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In this work, coacervation in primary amines solutions with hydrophobic natural deep eutectic solvents based on terpenoids and carboxylic acids was demonstrated for the first time. A liquid-phase microextraction approach was developed based on supramolecular solvent formation with primary amine acting as amphiphile and hydrophobic deep eutectic solvent making up mixed vesicles and serving as coacervation agent. Such supramolecular solvents could be used to separate wide range of substances from different aqueous media, such as food products, biological liquids and wastewaters. It is important that both hydrophobic and ionic interactions with supramolecular aggregates take place ensuring synergetic effect and better extraction ability, which is significant in separating relatively polar analytes. Different primary amines and deep eutectic solvents were investigated for liquid-phase microextraction of proof-of-concept amphoteric analyte (enrofloxacin, widely used veterinary fluoroquinolone antibiotic) and its determination by high-performance liquid chromatography with fluorescence detection using Shimadzu LC-20 Prominence chromatograph and RF-20A fluorescence detector. It was found that the supramolecular solvent based on 1-nonylamine, formed after addition of a deep eutectic solvent based on menthol and hexanoic acid (molar ratio of 1:1), provided maximum extraction recovery (85 %) and maximum enrichment factor (34). To characterize the extraction system, the composition of the phases was investigated, and cryo-transmission electron microscopy images were obtained. Vesicular aggregates were observed in the supramolecular solvent. The extraction mechanism was proposed in terms of formation of mixed aggregates to capture the analyte. Limit of detection was found to be 7 μg kg -1 , while linear range of 20-250 μg kg -1 was established. Relative standard deviation values were lower than 7 %. Relative bias did not exceed 12 %., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Trichomes are specialized hair-like structures in the epidermal cells of the above-ground parts of plants and help to protect them from pests and pathogens, and produce valuable metabolites. Chrysanthemum morifolium, which is used in tea products, has both ornamental and medicinal value; however, it is susceptible to infection by the fungus Alternaria alternata, which can result in substantial economic losses. Increasing the density of glandular trichomes enhances disease resistance and improves the production of medicinal metabolites in chrysanthemums, and jasmonate (JA) is known to promote the formation of trichomes in various plants. However, it remains unclear whether glandular trichomes in chrysanthemums are regulated by JA. In addition, grafting, a technique that can improve plant resistance to biotic stresses, has been poorly examined for its impact on glandular trichomes, terpenoids, and disease resistance. In this study, we demonstrate that grafting with Artemisia vulgaris rootstocks improves the resistance of chrysanthemum scions to A. alternata. Heterografted chrysanthemums exhibited higher trichome density and terpenoid content compared to self-grafted counterparts. Transcriptome analysis highlighted the significant role of CmJAZ1-like in disease resistance in heterografted chrysanthemums. Lines overexpressing CmJAZ1-like exhibited sensitivity to A. alternata, and this was characterized by reduced glandular trichome density and limited terpenoid content. Conversely, CmJAZ1-like silenced lines exhibited resistance to A. alternata and showed increased glandular trichome density and terpenoid content. Higher JA content was found in the heterografted chrysanthemum scions compared to self-grafted ones. Furthermore, we established that JA promoted the development of glandular trichomes and the synthesis of terpenoids while also inducing the degradation of CmJAZ1-like proteins in chrysanthemums. Our findings suggest that higher JA increases trichome density and terpenoid content, thereby enhancing resistance to A. alternata by regulating CmJAZ1-like in heterografted chrysanthemums., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Diterpenoid natural products serve critical functions in plant development and ecological adaptation and many diterpenoids have economic value as bioproducts. The family of class II diterpene synthases catalyzes the committed reactions in diterpenoid biosynthesis, converting a common geranylgeranyl diphosphate precursor into different bicyclic prenyl diphosphate scaffolds. Enzymatic rearrangement and modification of these precursors generates the diversity of bioactive diterpenoids. We report the crystal structure of Grindelia robusta 7,13-copalyl diphosphate synthase, GrTPS2, at 2.1 Å of resolution. GrTPS2 catalyzes the committed reaction in the biosynthesis of grindelic acid, which represents the signature metabolite in species of gumweed (Grindelia spp., Asteraceae). Grindelic acid has been explored as a potential source for drug leads and biofuel production. The GrTPS2 crystal structure adopts the conserved three-domain fold of class II diterpene synthases featuring a functional active site in the γβ-domain and a vestigial ɑ-domain. Substrate docking into the active site of the GrTPS2 apo protein structure predicted catalytic amino acids. Biochemical characterization of protein variants identified residues with impact on enzyme activity and catalytic specificity. Specifically, mutagenesis of Y457 provided mechanistic insight into the position-specific deprotonation of the intermediary carbocation to form the characteristic 7,13 double bond of 7,13-copalyl diphosphate., Competing Interests: Conflict of interest statement The authors declare no competing interests, (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Amomum tsao-ko is an important aromatic crop used in medicines and food. It can be categorized into three main types based on the fruit shape: long (L), oval (O), and round (R). However, limited information is available on the volatile substances present in differently shaped A. tsao-ko fruits. This study investigated the characteristics and biosynthesis of volatile organic compounds (VOCs) in fresh and dried A. tsao-ko fruits of different shapes using widely targeted volatilomics and transcriptome analyses., Results: In total, 978 VOCs, primarily terpenoids, esters, and heterocyclic compounds, were detected. The number of differentially accumulated volatile organic compounds (DAVOCs) in dried fruits of various shapes was significantly higher than that in fresh fruits, with terpenoids, esters, and heterocyclic compounds accounting for approximately 50% of the total DAVOCs. Notably, α-phellandrene, identified as a shared differential accumulated terpenoid across various fruit shapes, was detected in both fresh and dried fruits. Through transcriptome analysis, 40 candidate genes implicated in the terpenoid biosynthesis pathway were screened. An integrated analysis of the metabolome and transcriptome revealed that the structural genes HMGR-2, TPS7, TPS5-10, TPS21-3, TPS21-5, TPS21-6, TPS21-7, and TPS21-9, along with 81 transcription factors (including 17 NACs, 16 MYBs, 16 AP2/ERFs, 13 WRKYs, 13 bHLHs, and 6 bZIPs), co-regulate the biosynthesis of volatile terpenoids., Conclusions: This study expands our understanding of the volatile metabolism profile of A. tsao-ko and provides a solid foundation for future investigations of the mechanisms governing fruit quality., (© 2024. The Author(s).)

Basil ( Ocimum basilicum L.) is a widely used culinary herb. In this study, ethanol, dichloromethane, and sunflower oil were used separately as solvents with distinct polarities for the extraction of basil aerial parts to simulate the different polarity conditions in domestic food processing. The oil extract (OE) was re-extracted with acetonitrile, and the chemical composition, antioxidant potential, and antimicrobial activities of the ethanol (EE), dichloromethane (DCME), and acetonitrile (ACNE) extracts were determined. A total of 109 compounds were tentatively identified in EE, DCME, and ACNE by HPLC-DAD/ESI-ToF-MS. Fatty acids were present in all extracts. Phenolic acids and flavonoids dominated in EE. DCME was characterised by triterpenoid acids, while diterpenoids were mainly found in ACNE. The extracts were analysed for their antioxidant capacity using the 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) assay. EE and DCME showed significant radical scavenging potential. Antimicrobial activity was explored in eight bacterial, two yeast, and one fungal species. All extracts exhibited high antifungal activity, comparable to or better than that of the commercial drug nistatin. Antibacterial activities were notable for EE and ACNE, while DCME showed no activity against bacteria in the applied concentration ranges. The different polarities of the solvents led to distinctive phytochemical compositions and bioactivities in the extracts.

Background: Phalaenopsis bellina, an orchid native to Borneo, is renowned for its unique appearance. It releases distinct fragrances, which have been linked to the presence of terpenoids. However, the identification and study of sesquiterpene synthase in P. bellina remain limited. In this study, we examines the functional characterisation of terpene synthase (TPS) from P. bellina, known as PbTS, through recombinant protein expression and its manifestation in the flower., Methods and Results: Gene annotation of PbTS revealed that the inferred peptide sequence of PbTS comprises 1,680 bp nucleotides encoding 559 amino acids with an estimated molecular mass of 65.2 kDa and a pI value of 5.4. A similarity search against GenBank showed that PbTS shares similarities with the previously published partial sequence of P. bellina (ABW98504.1) and Phalaenopsis equestris (XP_020597359.1 and ABW98503.1). Intriguingly, the phylogenetic analysis places the PbTS gene within the TPS-a group. In silico analysis of PbTS demonstrated stable interactions with farnesyl pyrophosphate (FPP), geranyl pyrophosphate (GPP), and geranylgeranyl pyrophosphate (GGPP). To verify this activity, an in vitro enzyme assay was performed on the PbTS recombinant protein, which successfully converted FPP, GPP, and GGPP into acyclic sesquiterpene β-farnesene, yielding approximately 0.03 mg/L. Expressional analysis revealed that the PbTS transcript was highly expressed in P. bellina, but its level did not correlate with β-farnesene levels across various flowering time points and stages., Conclusion: The insights gained from this study will enhance the understanding of terpenoid production in P. bellina and aid in the discovery of novel fragrance-related genes in other orchid species., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Plant-associated streptomycetes play important roles in plant growth and development. However, knowledge of volatile-mediated crosstalk between Streptomyces spp. and plants remains limited. In this study, we investigated the impact of volatiles from nine endophytic Streptomyces strains on the growth and development of plants. One versatile strain, Streptomyces setonii WY228, was found to significantly promote the growth of Arabidopsis thaliana and tomato seedlings, confer salt tolerance, and induce early flowering and increased fruit yield following volatile treatment. Analysis of plant growth-promoting traits revealed that S. setonii WY228 could produce indole-3-acetic acid, siderophores, ACC deaminase, fix nitrogen, and solubilize inorganic phosphate. These capabilities were further confirmed through genome sequencing and analysis. Volatilome analysis indicated that the volatile organic compounds emitted from ISP-2 medium predominantly comprised sesquiterpenes and 2-ethyl-5-methylpyrazine. Further investigations showed that 2-ethyl-5-methylpyrazine and sesquiterpenoid volatiles were the primary regulators promoting growth, as confirmed by experiments using the terpene synthesis inhibitor phosphomycin, pure compounds, and comparisons of volatile components. Transcriptome analysis, combined with mutant and inhibitor studies, demonstrated that WY228 volatiles promoted root growth by activating Arabidopsis auxin signaling and polar transport, and enhanced root hair development through ethylene signaling activation. Additionally, it was confirmed that volatiles can stimulate plant abscisic acid signaling and activate the MYB75 transcription factor, thereby promoting anthocyanin synthesis and enhancing plant salt stress tolerance. Our findings suggest that aerial signaling-mediated plant growth promotion and abiotic stress tolerance represent potentially overlooked mechanisms of Streptomyces-plant interactions. This study also provides an exciting strategy for the regulation of plant growth and the improvement of horticultural crop yields within sustainable agricultural practices., Competing Interests: Declaration of Competing Interest The authors report no declarations of interest., (Copyright © 2024 Elsevier GmbH. All rights reserved.)

Terpenoids are widely used in the food, beverage, cosmetics, and pharmaceutical industries. Microorganisms have been extensively studied for terpenoid production. In yeast, the introduction of the mevalonate (MVA) pathway in organelles in addition to the augmentation of its own MVA pathway have been challenging. Introduction of the MVA pathway into mitochondria is considered a promising approach for terpenoid production because acetyl-CoA, the starting molecule of the MVA pathway, is abundant in mitochondria. However, mitochondria comprise only a small percentage of the entire cell. Therefore, we hypothesized that increasing the total mitochondrial volume per cell would increase terpenoid production. First, we ascertained that the amounts of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), the final molecules of the MVA pathway, were 15-fold higher of the strain expressing the MVA pathway in mitochondria than in the wild-type yeast strain. Second, we found that different deletion mutants induced different mitochondrial volumes by measuring the mitochondrial volume in various deletion mutants affecting mitochondrial morphology; for example,Δmdm32 increased mitochondrial volume, and Δfzo1 decreased it. Finally, the effects of mitochondrial volume on amounts of IPP/DMAPP and terpenoids (squalene or β-carotene) were investigated using mutants harboring large or small mitochondria expressing the MVA pathway in mitochondria. Amounts of IPP/DMAPP and terpenoids (squalene or β-carotene) increased when the mitochondrial volume expanded. Introducing the MVA pathway into mitochondria for terpenoid production in yeast may become more attractive by enlarging the mitochondrial volume. KEY POINTS: • IPP/DMAPP content increased in the strain expressing the MVA pathway in mitochondria • IPP/DMAPP and terpenoid contents are positively correlated with mitochondrial volume • Enlarging the mitochondria may improve mitochondria-mediated terpenoid production., (© 2024. The Author(s).)

Terpenoids possess significant physiological activities and are rich in essential oils. Some terpenoids have chiral centers and could form enantiomers with distinct physiological activities. Therefore, the extraction and separation of terpenoids enantiomers are very important and have attracted extensive attention in recent years. Meanwhile, the specific distribution and enantiomer excess results (the excess of one enantiomer over the other in a mixture of enantiomers) could be used as quality markers for illegitimate adulteration, origin identification, and exploring component variations and functional interrelations across different plant tissues. In this study, an overview of the progress in the extraction of terpenoids from essential oils and the separation of their enantiomers over the past two decades has been made. Extraction methods were retrieved by the resultant network visualization findings. The results showed that the predominant methods are hydrodistillation, solvent-free microwave extraction, headspace solid-phase microextraction and supercritical fluid extraction methods. GC-MS combined with chiral chromatography columns is commonly used for the separation of enantiomers, while 2D GC is found to have stronger resolution ability. Finally, some prospects for future research directions in the extraction and separation identification of essential oils are proposed., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Introduction: Ibrexafungerp is a new triterpenoid antifungal agent with activity against a variety of fungal species, including Aspergillus spp. and echinocandin-resistant Candida spp., Areas Covered: This evaluation will summarize currently available clinical evidence on the use of ibrexafungerp in the treatment/prevention of vulvovaginal candidiasis (VVC) and detail the mechanism of action, pharmacokinetic/pharmacodynamic parameters, and ongoing/latest research involving ibrexafungerp., Expert Opinion: The evidence involving the utilization of ibrexafungerp for the treatment of VVC shows that it is superior when compared to placebo and has comparable clinical cure rates when compared with fluconazole. Ibrexafungerp demonstrates reliable coverage against several Candida spp. including echinocandin-resistant strains, Candida auris , and Aspergillus spp. For VVC, a dose of 300 mg (two 150 mg tablets) twice daily is recommended and does not require dose adjustments based on renal or hepatic function. The use of ibrexafungerp outside of VVC is currently under study with several ongoing trials showing promising interim data.

Introduction: Croton crassifolius Geisel. (CCG) is a traditional Chinese medicine widely used in South China. It has various pharmacological effects and is often used in treating rheumatoid arthritis and gastric and duodenal ulcers. However, the chemical characteristics and its effective constituents are still scarcely studied., Objective: To determine the phytochemical profile of the CCG extract and to investigate the chemical characteristics of terpenoids extracted from rat plasma following oral administration of CCG extract based on UPLC-Q/TOF-MS. Moreover, six terpenoids in CCG were quantified, and in vivo pharmacokinetic behavior after oral CCG extract was further explored., Results: In total, 56 terpenoids were tentatively identified in the CCG extract and 16 terpenoids were detected in rat plasma after oral CCG extract. In addition, the contents of six terpenoids in CCG were clarified. The plasma quantification method of six terpenoids was further established, validated, and confirmed to have good sensitivity and specificity. The six analytes exhibited excellent linearity in respective concentration ranges (r ≥ 0.998). The intra-day and inter-day precisions relative standard deviation (RSD, %) were less than 11.27%, and the accuracies ranged from -7.06% to 9.91%. Stability, extraction recovery, and matrix effect in plasma were within the required limits (RSD < 15%)., Conclusion: A total of 56 terpenoids were identified in CCG and 16 prototype components in plasma after oral CCG. The validated quantitative method was successfully applied to the simultaneous determination of six major terpenoids in plasma. The pharmacokinetic parameters are clarified, which can guide the clinical application of CCG., (© 2024 John Wiley & Sons Ltd.)

Introduction: Camphora longepaniculata , a crucial commercial crop and a fundamental component of traditional Chinese medicine, is renowned for its abundant production of volatile terpenoids. However, the lack of available genomic information has hindered pertinent research efforts in the past., Methods: To bridge this gap, the present study aimed to use PacBio HiFi, short-read, and highthroughput chromosome conformation capture sequencing to construct a chromosome-level assembly of the C. longepaniculata genome., Results and Discussion: With twelve chromosomes accounting for 99.82% (766.69 Mb) of the final genome assembly, which covered 768.10 Mb, it was very complete. Remarkably, the assembly's contig and scaffold N50 values are exceptional as well-41.12 and 63.78 Mb, respectively-highlighting its excellent quality and intact structure. Furthermore, a total of 39,173 protein-coding genes were predicted, with 38,766 (98.96%) of them being functionally annotated. The completeness of the genome was confirmed by the Benchmarking Universal Single-Copy Ortholog evaluation, which revealed 99.01% of highly conserved plant genes. As the first comprehensive assembly of the C. longepaniculata genome, it provides a crucial starting point for deciphering the complex pathways involved in terpenoid production. Furthermore, this excellent genome serves as a vital resource for upcoming research on the breeding and genetics of C. longepaniculata ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Yan, Zhu, Cao, Meng, Li, Zhang, Liu, Wang, Feng, Soaud, Elhamid, Heakel, Wei, El-Sappah and Ru.)

In plants, secondary metabolites change in response to environmental conditions. These changes co-regulate resilience to stressful environmental conditions, plant growth and development, and interactions between plants and the wider ecosystem, while also affecting soil carbon storage and atmospheric and climatic conditions. The objective of this study was to determine the association between UV exposure and the contents of key metabolites, including amino acids, phenolics, flavonoids, terpenoids, carotenoids, tocopherols, and phytosterols. Mentha spicata plantlets were grown in tissue culture boxes for 30 days and then exposed to a low dose of broadband UV-B (291-315 nm; 2.8 kJm -2 biologically effective UV) enriched light for eight days. Metabolite contents were quantified either immediately after the final UV exposure, or after seven days of recovery under photosynthetically active radiation. It was found that UV promoted the production of flavonoids (1.8-fold) ahead of phenolic acids (unchanged). Furthermore, the majority of monoterpenes and sesquiterpenes, constituents of valuable mint essential oil, were significantly increased through UV treatment (up to 90-fold for α-linalool). In contrast, the contents of carotenoids and tocopherols did not increase following UV exposure. A comparison between plants sampled immediately after UV exposure and after seven days of recovery showed that there was an overall increase in the content of carotenoids, mono- and sesquiterpenes, phenolics, and amino acids following recovery, while the contents of sterols and tocopherols decreased. These UV-induced changes in metabolite profile may have important consequences for agriculture, ecology, and even the global climate, and they also provide an exciting opportunity to enhance crop value, facilitating the development of improved products with higher levels of essential oils and added benefits of enhanced flavour, colour, and bioactive content., Competing Interests: The authors declare no conflict of interest.

Background: Despite the availability of chemotherapy drugs such as 5-fluorouracil (5-FU), the treatment of some cancers such as gastric cancer remains challenging due to drug resistance and side effects. This study aimed to investigate the effect of celastrol in combination with the chemotherapy drug 5-FU on proliferation and induction of apoptosis in human gastric cancer cell lines (AGS and EPG85-257)., Materials and Methods: In this in vitro study, AGS and EPG85-257 cells were treated with different concentrations of celastrol, 5-FU, and their combination. Cell proliferation was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. The synergistic effect of 5-FU and celastrol was studied using Compusyn software. The DNA content at different phases of the cell cycle and apoptosis rate was measured using flow cytometry., Results: Co-treatment with low concentrations (10% inhibitory concentration (IC10)) of celastrol and 5-FU significantly reduced IC50 ( p < 0.05) so that 48 h after treatment, IC50 was calculated at 3.77 and 6.9 μM for celastrol, 20.7 and 11.6 μM for 5-FU, and 5.03 and 4.57 μM for their combination for AGS and EPG85-257 cells, respectively. The mean percentage of apoptosis for AGS cells treated with celastrol, 5-FU, and their combination was obtained 23.9, 41.2, and 61.9, and for EPG85-257 cells 5.65, 46.9, and 55.7, respectively. In addition, the 5-FU and celastrol-5-FU combination induced cell cycle arrest in the synthesis phase., Conclusions: Although celastrol could decrease the concentration of 5-fluorouracil that sufficed to suppress gastric cancer cells, additional studies are required to arrive at conclusive evidence on the anticancer effects of celastrol., Competing Interests: The authors declare that they have no conflicts of interest to report regarding the present study., (© 2024 Moradi et al.)

As global agricultural challenges intensify, particularly drought stress, the exploration of innovative strategies for crop resilience has become crucial. This study focuses on the role of the microbial endophyte metabolite Albaflavenone in enhancing drought resistance in tomato (Solanum lycopersicum L.) through the activation of the SlMAPK1 protein in the MAPK pathway. To computationally analyze the interaction between Albaflavenone and SlMAPK1 and to elucidate the potential enhancement of drought tolerance in tomato plants through this interaction. We utilized molecular docking, homology modeling, and molecular dynamics simulations to investigate the binding affinities and interaction dynamics between SlMAPK1 and Albaflavenone. Functional network analysis was employed to examine protein-protein interactions within the MAPK pathway, while the MM-GBSA method was used to calculate binding free energies. Our computational analyses revealed that Albaflavenone exhibited a high binding affinity to SlMAPK1 with a binding energy of - 8.9 kcal/mol. Molecular dynamics simulations showed this interaction significantly stabilized SlMAPK1, suggesting enhanced activity. Specifically, the root mean square deviation (RMSD) of the Albaflavenone-SlMAPK1 complex stabilized at around 3.1 Å, while the root mean square fluctuations (RMSF) indicated consistent amino acid conformations. Additionally, the radius of gyration (R g ) analysis demonstrated minimal variance, suggesting a compact and stable protein-ligand complex. The significant binding affinity between Albaflavenone and SlMAPK1 highlights the potential of leveraging plant-microbe interactions in developing sustainable agricultural practices. These findings also demonstrate the effectiveness of computational methods in dissecting complex biological interactions, contributing to a deeper understanding of plant resilience strategies against environmental stresses., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Sclareol, a diterpene alcohol, is the most common starting material for the synthesis of ambrox, which serves as a sustainable substitute for ambergris, a valuable fragrance secreted by sperm whales. Sclareol has also been proposed to possess antibacterial, antifungal, and anticancer activities. However, in nature, sclareol is only produced by a few plant species, including Cistus creticus , Cleome spinosa , Nicotiana glutinosa , and Salvia sclarea , which limits its commercial application. In this study, we cloned the two genes responsible for sclareol biosynthesis in S. sclarea , labda-13-en-8-ol diphosphate synthase ( LPPS ) and sclareol synthase ( SS ), and overexpressed them in tobacco ( Nicotiana tabacum L.). The best transgenic tobacco lines accumulated 4.1 μg/cm 2 of sclareol, which is comparable to the sclareol production of N. glutinosa , a natural sclareol producer. Thus, sclareol synthesis in tobacco represents a potential alternative means for the production of this high-value compound.

Numerous herbal products have been the subject of research regarding their potential role in cancer prevention or adjuvant therapy. Pistacia atlantica and its main phytochemicals have garnered significant attention for their potential anti-cancer effects. The study aimed to assess the growth inhibitory effects of P. atlantica essential oil (PAEO) on MKN-45 and AGS cells. This study quantified the volatile compounds in PAEO using Gas Chromatography-Mass Spectrometry (GC-MS). Subsequently, MKN-45 and AGS cells were treated with varying concentrations of PAEO (5%, 2.5%, 1.25%, 0.625%, 0.3125%, 0.156%, 0.0781%, 0.0391%, 0.0195%) for 24 h. Cell viability was evaluated through the MTT assay. The impact of PAEO on gene expression was investigated by quantifying the mRNA levels of Bax and Bcl2 in the various experimental groups using quantitative Real-Time PCR (qRT-PCR) analysis. Additionally, flow cytometry was utilized to evaluate apoptosis in the treated cells. The analysis of PAEO revealed that α-pinene was the predominant monoterpene, constituting 87.9% of the oil composition. The cytotoxic effects of PAEO were evaluated, and it was found that the oil significantly reduced the viability of MKN-45 and AGS cells. The IC 50 for MKN-45 cells was determined to be 1.94 × 10 -3 % after 24 h of treatment, while for AGS cells the IC 50 was 2.8 × 10 -3 % after 24 h. Additionally, the research revealed that PAEO triggered a notable rise in apoptotic cells in both AGS and MKN-45 cell lines. Moreover, at the molecular level, the findings indicated an increase in Bax expression and a decrease in Bcl2 mRNA expression, providing further evidence of the induction of apoptosis in both MKN-45 and AGS cell lines following PAEO treatment. The findings of this study offer evidence supporting the cytotoxic effects of PAEO on gastric cancer cell lines by promoting apoptosis. The findings suggest that PAEO may offer potential as a therapeutic candidate in managing and treating gastric cancer., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Terpenoids are one of the largest class of natural products with diverse structures and activities. This enormous diversity is embedded in enzymes called terpene synthases (TSs), which generate diverse terpene skeletons via sophisticated cyclization cascades. In addition to the many highly selective TSs, there are many promiscuous TSs that accept multiple prenyl substrates, or even noncanonical ones, with 6, 7, 8, 11, and 16 carbon atoms, synthesized via chemical approaches, C -methyltransferases, or engineered lepidopteran mevalonate pathways. The substrate promiscuity of TSs not only expands the structural diversity of terpenes but also highlights their potential for the discovery of novel terpenoids via combinatorial biosynthesis. In this review, we focus on the current knowledge on multisubstrate terpene synthases (MSTSs) and highlight their potential applications., (Copyright © 2024, Li and Tao.)

The study investigated the impact of intraspecific plant chemodiversity on plant growth and reproductive traits at both the plant and plot levels. It also aimed to understand how chemodiversity at stand level affects ecosystem functioning and plant-plant interactions. We describe a biodiversity experiment in which we manipulated intraspecific plant chemodiversity at the plot level using six different chemotypes of common tansy (Tanacetum vulgare L., Asteraceae). We tested the effects of chemotype identity and plot-level chemotype richness on plant growth and reproductive traits and plot-level headspace emissions. The study found that plant chemotypes differed in growth and reproductive traits and that traits were affected by the chemotype richness of the plots. Although morphological differences among chemotypes became less pronounced over time, reproductive phenology patterns persisted. Plot-level trait means were also affected by the presence or absence of certain chemotypes in a plot, and the direction of the effect depended on the specific chemotype. However, chemotype richness did not lead to overyielding effects. Lastly, chemotype blends released from plant communities were neither richer nor more diverse with increasing plot-level chemotype richness, but became more dissimilar as they became more dissimilar in their leaf terpenoid profiles. We found that intraspecific plant chemodiversity is crucial in plant-plant interactions. We also found that the effects of chemodiversity on plant growth and reproductive traits were complex and varied depending on the chemotype richness of the plots. This long-term field experiment will allow further investigation into plant-insect interactions and insect community assembly in response to intraspecific chemodiversity., (© 2024 The Authors. Plant Biology published by John Wiley & Sons Ltd on behalf of German Society for Plant Sciences, Royal Botanical Society of the Netherlands.)

Aster tataricus L.f. is highly valued for its rich reserves of bioactive compounds. Our research focused on the identification of previously unreported compounds found within the ethanol extract of A. tataricus. Through meticulous spectroscopic analyses and computational methods like NMR calculations and ECD, we successfully elucidated the structures of five novel compounds termed tatarisides A-E (1-5), alongside two known compounds (6, 7). The anti-inflammatory assays conducted yielded noteworthy results, particularly in relation to compounds 1 and 5. These compounds exhibited significant potential in inhibiting the release of NO in LPS-induced RAW 264.7 cells, as evidenced by their respective IC 50 values of 17.81 ± 1.25 μM and 13.32 ± 0.84 μM. The discovery of these new compounds adds to the existing knowledge of A. tataricus's chemical composition and potential applications., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2023. Published by Elsevier B.V.)

Cyanobacteria are attractive hosts for photosynthetic terpenoid production, using CO 2 as the sole carbon source. Although the methylerythritol phosphate (MEP) pathway is superior to the mevalonate pathway for cyanobacterial terpenoid synthesis, the first reaction of the MEP pathway, which is catalyzed by 1-deoxy-d-xylulose-5-phosphate (DXP) synthase, involves complex regulation and carbon loss. Here, we constructed a direct route linking ribulose-5-phosphate (Ru5P) in the Calvin-Benson (CB) cycle with DXP in the MEP pathway in a cyanobacterium to increase the terpenoid yield from CO 2 and bypass the DXS-targeted regulations. By employing the adaptive laboratory evolution, we identified new RibB variants including RibB 90-92del with a high activity of synthesizing DXP from Ru5P. These RibB variants were introduced into Synechococcus elongatus , resulting in the significantly increased photosynthetic production of isopentenol. The 13 C tracer experiments demonstrated a direct carbon flow from Ru5P in the CB cycle to the MEP pathway; thus, this direct route was denoted as the Ru5P shunt. The strain harboring the Ru5P shunt produced 105.2 mg L -1 of isopentenol with an average rate of 17.5 mg L -1 d -1 under continuous light conditions, which is higher than those ever reported for five-carbon alcohol production by photoautotrophic microorganisms. Utilization of the Ru5P shunt in cyanobacterial cells also improved the pinene production, which demonstrates that this shunt can be used to enhance the photosynthetic production of diverse terpenoids.

Main Conclusion: Protein modeling, carbocation docking, and molecular dynamics along with structure-based mutability landscapes provided insight into taxadiene synthase catalysis (first step of the anticancer Taxol biosynthesis), protein structure-function correlations, and promiscuity. Plant terpenes belong to one of the largest and most diverse classes of natural products. This diversity is driven by the terpene synthase enzyme family which comprises numerous different synthases, several of which are promiscuous. Taxadiene synthase (TXS) is a class I diterpene synthase that catalyzes the first step in the biosynthesis pathway of the diterpene Taxol, an anticancer natural product produced by the Taxus plant. Exploring the molecular basis of TXS catalysis and its promiscuous potential garnered interest as a necessary means for understanding enzyme evolution and engineering possibilities to improve Taxol biosynthesis. A catalytically active closed conformation TXS model was designed using the artificial intelligence system, AlphaFold, accompanied by docking and molecular dynamics simulations. In addition, a mutability landscape of TXS including 14 residues was created to probe for structure-function relations. The mutability landscape revealed no mutants with improved catalytic activity compared to wild-type TXS. However, mutations of residues V584, Q609, V610, and Y688 showed high degree of promiscuity producing cembranoid-type and/or verticillene-type major products instead of taxanes. Mechanistic insights into V610F, V584M, Q609A, and Y688C mutants compared to the wild type revealed the trigger(s) for product profile change. Several mutants spanning residues V584, Q609, Y688, Y762, Q770, and F834 increased production of taxa-4(20),11(12)-diene which is a more favorable substrate for Taxol production compared to taxa-4(5),11(12)-diene. Finally, molecular dynamics simulations of the TXS reaction cascade revealed residues involved in ionization, carbocation stabilization, and cyclization ushering deeper understanding of the enzyme catalysis mechanism., (© 2024. The Author(s).)

Seaweeds are a treasure trove of natural secondary metabolites. Terpenoids extracted from seaweeds are shown to possess a variety of antitumor cellular activities. However, due to the complex and diverse structures of terpenoids, their therapeutic targets and complex mechanisms of action have not been clarified. The present study summarises the research on terpenoids from seaweeds in oncological diseases over the last 20 years. Terpenoids show different degrees of inhibitory effects on different types of tumor cells, suggesting that terpenoids in seaweeds may have potential antitumor disease potential. Terpenoids with potential antitumor activity and their mechanism of action are investigated using network pharmacology. A total of 125 terpenoids and 286 targets are obtained. Proto-oncogene tyrosine-protein kinase Src(SRC), Signal transducer and activator of transcription 3 (STAT3), Mitogen-activated protein kinase (MAPK3, MAPK1), Heat shock protein HSP 90-alpha (HSP90AA1), Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), and RAC-alpha serine/threonine-protein kinase (AKT1) are defined as core targets. According to GO function and Kyoto encyclopedia of genes and genomes(KEGG) enrichment analysis, terpenoids may affect the Phoshatidylinositol 3'-kinase (PI3K)-Akt signaling pathway, Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor resistance, Prostate cancer, MAPK signaling pathway, and Proteoglycans in cancer. In addition, the molecular docking results show that the selected terpenoids are all able to bind strongly to the active protein. Terpenoids may slow down the progression of cancer by controlling apoptosis, proliferation, and protein and enzyme binding., (© 2023 Wiley-VCH GmbH.)

Dehydrosqualene synthase (CrtM), as a squalene synthase-like enzyme from Staphylococcus aureus , can naturally utilize farnesyl diphosphate to produce dehydrosqualene (C 30 H 48 ). However, no study has documented the natural production of squalene (C 30 H 50 ) by CrtM. Here, based on an HPLC-Q-Orbitrap-MS/MS study, we report that the expression of crtM in vitro or in Bacillus subtilis 168 both results in the output of squalene, dehydrosqualene, and phytoene (C 40 H 64 ). Notably, wild-type CrtM exhibits a significantly higher squalene yield compared to squalene synthase (SQS) from Bacillus megaterium with an approximately 2.4-fold increase. Moreover, the examination of presqualene diphosphate's stereostructures in both CrtM and SQS enzymes provides further understanding into the presence of multiple identified terpenoids. In summary, this study not only provides insights into the promiscuity demonstrated by squalene synthase-like enzymes but also highlights a new strategy of utilizing CrtM as a potential replacement for SQS in cell factories, thereby enhancing squalene production.

In this review out of 300 selected articles 70 articles were evaluated, and the most significant compounds impacting COVID-19 and their mechanism of action were introduced. The compounds belong to four categories as follow: Phenolic, Flavonoid, Terpenoid, and Alkaloid compounds. In the phenol groups, the most effective compounds are scutellarin (suppressor of COVID-19 virus), thymol and carvacrol (the most inhibitory effect on COVID-19 virus), in the flavonoid groups, hesperdin (a strong inhibitor on COVID-19), in the terpenoids, methyl tanshinonate and sojil COVID-19 inhibitory effect) and 1,8-cineol (COVID-19 inhibitory effect) and in the last group, niglidine and quinoline alkaloid compounds (COVID-19 inhibitory effect) have been identified and introduced. These compounds have shown promising results due to their structure and effective mechanisms on COVID-19, so it can be an idea for researchers in this field to try to produce drugs by using natural compounds against the COVID-19 and Corona viruses.

A 1,2:3,4:9,10:9,19-tetraseco-cycloartane triterpene spiroketal lactone, pseudoamaolide P ( 1 ), two new labdane-type diterpenoids, pseudoamains A and B ( 2-3 ), and four known cembrane-type diterpenoids ( 4-7 ) were isolated from the seeds of Pseudolarix amabilis . The structures of these compounds were elucidated by spectroscopic analyses, including HRESIMS, 1D-, and 2D-NMR. The anti-inflammatory activities of the compounds were evaluated by suppressing the transcription of the NF-κB-dependent reporter gene in LPS-induced 293 T/NF-κB-luc cells. All compounds do not show potent activity.

Satureja hortensis L. (SH) is a medicinal and aromatic herb utilized markedly in the food, pharmaceutical, and cosmetic industries because of its specific secondary metabolites (SMs), especially monoterpenoids. However, the impact of elicitors on the expression of monoterpene synthase genes in SH remains unexplored. Therefore, the supreme objective of this research was to elucidate the phyto-biochemical and targeted genes expression responses of SH to the exogenous application of methyl jasmonate (MeJA). Accordingly, having chosen four concentrations of MeJA, they sprayed at the five-leaf stage for two weeks, and then leaves were harvested at two exposure times of 24 and 48 hrs for further analysis, using spectrophotometric, real time-quantitative polymerase chain reaction (RT-qPCR), and gas chromatography-mass spectrometric (GC-MS) methods. The results highlight that not only were the highest contents of phenolic, flavonoids, peroxidase, superoxide dismutase, proline, and malondialdehyde in the elicited SHs but also the highest expression of β-farnesene synthase (BFS) and 1-deoxy-D-xylulose 5-phosphate reductase (DXR) genes together with the highest amount of carvacrol were up-regulated in them compared to their respective control. In conclusion, these findings demonstrate the great industrial potential of MeJA in improving the production of phytochemicals in medicinal plants, particularly in SH., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Cadmium (Cd) contamination poses a substantial threat the environment, necessitating effective remediation strategies. Phytoremediation emerges as a cost-efficient and eco-friendly approach for reducing Cd levels in the soil. In this study, the suitability of A. venetum for ameliorating Cd-contaminated soils was evaluated. Mild Cd stress promoted seedling and root growth, with the root being identified as the primary tissue for Cd accumulation. The Cd content of roots ranged from 0.35 to 0.55 mg/g under treatment with 10-50 µM CdCl 2 ·2.5 H 2 O, and the bioaccumulation factor ranged from 28.78 to 84.43. Transcriptome sequencing revealed 20,292 unigenes, and 7507 nonredundant differentially expressed genes (DEGs) were identified across five comparison groups. DEGs belonging to the "MAPK signaling pathway-plant," "monoterpenoid biosynthesis," and "flavonoid biosynthesis pathway" exhibited higher expression levels in roots compared to stems and leaves. In addition, cytokinin-related DEGs, ROS scavenger genes, such as P450, glutathione-S-transferase (GST), and superoxide dismutase (SOD), and the cell wall biosynthesis-related genes, CSLG and D-GRL, were also upregulated in the root tissue, suggesting that Cd promotes root development. Conversely, certain ABC transporter genes, (e.g, NRAMP5), and some vacuolar iron transporters, predominantly expressed in the roots, displayed a strong correlation with Cd content, revealing the mechanism underlying the compartmentalized storage of Cd in the roots. KEGG enrichment analysis of DEGs showed that the pathways associated with the biosynthesis of flavonoids, lignin, and some terpenoids were significantly enriched in the roots under Cd stress, underscoring the pivotal role of these pathways in Cd detoxification. Our study suggests A. venetum as a potential Cd-contaminated phytoremediation plant and provides insights into the molecular-level mechanisms of root development promotion and accumulation mechanism in response to Cd stress., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Gibberellins (GAs) are plant hormones widely used in agriculture. At present, GAs are produced by fermentation of the fungus Fusarium fujikuroi . However, fungal growth is too slow, resulting in slow fungal fermentation and a low yield. Here, to develop an alternative production source of GAs, an artificial pathway was engineered in Escherichia coli . By selecting and combining enzymes derived from plants and bacteria, a novel 4-enzyme pathway was successfully constructed to produce GAs using steviol, a readily available and less valuable byproduct during enzymatic refining of rebaudioside A, as a feedstock. Whole-cell biotransformation with E. coli strain expressing the novel pathway produced 71.17 ± 2.00 mg/L GA 1 and a trace amount of GA 3 from steviol in 48 h. This report presents a significant advancement in the fast production of GAs and establishes a method for the metabolism of terpenoids to produce target products in microbial hosts.

Batrachotoxin (1) is a potent cardio- and neurotoxic steroid isolated from certain species of frogs, birds, and beetles. We previously disclosed two synthetic routes to 1. During our synthetic studies toward 1, we explored an alternative strategy for efficiently assembling its 6/6/6/5-membered steroidal skeleton (ABCD-ring). Here we report the application of intermolecular Weix and intramolecular pinacol coupling reactions. While Pd/Ni-promoted Weix coupling linked the AB-ring and D-ring fragments, SmI 2 -mediated pinacol coupling did not cyclize the C-ring. Instead, we discovered that SmI 2 promoted a 1,4-addition of the α-alkoxy radical intermediate to produce the unusual 11(9→7)-abeo-steroid skeleton. Thus, this study demonstrates the convergent assembly of the skeleton of the natural product matsutakone in 11 steps from 2-allyl-3-hydroxycyclopent-2-en-1-one.

Terpenes are a diverse class of natural products which have long been sought after for their chemical properties as medicine, perfumes, and for food flavoring. Computational docking studies of terpene mechanisms have been a challenge due to the lack of strong directing groups which many docking programs rely on. In this chapter, we dive into our computational method Terdockin (Terpene-Docking) as a successful methodology in modeling terpene synthase mechanisms. This method could also be used as inspiration for any multi-ligand docking project., (Copyright © 2024. Published by Elsevier Inc.)

Terpenoids are used in various industries, and Saccharomyces cerevisiae is a promising microorganism for terpenoid production. Introducing the mevalonate (MVA) pathway into the mitochondria of a strain with an augmented inherent cytosolic MVA pathway increased terpenoid production but also led to the accumulation of toxic pyrophosphate intermediates that negatively affected terpenoid production. We first engineered the inherent MVA pathway in the cytosol and then introduced the MVA pathway into the mitochondria using several promoter combinations, considering the toxicity of pyrophosphate intermediates. However, the highest titer, 183 mg/L, tends to be only 5% higher than that of the strain that only augmented the inherent MVA pathway (SYCM1; 174 mg/L). Next, we hypothesized that, in addition to the toxicity of pyrophosphate, other compounds in the MVA pathway could affect the squalene titer. Thus, we constructed a combinatorial strain library expressing MVA pathway enzymes in the mitochondria with various promoter combinations. The highest squalene titer (230 mg/L) was 32% higher than that of SYCM1. The promoter set revealed that mitigation of mono- and pyrophosphate compound accumulation was important for mitochondrial usage. This study demonstrated that a combinatorial strain library is useful for discovering the optimal gene expression balance in engineering yeast., (Copyright © 2023 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

Long non-coding RNAs (lncRNAs) have emerged as curial regulators of diverse biological processes in plants. Jasmine ( Jasminum sambac ) is a world-renowned ornamental plant for its attractive and exceptional flower fragrance. However, to date, no systematic screening of lncRNAs and their regulatory roles in the production of the floral fragrance of jasmine flowers has been reported. In this study, we identified a total of 31,079 novel lncRNAs based on an analysis of strand-specific RNA-Seq data from J. sambac flowers at different stages. The lncRNAs identified in jasmine flowers exhibited distinct characteristics compared with protein-coding genes (PCGs), including lower expression levels, shorter transcript lengths, and fewer exons. Certain jasmine lncRNAs possess detectable sequence conservation with other species. Expression analysis identified 2752 differentially expressed lncRNAs (DE&#95;lncRNAs) and 8002 DE&#95;PCGs in flowers at the full-blooming stage. DE&#95;lncRNAs could potentially cis - and trans -regulate PCGs, among which DE&#95;lincRNAs and their targets showed significant opposite expression patterns. The flowers at the full-blooming stage are specifically enriched with abundant phenylpropanoids and terpenoids potentially contributed by DE&#95;lncRNA cis -regulated PCGs. Notably, we found that many cis -regulated DE&#95;lncRNAs may be involved in terpenoid and phenylpropanoid/benzenoid biosynthesis pathways, which potentially contribute to the production of jasmine floral scents. Our study reports numerous jasmine lncRNAs and identifies floral-scent-biosynthesis-related lncRNAs, which highlights their potential functions in regulating the floral scent formation of jasmine and lays the foundations for future molecular breeding.

Little is known concerning terpenoids produced by members of the fungal order Ophiostomales, with the member Harringtonia lauricola having the unique lifestyle of being a beetle symbiont but potentially devastating tree pathogen. Nine known terpenoids, including six labdane diterpenoids ( 1 - 6 ) and three hopane triterpenes ( 7 - 9 ), were isolated from H. lauricola ethyl acetate (EtOAc) extracts for the first time. All compounds were tested for various in vitro bioactivities. Six compounds, 2 , 4 , 5 , 6 , 7 , and 9 , are described functionally. Compounds 2 , 4 , 5 , and 9 expressed potent antiproliferative activity against the MCF-7, HepG2 and A549 cancer cell lines, with half-maximal inhibitory concentrations (IC 50 s) ~12.54-26.06 μM. Antimicrobial activity bioassays revealed that compounds 4 , 5 , and 9 exhibited substantial effects against Gram-negative bacteria ( Escherichia coli and Ralstonia solanacearum ) with minimum inhibitory concentration (MIC) values between 3.13 and 12.50 μg/mL. Little activity was seen towards Gram-positive bacteria for any of the compounds, whereas compounds 2 , 4 , 7 , and 9 expressed antifungal activities ( Fusarium oxysporum ) with MIC values ranging from 6.25 to 25.00 μg/mL. Compounds 4 , 5 , and 9 also displayed free radical scavenging abilities towards 2,2-diphenyl-1-picrylhydrazyl (DPPH) and superoxide (O 2- ), with IC 50 values of compounds 2 , 4 , and 6 ~3.45-14.04 μg/mL and 22.87-53.31 μg/mL towards DPPH and O 2- , respectively. These data provide an insight into the biopharmaceutical potential of terpenoids from this group of fungal insect symbionts and plant pathogens.

Background: Cannabis sativa, a dioecious plant that has been cultivated worldwide for thousands of years, is known for its secondary metabolites, especially cannabinoids, which possess several medicinal effects. In this study, we investigated the autopolyploidization effects on the biosynthesis and accumulation of these metabolites, transcriptomic and metabolomic analyses were performed to explore the gene expression and metabolic variations in industrial hemp autotetraploids and their diploid progenitors., Results: Through these analyses, we obtained 1,663 differentially expressed metabolites and 1,103 differentially expressed genes. Integrative analysis revealed that phenylpropanoid and terpenoid biosynthesis were regulated by polyploidization. No substantial differences were found in the cannabidiol or tetrahydrocannabinol content between tetraploids and diploids. Following polyploidization, some transcription factors, including nine bHLH and eight MYB transcription factors, affected the metabolic biosynthesis as regulators. Additionally, several pivotal catalytic genes, such as flavonol synthase/flavanone 3-hydroxylase, related to the phenylpropanoid metabolic pathway, were identified as being modulated by polyploidization., Conclusions: This study enhances the overall understanding of the impact of autopolyploidization in C. sativa and the findings may encourage the application of polyploid breeding for increasing the content of important secondary metabolites in industrial hemp., (© 2023. The Author(s).)

Key Message: This review summarized how TFs function independently or in response to environmental factors to regulate terpenoid biosynthesis via fine-tuning the expression of rate-limiting enzymes. Terpenoids are derived from various species and sources. They are essential for interacting with the environment and defense mechanisms, such as antimicrobial, antifungal, antiviral, and antiparasitic properties. Almost all terpenoids have high medicinal value and economic performance. Recently, the control of enzyme genes on terpenoid biosynthesis has received a great deal of attention, but transcriptional factors regulatory network on terpenoid biosynthesis and accumulation has yet to get a thorough review. Transcription factors function as activators or suppressors independently or in response to environmental stimuli, fine-tuning terpenoid accumulation through regulating rate-limiting enzyme expression. This study investigates the advancements in transcription factors related to terpenoid biosynthesis and systematically summarizes previous works on the specific mechanisms of transcription factors that regulate terpenoid biosynthesis via hormone signal-transcription regulatory networks in plants. This will help us to better comprehend the regulatory network of terpenoid biosynthesis and build the groundwork for terpenoid development and effective utilization., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Cannabinoids are important therapeutical molecules for human ailments, cancer treatment, and SARS-CoV-2. The central cannabinoid, cannabigerolic acid (CBGA), is generated from geranyl pyrophosphate and olivetolic acid by Cannabis sativa prenyltransferase (CsPT4). Despite efforts to engineer microorganisms such as Saccharomyces cerevisiae (S. cerevisiae) for CBGA production, their titers remain suboptimal because of the low conversion of hexanoate into olivetolic acid and the limited activity and stability of the CsPT4. To address the low hexanoate conversion, we eliminated hexanoate consumption by the beta-oxidation pathway and reduced its incorporation into fatty acids. To address CsPT4 limitations, we expanded the endoplasmic reticulum and fused an auxiliary protein to CsPT4. Consequently, the engineered S. cerevisiae chassis showed a marked improvement of 78.64-fold in CBGA production, reaching a titer of 510.32 ± 10.70 mg l -1 from glucose and hexanoate., (Copyright © 2023 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.)

Enzyme spatial organization is an evolved mechanism for facilitating multi-step biocatalysis and can play an important role in the regulation of promiscuous enzymes. The latter function suggests that artificial spatial organization can be an untapped avenue for controlling the specificity of bioengineered metabolic pathways. A promiscuous terpene synthase (nerolidol synthase) is co-localized and spatially organized with the preceding enzyme (farnesyl diphosphate synthase) in a heterologous production pathway, via translational protein fusion and/or co-encapsulation in a self-assembling protein cage. Spatial organization enhances nerolidol production by ≈11- to ≈62-fold relative to unorganized enzymes. More interestingly, striking differences in the ratio of end products (nerolidol and linalool) are observed with each spatial organization approach. This demonstrates that artificial spatial organization approaches can be harnessed to modulate the product profiles of promiscuous enzymes in engineered pathways in vivo. This extends the application of spatial organization beyond situations where multiple enzymes compete for a single substrate to cases where there is competition among multiple substrates for a single enzyme., (© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Medicinal plants are rich in secondary metabolites with beneficial pharmacological effects. The production of plant secondary metabolites is subjected to the influences by environmental factors including the plant-associated microbiome, which is crucial to the host's fitness and survival. As a result, research interests are increasing in exploiting microbial capacities for enhancing plant production of pharmacological metabolites. A growing body of recent research provides accumulating evidence in support of developing microbe-based tools for achieving this objective. This mini review presents brief summaries of recent studies on medicinal plants that demonstrate microbe-augmented production of pharmacological terpenoids, polyphenols, and alkaloids, followed by discussions on some key questions beyond the promising observations. Explicit molecular insights into the underlying mechanisms will enhance microbial applications for metabolic fortification in medicinal plants., Competing Interests: Authors declare that they have no competing interests., (© 2023 The Authors.)

Having a spiral grain is considered to be one of the most important wood properties influencing wood quality. Here, transcriptome profiles and metabolome data were analyzed in the straight grain and twist grain of Pinus yunnanensis . A total of 6644 differential expression genes were found between the straight type and the twist type. A total of 126 differentially accumulated metabolites were detected. There were 24 common differential pathways identified from the transcriptome and metabolome, and these pathways were mainly annotated in ABC transporters, arginine and proline metabolism, flavonoid biosynthesis, isoquinoline alkaloid biosynthesis, linoleic acid metabolism, phenylpropanoid, tryptophan metabolism, etc. A weighted gene coexpression network analysis showed that the lightblue4 module was significantly correlated with 2'-deoxyuridine and that transcription factors (basic leucine zipper (bZIP), homeodomain leucine zipper (HD-ZIP), basic helix-loop-helix (bHLH), p-coumarate 3-hydroxylase (C3H), and N-acetylcysteine (NAC)) play important roles in regulating 2'-deoxyuridine, which may be involved in the formation of spiral grains. Meanwhile, the signal transduction of hormones may be related to spiral grain, as previously reported. ARF7 and MKK4&#95;5 , as indoleacetic acid (IAA)- and ethylene (ET)-related receptors, may explain the contribution of plant hormones in spiral grain. This study provided useful information on spiral grain in P. yunnanensis by transcriptome and metabolome analyses and could lay the foundation for future molecular breeding.

Background: Cannabis and its primary psychoactive constituent delta-9-tetrahydrocannabinol (D9-THC) produce biphasic, dose-dependent effects on anxiety. In addition to D9-THC, cannabis contains other "minor" cannabinoids and terpenes with purported therapeutic potential for the treatment of anxiety. Empirical data on potential therapeutic effects of these compounds is limited. The current study evaluated the effects of selected minor cannabinoids and terpenes in a battery of tests sensitive to anxiolytic and anxiogenic drugs. Methods: In Experiment 1, adult male Sprague Dawley rats ( N =7-8/group) were administered acute oral doses of one of five minor cannabinoids: delta-8-tetrahydrocannabinol (D8-THC; 10 mg/kg), tetrahydrocannabivarin (32 mg/kg), cannabidiolic acid (32 mg/kg), cannabidivarin (32 mg/kg), and cannabigerol (100 mg/kg), or one of five terpenes: D-limonene (17 mg/kg), ⍺-pinene (100 mg/kg), ⍺-terpineol (10 mg/kg), bisabolol (100 mg/kg), and β-caryophyllene (17 mg/kg), or vehicle (medium-chain triglycerides [MCT] oil). Ethyl alcohol was tested as an active comparator. Thirty minutes post-administration, the marble burying test, the three-chamber social interaction test, and the novelty-induced hypophagia test were completed; motor activity was assessed throughout testing. Experiment 2 examined the potential anxiolytic effects of minor cannabinoids when administered chronically; rats administered MCT oil or minor cannabinoids in Experiment 1 continued receiving once-daily doses for 21 days and were assessed using the same test battery after 7, 14, and 21 days of administration. Results and Conclusions: When compared to vehicle, acute administration of bisabolol and D-limonene increased the amount of food consumed and bisabolol-, D-limonene-, ⍺-pinene-, and β-caryophyllene decreased percent time spent in the outer zone in the novelty-induced hypophagia test, suggestive of an anxiolytic effect. Only ethanol increased social interaction. After acute administration, anxiogenic effects in the marble burying test were observed for D8-THC, but not for other minor cannabinoids and terpenes. Throughout chronic administration, only D8-THC displayed anxiogenic effects in the novelty-induced hypophagia test. The other cannabinoids did not show anxiolytic or anxiogenic effects in any of the tests at the doses or times tested. The minor cannabinoids and terpenes did not impair or stimulate general motor activity. These data provide a foundation for future studies investigating cannabinoid/terpene interactions.

Five new lipids, tricholixins A-E ( 1 - 5 ), and two known terpenoids, brasilane A ( 6 ) and harzianone A ( 7 ), were discovered from a deep-sea strain (R22) of the fungus Trichoderma lixii isolated from the cold seep sediments of the South China Sea. Their structures and relative configurations were identified by meticulous analysis of MS and IR as well as NMR data. The absolute configuration of 5 was ascertained by dimolybdenum-induced ECD data in particular. Compounds 1 and 2 represent the only two new butenolides from marine-derived Trichoderma , and they further add to the structural diversity of these molecules. Although 6 has been reported from a basidiomycete previously, it is the first brasilane aminoglycoside of Trichoderma origin. During the assay against wheat-pathogenic fungi, both 1 and 2 inhibited Fusarium graminearum with an MIC value of 25.0 μg/mL, and 6 suppressed Gaeumannomyces graminis with an MIC value of 12.5 μg/mL. Moreover, the three isolates also showed low toxicity to the brine shrimp Artemia salina .

Plants from the genus Styrax have been extensively used in folk medicines to treat diseases such as skin diseases and peptic ulcers and as an antiseptic and analgesic. Most Styrax species, especially Styrax tonkinensis , which is used as an expectorant, antiseptic, and analgesic in Chinese traditional medicine, could screen resin after external injury. Styrax is also used in folk medicines in Korea to treat sore throat, bronchitis, cough, expectoration, paralysis, laryngitis, and inflammation. Different parts of various Styrax species can be widely employed for ethnopharmacological applications. Moreover, for ethnopharmacological use, these parts of Styrax species can be applied in combination with other folk medicines. Styrax species consist of versatile natural compounds, with some of them exhibiting particularly excellent pharmacological activities, such as cytotoxic, acetylcholinesterase inhibitory, antioxidant, and antifungal activities. Altogether, these exciting results indicate that a comprehensive review of plants belonging to this genus is essential for helping researchers to continuously conduct an in-depth investigation. In this review, the traditional uses, phytochemistry, corresponding pharmacological activities, and structure-activity relationships of different Styrax species are clarified and critically discussed. More insights into potential opportunities for future research are carefully assessed., Competing Interests: The authors declare no conflicts of interest.

Terpenoids are a large class of plant-derived compounds, that constitute the main components of essential oils and are widely used as natural flavors and fragrances. The biosynthesis approach presents a promising alternative route in terpenoid production compared to plant extraction or chemical synthesis. In the past decade, the production of terpenoids using biotechnology has attracted broad attention from both academia and the industry. With the growing market of flavor and fragrance, the production of terpenoids directed by synthetic biology shows great potential in promoting future market prospects. Here, we reviewed the latest advances in terpenoid biosynthesis. The engineering strategies for biosynthetic terpenoids were systematically summarized from the enzyme, metabolic, and cellular dimensions. Additionally, we analyzed the key challenges from laboratory production to scalable production, such as key enzyme improvement, terpenoid toxicity, and volatility loss. To provide comprehensive technical guidance, we collected milestone examples of biosynthetic mono- and sesquiterpenoids, compared the current application status of chemical synthesis and biosynthesis in terpenoid production, and discussed the cost drivers based on the data of techno-economic assessment. It is expected to provide critical insights into developing translational research of terpenoid biomanufacturing., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Five undescribed terpenoids including a polyketide-terpenoid hybrid paraphaone, and four eremophilane sesquiterpenoids, paraphaterpenes A-D, as well as two known compounds were isolated from the endophytic fungus Paraphaeosphaeria sp. cultured by extract of host Ginkgo biloba L. The structures were established by spectroscopic analyses, and the single-crystal X-ray diffraction. The antifungal activity of Paraphaeosphaeria sp. cultured by extract of G. biloba against the plant pathogen Alternaria alternata was significant and higher than that of PDB medium. Tested compounds indicated antifeedant activities against silkworms with feeding deterrence index at 10-70%. Paraphaone, paraphaterpenes A, C, D and alternariol methyl ether showed significant antifungal activities against the phytopathogens A. alternata, Aspergillus fumigatus, and entomopathogen Beauveria bassiana with MICs ≤4 μg/mL. And the preliminary structure-activity relationship of eremophilane sesquiterpenoids was exhibited. The culture of Paraphaeosphaeria sp. by host G. biloba medium afforded agricultural antibiotics., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Due to the complexity of metabolic and regulatory networks in microorganisms, it is difficult to obtain robust phenotypes through artificial rational design and genetic perturbation. Adaptive laboratory evolution (ALE) engineering plays an important role in the construction of stable microbial cell factories by simulating the natural evolution process and rapidly obtaining strains with stable traits through screening. This review summarizes the application of ALE technology in microbial breeding, describes the commonly used methods for ALE, and highlights the important applications of ALE technology in the production of lipids and terpenoids in yeast and microalgae. Overall, ALE technology provides a powerful tool for the construction of microbial cell factories, and it has been widely used in improving the level of target product synthesis, expanding the range of substrate utilization, and enhancing the tolerance of chassis cells. In addition, in order to improve the production of target compounds, ALE also employs environmental or nutritional stress strategies corresponding to the characteristics of different terpenoids, lipids, and strains.

The roots of F. haussknechtii are used by local people in order to treat wounds and urinary infections. Ferula species are rich in bioactive compounds with biological effects. In line with our previous studies about screening antibacterial natural products, five terpenoid derivatives were purified from Ferula haussknechtii . The separation and purification were performed by column chromatography. Their structures were determined by 1 D and 2 D NMR as hawraman 8- p -hydroxybenzoyl-tovarol ( 1 ), ferutinin ( 2 ), lancerotriol 6-( p -hydroxybenzoate) ( 3 ), chimganin ( 4 ), and chimgin ( 5 ). Then, the antibacterial effects of the purified compounds were evaluated by measuring their MIC values against Gram-positive and Gram-negative bacteria. The results showed that compound ( 1 ) had the most antibacterial effect on Bacillus cereus (MIC = 16 µg/mL). The antibacterial effects of F. haussknechtii compounds are in line with their local application and it is suggested that further studies should be conducted to determine their mechanism of action.

Modern tools in DNA synthesis and assembly give genetic engineers control over the nucleotide-level design of complex, multi-gene systems. Systematic approaches to explore genetic design space and optimize the performance of genetic constructs are lacking. Here we explore the application of a five-level Plackett-Burman fractional factorial design to improve the titer of a heterologous terpene biosynthetic pathway in Streptomyces. A library of 125 engineered gene clusters encoding the production of diterpenoid ent-atiserenoic acid (eAA) via the methylerythritol phosphate pathway was constructed and introduced into Streptomyces albidoflavus J1047 for heterologous expression. The eAA production titer varied within the library by over two orders of magnitude and host strains showed unexpected and reproducible colony morphology phenotypes. Analysis of Plackett-Burman design identified expression of dxs, the gene encoding the first and the flux-controlling enzyme, having the strongest impact on eAA titer, but with a counter-intuitive negative correlation between dxs expression and eAA production. Finally, simulation modeling was performed to determine how several plausible sources of experimental error/noise and non-linearity impact the utility of Plackett-Burman analyses., (Copyright © 2023 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.)

Terpenoid is an important group of compounds not only as biocomponents but also as useful secondary metabolites. A volatile terpenoid 1,8-cineole, which is used as a food additive, flavoring agent, cosmetic, etc., is also attracting attention from a medical perspective due to its antiinflammation and antioxidation. The 1,8-cineole fermentation using a recombinant Escherichia coli strain has been reported, although a carbon source supplement is necessary for a high-yield 1,8-cineole production. We constructed the 1,8-cineole-producing cyanobacteria toward a carbon-free and sustainable 1,8-cineole production. cnsA, the 1,8-cineole synthase gene in Streptomyces clavuligerus ATCC 27064, was introduced and overexpressed in the cyanobacterium Synechococcus elongatus PCC 7942. We succeeded in producing an average of 105.6 µg g-1 wet cell weight of 1,8-cineole in S. elongatus 7942 without supplementing any carbon source. Using the cyanobacteria expression system is an efficient approach to producing 1,8-cineole by photosynthesis., (© The Author(s) 2023. Published by Oxford University Press on behalf of Japan Society for Bioscience, Biotechnology, and Agrochemistry.)