wall weakening, which exacerbated cell membrane rupture.

Morphological, physiological, and transcriptomic analyses indicate that cell wall properties and antioxidant processes are potential targets for improving the aluminium tolerance of broad beans

Aluminium (Al) stress is the second-leading abiotic stress on crops. An improved understanding of the response mechanisms of plants to Al stress will provide scientific guidance for enhancing the crops’ tolerance to Al stress. In this study, Al stress (50–200 μM AlCl3) caused visible damage to broad bean (Vicia faba L.) roots rather than shoots, which was attributed to Al accumulation and distribution in different tissues. Root transcriptomic analysis revealed that Al stress altered cell wall properties by downregulating lignin synthesis and several xyloglucan endotransglucosylase/hydrolase-, expansin- and peroxidase (POD)-encoding genes, which likely weakened cell extensibility to inhibit root growth. Additionally, Al stress impeded reactive oxygen species scavenging pathways involving POD activity and flavonoid biosynthesis, leading to oxidative damage characterised by malondialdehyde accumulation. These results indicate that optimising cell wall properties and/or enhancing antioxidant processes are crucial for alleviating Al toxicity to broad beans. Interestingly, exogenous application (500 and 1000 μM) of the flavonoid apigenin effectively alleviated Al toxicity in broad bean roots by partially improving the total antioxidant capacity of the roots. This study contributes to understanding the interaction between plants and Al and provides new strategies to alleviate Al toxicity in crops.

Penicillin-binding proteins exhibit catalytic redundancy during asymmetric cell division in Clostridioides difficile.

Peptidoglycan synthesis is an essential driver of bacterial growth and division. The final steps of this crucial process involve the activity of SEDS family glycosyltransferases that polymerize glycan strands and class B penicillin-binding protein (bPBP) transpeptidases that cross-link them. While most bacteria encode multiple bPBPs that perform specialized roles during specific cellular processes, some bPBPs can play redundant roles that are important for resistance against certain cell wall stresses. Our understanding of these compensatory mechanisms, however, remains incomplete. Endospore-forming bacteria typically encode multiple bPBPs that drive morphological changes required for sporulation. The sporulation-specific bPBP, SpoVD, is important for synthesizing the asymmetric division septum and spore cortex peptidoglycan during sporulation in the pathogen Clostridioides difficile. Although SpoVD catalytic activity is essential for cortex synthesis, we show that it is unexpectedly dispensable for SpoVD to mediate asymmetric division. The dispensability of SpoVD's catalytic activity requires the presence of its SEDS partner, SpoVE, and is facilitated by the catalytic activity of another sporulation-specific bPBP, PBP3. Our data further suggest that PBP3 interacts with components of the asymmetric division machinery, including SpoVD. These findings suggest a possible mechanism by which bPBPs can be functionally redundant in diverse bacteria and facilitate antibiotic resistance.

Carbon and Nitrogen Sources Influence Parasitic Responsiveness in Trichoderma atroviride NI-1.

Parasitic species of Trichoderma use hydrolytic enzymes to destroy the host cell wall. Preferent carbon and nitrogen sources suppress the expression of genes related to parasitism. Here, different nutrients were evaluated in the parasitic isolated NI-1, which was identified as Trichoderma atroviride. The genes cbh1 and chb2 (cellobiohydrolases), bgl3.1 (endoglucanase), and pra1 and prb1 (proteinases) were poorly expressed during the interaction between NI-1 and Phytophthora capsici on PDA. However, gene expression improved on minimal medium with preferent and alternative carbon sources. Dextrin and glucose stimulated higher transcript levels than cellulose, sucrose, and glycerol. Also, ammonium stimulated a stronger parasitic responsiveness than the alternative nitrogen sources. During interaction against different phytopathogens, NI-1 detects their host differentially from a distance due to the cbh1 and cbh2 genes being only induced by P. capsici. The pra1 and ech42 genes were induced before contact with Botrytis cinerea and Rhizoctonia solani, while when confronted with P. capsici they were stimulated until contact and overgrowth. The prb1 and bgl3.1 genes were induced before contact against the three-host assayed. Overall, T. atroviride prefers to parasitize and has the capacity to distinguish between an oomycete and a fungus, but nutrient quality regulates its parasitic responsiveness.

A Klebsiella rhizobacterium deregulates the metabolism of phytopathogenic Aspergillus flavus during in-vitro assays and confers protective functions

In previous investigations, we have identified a rhizobacterium (Klebsiella sp. MBE02) that confers host protection against several phytopathogenic fungi. For instance, this rhizobacterium prevents Aspergillus flavus infection and promotes peanut growth and fitness in controlled and field-conditions. The mechanistic basis of the protective function offered by this rhizobacterium is not completely understood. MBE02 directly restricts the growth of the pathogenic fungi, which led us to hypothesize that it may strongly dysregulate the metabolism of A. flavus, and inhibit critical metabolic processes of the fungus, which severely restricts pathogen growth. We have tested this hypothesis by using untargeted metabolite profiling. Sixty-nine A. flavus metabolites accumulated differentially due to the presence of the MBE02. MBE02 could inhibit several important metabolic pathways, which include the biosynthesis of critical primary metabolites such as amino acids and fatty acids. It also impacts energy metabolism of the fungus, and that the accumulation of several structural components, including of the cell wall, were strongly inhibited. MBE02 abrogated the accumulation of disease-causing metabolites in A. flavus, whereas the accumulation of metabolites that inhibit fungal growth were enhanced. On the other hand, A. flavus did not strikingly impact the accumulation of metabolites of the MBE02. Our investigation supports the hypothesis that Klebsiella sp. MBE02 mediates protective function by directly impairing the pathogen’s metabolism.

Characterization of water states and pore size distribution in Beijing poplar using nuclear magnetic resonance techniques

ABSTRACT This study employs nuclear magnetic resonance (NMR) techniques to elucidate the water states and pore size distribution within Beijing poplar (Populus beijingensis W. Y. Hsu) across various temperatures. By analyzing the variation in moisture content (MC) at different freezing temperatures, the proportion and distribution of cell wall pores were determined using the NMR Cryoporometry (NMR-C) method. The results show approximately 60% of the pores in both the heartwood (HW) and sapwood (SW) of Beijing poplar have diameters were 1.5 nm, while the proportion of pores with diameters greater than 4.6 nm was less than 10%. Moreover, the MC of both more freely bound water (C-water) and OH bound water (B-water) is observed to decrease exponentially with decreasing temperature. B-water in Beijing poplar is found to maintain a higher MC than C-water across the temperature range from −3 to −60°C, indicating greater resistance to freezing. The findings of this study contribute to a comprehensive understanding of water states in Beijing poplar. Also the pore size distribution of Beijing poplar provides reference data for the selection of wood chemical modification groups.

Storage temperature affects metabolism of sweet corn

Temperature is one of the critical factors affecting the postharvest storage quality of sweet corn, but effects of storage temperatures have not been elucidated at the molecular level. In this study, the effects of storage at 0 °C, 4 °C and 20 °C, on quality and associated effects on the transcriptome and metabolome were investigated. Compared with storage at 20 °C, cobs stored at 0 °C and 4 °C had up-regulated expression of genes associated with cell wall disassembly (PG, PE, BMY, β-Gal) and down-regulated expression of genes involved in lignin synthesis, the up-regulation of SPP, SPS and SUS involved in sucrose synthesis, and the down-regulation of IN, FK, HXK and PFK involved in sucrose catabolism. Most of the genes involved in ethylene biosynthesis and signaling were inhibited at 0 °C, except for ERFs, while abscisic acid synthesis and signal transduction were most active at this temperature. Near-freezing temperature also maintains the integrity of the cell membrane by inhibiting the expression of PLD, PLC and LOX, which encode membrane lipid-degrading enzymes. Metabolomics and combined analysis showed that the contents of lysophospholipid, glutathione and L-ascorbic acid in sweet corn increased at low temperatures. These results provide insight into the effects of low temperatures on sweet corn quality.

High cycle fatigue behavior and strengthening mechanisms of laser powder bed fusion pure tantalum

The fatigue behavior and strengthening mechanisms of the laser powder bed fusion pure tantalum (LPBF-Ta), coupled with a fatigue life prediction model, were originally explored. The fatigue strength of LPBF-Ta fabricated at the optimal laser energy density (154.16 J/mm3) is 348 MPa (106 cycles), showing a remarkable improvement of about 80 % and 50 % compared with that of the annealed and cold worked commercially pure tantalum, respectively. The microstructural analysis identifies that cellular structures averaging 300 nm in size contribute 438.2 MPa to strengthening, serving as the main contributor to fatigue strength. The breaching of cell walls by dislocations, along with the activation of cross-slip, ensures plasticity and ultimately enhances fatigue strength. Besides, the analysis of the defects reveals that the internal lack of fusion pore can induce the secondary fatigue crack, resulting in a region of brittle crack propagation. Furthermore, a finite life prediction model is established by originally incorporating the size of the crack source defect, and it significantly improves the prediction accuracy of the fatigue life. These results provide guidance for the development of fatigue strengthening strategies for LPBF-Ta.

Disruption to de novo uridine biosynthesis alters β-1,3-glucan masking in Candida albicans.

The uridine derivatives UDP-glucose and UDP-N-acetylglucosamine are important for cell wall construction as they are the precursors for the synthesis of β-1,3-glucan and chitin, respectively. Previous studies have demonstrated attenuated virulence of uridine auxotrophs in mice, which has been attributed to insufficient uridine levels for growth in the host. We have discovered that uridine deprivation in the uridine auxotroph ura3ΔΔ disrupts cell wall architecture by increasing surface mannans, exposing β-1,3-glucan and chitin, and decreasing UDP-sugar levels. Cell wall architecture and UDP-sugars can be rescued with uridine supplementation. The cell wall architectural disruptions in the ura3ΔΔ mutant also impact immune activation since the mutant elicited greater TNFα secretion from RAW264.7 macrophages than wild type. To determine if cell wall defects contributed to decreased virulence in the ura3ΔΔ mutant, we used a murine model of systemic infection. Mice infected with the ura3ΔΔ mutant exhibited increased survival and reduced kidney fungal burden compared with mice infected with wild type. However, suppression of the immune response with cyclophosphamide did not rescue virulence in mice infected with the ura3ΔΔ mutant, indicating the attenuation in virulence of uridine auxotrophs can be attributed to decreased growth in the host but not increased exposure of β-1,3-glucan. Moreover, the ura3ΔΔ mutant is unable to grow on ex vivo kidney agar, which demonstrates its inability to colonize the kidneys due to poor growth. Thus, although uridine auxotrophy elicits changes to cell wall architecture that increase the exposure of immunogenic polymers, metabolic fitness costs more strongly drive the observed virulence attenuation.IMPORTANCECandida albicans is a common cause of bloodstream infections (candidemia). Treatment of these bloodstream infections is made difficult because of increasing antifungal resistance and drug toxicity. Thus, new tactics are needed for antifungal drug development, with immunotherapy being of particular interest. The cell wall of C. albicans is composed of highly immunogenic polymers, particularly β-1,3-glucan. However, β-1,3-glucan is naturally masked by an outer layer of mannoproteins, which hampers the detection of the fungus by the host immune system. Alteration in cell wall components has been shown to increase β-1,3-glucan exposure; however, it is unknown how the inability to synthesize precursors to cell wall components affects unmasking. Here, we demonstrate how cell wall architecture is altered in response to a deficit in precursors for cell wall synthesis and how uridine is a crucial component of these precursors.

H3K4me3 changes occur in cell wall genes during the development of Fagopyrum tataricum morphogenic and non-morphogenic calli.

Epigenetic changes accompany the dynamic changes in the cell wall composition during the development of callus cells. H3K4me3 is responsible for active gene expression and reaction to environmental cues. Chromatin immunoprecipitation (ChIP) is a powerful technique for studying the interplay between epigenetic modifications and the DNA regions of interest. In combination with sequencing, it can provide the genome-wide enrichment of the specific epigenetic mark, providing vital information on its involvement in the plethora of cellular processes. Here, we describe the genome-wide distribution of H3K4me3 in morphogenic and non-morphogenic callus of Fagopyrum tataricum. Levels of H3K4me3 were higher around the transcription start site, in agreement with the role of this mark in transcriptional activation. The global levels of methylation were higher in the non-morphogenic callus, which indicated increased gene activation compared to the morphogenic callus. We also employed ChIP to analyse the changes in the enrichment of this epigenetic mark on the cell wall-related genes in both calli types during the course of the passage. Enrichment of H3K4me3 on cell wall genes was specific for callus type, suggesting that the role of this mark in cell-wall remodelling is complex and involved in many processes related to dedifferentiation and redifferentiation. This intricacy of the cell wall composition was supported by the immunohistochemical analysis of the cell wall epitopes' distribution of pectins and extensins. Together, these data give a novel insight into the involvement of H3K4me3 in the regeneration processes in F. tataricum in vitro callus tissue culture.

MmpL3, Wag31, and PlrA are involved in coordinating polar growth with peptidoglycan metabolism and nutrient availability.

This study is performed in Mycobacterium smegmatis, which is used as a model to understand the basic physiology of pathogenic mycobacteria such as Mycobacterium tuberculosis. In this work, we examine the function and regulation of three proteins involved in regulating cell wall elongation in mycobacterial cells, which occurs at the cell tips or poles. We find that Wag31, a regulator of polar elongation, works partly through the regulation of MmpL3, a transporter of cell wall constituents and an important drug target. Our work suggests that, beyond its transport function, MmpL3 has another function in controlling cell wall synthesis broadly in response to stress.

Suitability of an ornamental tree, Sorbus alnifolia, as a source of industrial wood: Properties and the juvenile to mature transition

Ornamental trees are being promoted to supplement wood for industrial applications in China. To determine the wood utilization potential of an ornamental tree species, Sorbus alnifolia, this study investigated the radial variation of anatomical characteristics of its wood cross-section. The results showed that S. alnifolia is porous with high cell wall percentage, fiber percentage, and vessel percentage, small fiber and vessel sizes, and low vessel frequency. The transition age between juvenile wood and mature wood is 7 to 11 years for vessels, 12 to 16 years for axial parenchyma, and 18 to 24 years for fibers. Mature wood exhibits a higher percentage of cell walls, thicker fiber walls, and a lower percentage of vessels than juvenile wood. This result implies that wood is easy to dry, has strong permeability, good physical and mechanical properties, and a high fiber yield.

Ventral body wall closure: Mechanistic insights from mouse models and translation to human pathology.

The ventral body wall (VBW) that encloses the thoracic and abdominal cavities arises by extensive cell movements and morphogenetic changes during embryonic development. These morphogenetic processes include embryonic folding generating the primary body wall; the initial ventral cover of the embryo, followed by directed mesodermal cell migrations, contributing to the secondary body wall. Clinical anomalies in VBW development affect approximately 1 in 3000 live births. However, the cell interactions and critical cellular behaviors that control VBW development remain little understood. Here, we describe the embryonic origins of the VBW, the cellular and morphogenetic processes, and key genes, that are essential for VBW development. We also provide a clinical overview of VBW anomalies, together with environmental and genetic influences, and discuss the insight gained from over 70 mouse models that exhibit VBW defects, and their relevance, with respect to human pathology. In doing so we propose a phenotypic framework for researchers in the field which takes into account the clinical picture. We also highlight cases where there is a current paucity of mouse models for particular clinical defects and key gaps in knowledge about embryonic VBW development that need to be addressed to further understand mechanisms of human VBW pathologies.

Enhancing emulsion stability and adsorption of Pickering emulsions using alkylated cellulose nanofibers

Cellulose nanofibers (CNFs) extracted from plant cell walls form a three-dimensional network in water and stabilize the dispersion of droplets, which are used as emulsion stabilizers. In this study, four types of alkyl-grafted CNFs (ACNFs) with different lengths of dialkyl chains (diC4, diC6, diC8, and diC10) were synthesized by the surface modification of 2,2,6,6-tetramethylpiperidine-1-oxyl radical (TEMPO)-oxidized CNF (TOCNF) to improve the stability of oil-in-water Pickering emulsions (PEs). The ACNF-stabilized emulsions with a 20/80 ratio of oil/water were prepared using two different oils (olive oil and eucalyptus oil) and the ACNF 1.0 % dispersion, and the changes over time were investigated at approximately 25 °C. Compared to TOCNF, the ACNFs provided more stable olive oil-PE (o-PE) and eucalyptus oil-PE (e-PE) although the stabilization behaviors of o-PE and e-PE differed. Olive oil, which contained rich long-chain fatty acids, was more likely to interact with the ACNFs, thereby promoting the interfacial adsorption of oil droplets. On the other hand, eucalyptus oil is mainly composed of 1,8-cineole and does not have alkyl chains, so the interaction with the ACNFs was weak and therefore, sufficient interfacial adsorption could not be obtained. The results of the particle size distribution and viscosity measurements also showed that the ACNFs contributed significantly to the stabilization of o-PE. Based on the overall experimental results, it was determined that the diC6-ACNF gave the most significant stability improvement. Therefore, we designed an alkylated CNF that mimics diC6-ACNF for MD simulations. MD simulations showed that hydrophobic modification created a hydrophobic interaction between the alkyl chain and dodecane molecules as the oil, leading to the enhancing adsorption of ACNF at the oil droplet interfaces.

Activated carbon fiber loaded nano zero-valent iron for Microcystis aeruginosa removal: Performance and mechanisms

Cyanobacterial blooms caused by Microcystis aeruginosa threaten environmental safety and daily life. In this study, an activated carbon fiber-supported nano zero-valent iron composite (ACF-nZVI) was developed to remove Microcystis aeruginosa. The results showed that nZVI was evenly distributed on the activated carbon fibers, preventing aggregation and oxidation. ACF-nZVI achieved a removal efficiency of more than 90 % within a pH range of 3–7. During the reaction, H2O2, which was generated by Fe0, was activated to form ·OH and ·O–2, which dismantled antioxidant enzymes and induced lipid peroxidation. Additionally, ACF-nZVI destroyed the cell wall and membrane, resulting in protein and humus leakage and causing 92.34 % cell damage and death. In this study, an environmentally friendly and stable nanomaterial was developed, offering a novel approach for the safe, cost-effective, and efficient removal of cyanobacteria.

Impact of particle size on the functionality of corn-derived dietary fiber-phenolic acid complexes

Milling and sieving were applied to modify corn-derived cell wall dietary fiber-phenolic acid complexes (CWDFPC) to enhance their functionality and gut fermentability. The physicochemical properties of three modified CWDFPCs (CWDFPC-M1, CWDFPC-M2, and CWDFPC-M3) were characterized, showing changes in particle size (430–73.55 μm) and bulk density (0.29–0.57 g/mL). Sieving altered the composition, with CWDFPC-M1 and CWDFPC-M3 exhibiting higher bound phenolic contents than CWDFPC-M2. Increased water holding capacity indicated improved functionalities. Modified CWDFPCs exhibited a 4-fold increase in glucose adsorption capacity, higher phenolic acid release during gastrointestinal digestion in vitro, and a greater proportion of short-chain fatty acids in fecal fermentation in vitro. Hemicellulose from CWDFPC-M3 was primarily composed of →4)-β-Xylp-(1→ and →3)-β-Xylp-(1→, and →3,4)-β-Xylp-(1→, with branches possibly including →5)-α-Araf-(1→ and →3)-α-Araf-(1→ units. These modifications highlight the potential of milling and sieving to convert CWDFPCs into valuable functional food ingredients.

Universal Receptive System as a novel regulator of transcriptomic activity of Staphylococcus aureus.

Our previous studies revealed the existence of a Universal Receptive System that regulates interactions between cells and their environment. This system is composed of DNA- and RNA-based Teazeled receptors (TezRs) found on the surface of prokaryotic and eukaryotic cells, as well as integrases and recombinases.. In the current study, we aimed to provide further insight into the regulatory role of TezR and its loss in Staphylococcus aureus gene transcription. To this end, transcriptomic analysis of S. aureus MSSA VT209 was performed following the destruction of TezRs. Bacterial RNA samples were extracted from nuclease-treated and untreated S. aureus MSSA VT209. After destruction of the DNA-based-, RNA-, or combined DNA- and RNA-based TezRs of S. aureus , 103, 150, and 93 genes were significantly differently expressed, respectively. The analysis revealed differential clustering of gene expression following the loss of different TezRs, highlighting individual cellular responses following the loss of DNA- and RNA-based TezRs. KEGG pathway gene enrichment analysis revealed that the most upregulated pathways following TezR inactivation included those related to energy metabolism, cell wall metabolism, and secretion systems. Some of the genetic pathways were related to the inhibition of biofilm formation and increased antibiotic resistance, and we confirmed this at the phenotypic level using in vitro studies. The results of this study add another line of evidence that the Universal Receptive System plays an important role in cell regulation, including cell responses to the environmental factors of clinically important pathogens, and that nucleic acid-based TezRs are functionally active parts of the extrabiome.

Indication of radioactive contamination of forest ecosystems in the zone of the east Ural radioactive trace using methods of quantitative wood anatomy

The anatomical structure of the annual rings of Scots pine, formed before and after the Kyshtym accident, is analyzed. In trees growing closer to the central axis of the East Ural radioactive trace (EURT), a decrease in the number of cells in the annual ring, as well as a decrease in the diameter of the lumens and the thickness of the cell walls, was noted. It is assumed that radiation-induced damage to the photo-assimilation apparatus of trees led to disturbances in physiological processes that were reflected in the anatomical structure of wood.

Translocon Subunits of the COP9 Signalosome Complex Are a Central Hub for Regulating Multiple Photoresponsive Processes and Autophagic Flux in Magnaporthe oryzae.

Photodependent processes, including circadian rhythm, autophagy, ubiquitination, neddylation/deneddylation, and metabolite biosynthesis, profoundly influence microbial pathogenesis. Although a photomorphogenesis signalosome (COP9/CSN) has been identified, the mechanism by which this large complex contributes to the pathophysiological processes in filamentous fungi remains unclear. Here, we identified eight CSN complex subunits in the rice blast fungus Magnaporthe oryzae and functionally characterized the translocon subunits containing a nuclear export or localization signal (NES/NLS). Targeted gene replacement of these CSN subunits, including MoCSN3, MoCSN5, MoCSN6, MoCSN7, and MoCSN12, attenuated vegetative growth and conidiation and rendered the deletion strains nonpathogenic. MoCSN7 deletion significantly suppressed arachidonic acid catabolism, and compromised cell wall integrity in M. oryzae. Surprisingly, we also discovered that MoCSN subunits, particularly MoCsn7, are required for the cAMP-dependent regulation of autophagic flux. Therefore, MoCSN significantly contributes to morphological, physiological, and pathogenic differentiation in M. oryzae by fostering cross-talk between multiple pathways.

Extraction and Synthesis of Typical Carotenoids: Lycopene, β-Carotene, and Astaxanthin.

Carotenoids are tetraterpene compounds acting as precursors to vitamin A, with functions that include protecting eyesight, enhancing immunity, promoting cell growth and differentiation, and providing antioxidative benefits. Lycopene, β-carotene, and astaxanthin are particularly critical for health and have diverse applications in food, health products, and medicine. However, natural carotenoids are encased within cell structures, necessitating mechanical methods to disrupt the cell wall for their extraction and purification-a process often influenced by environmental conditions. Thus, improving the efficiency of carotenoid extraction from natural resources is of great interest. This review delves into the research progress made on the extraction processes, structures, and biological functions of carotenoids, focusing on lycopene, β-carotene, and astaxanthin. Traditional extraction methods primarily involve organic solvent-assisted mechanical crushing. With deeper research and technological advancements, more environmentally friendly solvents, advanced machinery, and suitable methods are being employed to enhance the extraction and purification of carotenoids. These improvements have significantly increased extraction efficiency, reduced preparation time, and lowered production costs, laying the groundwork for new carotenoid product developments.