Search Results (14)

Eperua oleifera Ducke (Fabaceae), commonly known as copaíba-jacaré, is traditionally used for therapeutic purposes, like Copaifera oleoresins. Previous GC-MS studies reported its chemical composition as mainly composed of diterpenic acids, consistent with species of the same genus. Although GC-MS remains widely used for comparing compound retention times and fragmentation patterns, its application to diterpenic acids requires a derivatization step to form methyl esters due to the poor chromatographic performance of carboxylic acids on methyl silicone stationary phases. This step may lead to misinterpretations, especially considering recent findings of naturally occurring methyl esters in oleoresins that may co-elute with derivatized acids. This study aimed to apply more sensitive analytical techniques to identify both target and untargeted compounds. The resin of E. oleifera was analyzed by GC-MS to assess the presence of volatile components. Additionally, UHPLC-HRMS was employed using full-scan MS, data-dependent acquisition (DDA), and parallel reaction monitoring (PRM) in both positive and negative ESI modes. GC-MS confirmed the absence of volatile sesquiterpenes, classifying E. oleifera as a resin. Targeted UHPLC-HRMS detected natural methyl esters of diterpenic acids, while untargeted analysis using Compound Discoverer 3.3 software revealed flavonoids and phenolic compounds not previously reported. These findings support the application of UHPLC-HRMS as a powerful tool in phytochemical studies. Full article

S. mutans is a key pathogen in dental caries initiation and progression. It promotes oral biofilm dysbiosis and biofilm acidification. Sodium resinate is a salt of pine-oil-derived resin which has antimicrobial properties. Pine-oil-derived resin consists of terpenes, diterpenes, and abietic acids. The aim of this study was to determine the effects of pine (Pinus sylvestris) oil resinate (RS) on growth and acid production of cariogenic S. mutans strains in planktonic form and biofilm. The S. mutans type strain NCTC10449 and clinical isolate CI2366 were grown on 96-well plates for testing of RS effects on growth and biofilm formation, and on plates with integrated pH-sensitive optical ensors for real-time measurements of the effects of RS on bacterial acid production. We found that even short-time exposure to RS inhibits the growth and acid production of S. mutans in the planktonic phase and biofilms. In addition, RS was able to penetrate the biofilm matrix and reduce acid production inside S. mutans biofilm. RS thus shows potential as a novel antibacterial agent against cariogenic bacteria in biofilm. Full article

Balms and resins of Picea abies, Larix decidua, and Pinus nigra are traditionally used to treat wounds. Three chromatographic techniques differing in separation capacity and technical demands were employed to distinguish among these plant exudates. A TLC method was established for fingerprint comparison, providing a quick overview of a large number of samples at low cost. HPLC-DAD (RP18) and UHPSFC-DAD (Torus 2-Picolylamin), hyphenated to ESI-MS, represented orthogonal chromatographic systems with high separation performance. The developed methods allow for the separation and detection of major and minor constituents belonging to different compound classes (phenyl carboxylic acids, lignans, diterpene resin acids). The qualitative compositions of the diterpene resin acids, the main compounds in the exudates, were comparable in all three genera. Differences were detected in the distribution of hydroxylated diterpene resin acids, pinoresinol, and hydroxycinnamic acids. The three tested chromatographic systems with varying demands on lab equipment offer appropriate tools for the quality assessment of Picea abies, Larix decidua, and Pinus nigra. The extracts were furthermore tested at three different concentrations (10 µg/mL, 3 µg/mL, and 1 µg/mL) for boosted re-epithelialization, a crucial step in the wound-healing process, in an in vitro HaCaT keratinocyte-based scratch assay. Lysophosphatidic acid (LPA, 10 µM) and extracts of several medicinal plants well known for their wound-healing properties (birch, marigold, St. John’s wort, manuka honey) were used as positive controls. Picea abies and Pinus nigra showed concentration dependency; significant activity was measured for Larix decidua at 3 µg/mL. Full article

Diterpene resin acids (DRAs) are major components of pine oleoresin that can effectively resist the invasion of insects and pathogenic microorganisms. The subfamily of cytochrome P450s, CYP720B, catalyzes diterpene products into DRAs. Identifying CYP720B subfamily members and revealing the characteristics of tissue-specific expression would help understand diterpene-rich structures and diverse types. Slash pine and loblolly pine are important pines that provide oleoresin products. In this study, we identified CYP720B candidate genes based on the Pinus taeda V2.0 genome and full-length transcriptome of slash pine by PacBio. A total of 17 genes in slash pine and 19 in loblolly pine were identified and classified into four main clades by phylogenetic analysis. An analysis of cis-acting elements showed that CYP720B genes were closely related to adversity resistance. The gene expression of these candidates in different tissues was quantified by real-time quantitative PCR (RT–qPCR) analysis. Most of the genes showed relatively higher expression levels in roots and stems than in the other tissues, corresponding with the results of DRA component detection by gas chromatography–mass spectrometry (GC–MS), which indicated that stems and roots might be important tissues in oleoresin biosynthesis. These results provide a valuable resource for a better understanding of the biological role of individual CYP720Bs in slash pine and loblolly pine. Full article

A quali-quantitative analysis of diterpenoid composition in tissues obtained from different organs of Pinus nigra subsp. laricio (Poiret) Maire (Calabrian pine) was carried out. Diterpene resin acids were the most abundant diterpenoids across all the examined tissues. The same nine diterpene resin acids were always found, with the abietane type prevailing on the pimarane type, although their quantitative distribution was found to be remarkably tissue-specific. The scrutiny of the available literature revealed species specificity as well. A phylogeny-based approach allowed us to isolate four cDNAs coding for diterpene synthases in Calabrian pine, each of which belonging to one of the four groups into which the d3 clade of the plants’ terpene synthases family can be divided. The deduced amino acid sequences allowed predicting that both monofunctional and bifunctional diterpene synthases are involved in the biosynthesis of diterpene resin acids in Calabrian pine. Transcript profiling revealed differential expression across the different tissues and was found to be consistent with the corresponding diterpenoid profiles. The isolation of the complete genomic sequences and the determination of their exon/intron structures allowed us to place the diterpene synthase genes from Calabrian pine on the background of current ideas on the functional evolution of diterpene synthases in Gymnosperms. Full article

With the aim of addressing the impact of extractive protocols in molecular characterization of ceramic content, sixteen archaeological shards and waterproofing coatings of Roman amphorae were studied to compare the extractive capacities of protocols prevalently mentioned in wine amphorae analysis. A microwave-assisted protocol is developed in order to esterify grape-derivative markers from archaeological pitch and shard. Gas chromatography-mass spectrometry is used to highlight the great capacities of a two-step protocol that combines organic extraction with BF₃-etherate complex butylation applied on archaeological shards. Instead, simultaneous alkaline fusion and direct-resin acid-catalyzed butylation are favored for the characterization of waterproofing material. The identification of tartaric acid, together with succinic, fumaric pyruvic and syringic acids provide valuable insights on the archaeological grape-derivative content, possibly wine. Diterpenic markers highlighted Pinus pitch and wood tar, originally used to waterproof the amphorae. Since markers are reliable tools in organic residue analyses, protocols exhibiting high extractive capacities are favored to avoid false conclusions drawn through the absence of markers. Full article

Fourteen triterpene acids, viz., three tirucallane-type (1–3), eight ursane-type (4–11), two oleanane-type (12, 13) and one lupane type (21), along with boswellic aldehyde (14), α-amyrine (15), epi-amyrine (16), straight chain acid (17), sesquiterpene (19) and two cembrane-type diterpenes (18, 20) were isolated, first time, from the methanol extract of Boswellia elongata resin. Compound (1) was isolated for first time as a natural product, while the remaining compounds (2‒21) were reported for first time from B. elongata. The structures of all compounds were confirmed by advanced spectroscopic techniques including mass spectrometry and also by comparison with the reported literature. Eight compounds (1–5, 11, 19 and 20) were further screened for in vitro α-glucosidase inhibitory activity. Compounds 3–5 and 11 showed significant activity against α-glucosidase with IC₅₀ values ranging from 9.9–56.8 μM. Compound 4 (IC₅₀ = 9.9 ± 0.48 μM) demonstrated higher inhibition followed by 11 (IC₅₀ = 14.9 ± 1.31 μM), 5 (IC₅₀ = 20.9 ± 0.05 μM) and 3 (IC₅₀ = 56.8 ± 1.30 μM), indicating that carboxylic acid play a key role in α-glucosidase inhibition. Kinetics studies on the active compounds 3–5 and 11 were carried out to investigate their mechanism (mode of inhibition and dissociation constants K_i). All compounds were found to be non-competitive inhibitors with K_i values in the range of 7.05 ± 0.17–51.15 ± 0.25 µM. Moreover, in silico docking was performed to search the allosteric hotspot for ligand binding which is targeted by our active compounds investigates the binding mode of active compounds and it was identified that compounds preferentially bind in the allosteric binding sites of α-glucosidase. The results obtained from docking study suggested that the carboxylic group is responsible for their biologic activities. Furthermore, the α-glucosidase inhibitory potential of the active compounds is reported here for the first time. Full article

A high-throughput quantitative Nuclear Magnetic Resonance ¹H-NMR method was developed and applied to screen the quantity of the diterpenic resin acids in the heartwood of black pine, due to the renewed scientific interest in their medicinal properties and use in various diseases treatment. The 260 samples were taken from Pinus nigra clones, selected from four provenances of the Peloponnese (Greece), participating in a 35-year-old clonal seed orchard. Total resin acids per dry heartwood weight (dhw) varied greatly, ranging from 30.05 to 424.70 mg/g_dhw (average 219.98 mg/g_dhw). Abietic was the predominant acid (76.77 mg/g_dhw), followed by palustric acid (47.94 mg/g_dhw), neoabietic acid (39.34 mg/g_dhw), and pimaric acid (22.54 mg/g_dhw). Dehydroabietic acid was at moderate levels (11.69 mg/g_dhw), while levopimaric, isopimaric, and sandaracopimaric acids were in lower concentrations. The resin acid fraction accounted for 72.33% of the total acetone extractives. Stilbenes were presented in significant quantities (19.70%). The resin acid content was composed mainly of the abietane type resin acids (83.56%). Peloponnesian Pinus nigra heartwood was found to be the richest source of resin acids identified to date and is considered the best natural source for the production of such bioactive extracts. The results indicate a high potential for effective selection and advanced breeding of pharmaceutical and high economic value bioactive substances from Pinus nigra clones. Full article

Dehydroabietic acid (DAA) is a naturally occurring diterpene resin acid derived from coniferous plants such as Pinus and Picea. Various bioactive effects of DAA have been studied including antibacterial, antifungal, and anticancer activities. However, the anti-inflammatory mechanism of DAA remains unclear. We evaluated the anti-inflammatory effect of DAA in macrophage cell lines. Dehydroabietic acid clearly reduced nitric oxide (NO) production and inflammatory gene expression decreased according to RT-PCR results. Dehydroabietic acid displayed anti-inflammatory activity at the transcriptional level in results from NF-κB- or AP-1-mediated luciferase assays. To identify the DAA target protein, we investigated NF-κB and AP-1 pathways by Western blotting analysis. Dehydroabietic acid suppressed the activity of proto-oncogene tyrosine protein kinase (Src) and spleen tyrosine kinase (Syk) in the NF-κB cascade and transforming growth factor beta-activated kinase 1 (TAK1) in the AP-1 cascade. Using overexpression strategies, we confirmed that DAA targeted these kinases. Our findings demonstrate the anti-inflammatory effects and molecular mechanism of DAA. This suggests that DAA has potential as a drug or supplement to ameliorate inflammation. Full article

The chemical composition of lipophilic bark extracts from Pinus pinaster and Pinus pinea cultivated in Portugal was evaluated using gas chromatography-mass spectrometry. Diterpenic resin acids were found to be the main components of these lipophilic extracts, ranging from 0.96 g kg⁻¹ dw in P. pinea bark to 2.35 g kg⁻¹ dw in P. pinaster bark. In particular, dehydroabietic acid (DHAA) is the major constituent of both P. pinea and P. pinaster lipophilic fractions, accounting for 0.45 g kg⁻¹ dw and 0.95 g kg⁻¹ dw, respectively. Interestingly, many oxidized compounds were identified in the studied lipophilic extracts, including DHAA-oxidized derivatives (7-oxo-DHAA, 7α/β-hydroxy-DHAA, and 15-hydroxy-DHAA, among others) and also terpin (an oxidized monoterpene). These compounds are not naturally occurring compounds, and their formation might occur by the exposure of the bark to light and oxygen from the air, and the action of micro-organisms. Some of these compounds have not been previously reported as lipophilic constituents of the bark of the referred pine species. Other constituents, such as aromatic compounds, fatty acids, fatty alcohols, and sterols, are also present in the studied extracts. These results can represent an opportunity to valorize P. pinaster and P. pinea by-products as a primary source of the bioactive resin acids that are integrated into the current uses of these species. Full article

The oleoresin of Copaifera trees has been widely used as a traditional medicine in Neotropical regions for thousands of years and remains a popular treatment for a variety of ailments. The copaiba resins are generally composed of a volatile oil made up largely of sesquiterpene hydrocarbons, such as β-caryophyllene, α-copaene, β-elemene, α-humulene, and germacrene D. In addition, the oleoresin is also made up of several biologically active diterpene acids, including copalic acid, kaurenoic acid, alepterolic acid, and polyalthic acid. This review presents a summary of the ecology and distribution of Copaifera species, the traditional uses, the biological activities, and the phytochemistry of copaiba oleoresins. In addition, several biomolecular targets relevant to the bioactivities have been implicated by molecular docking methods. Full article

Little is known about the changes in resin composition in South American gymnosperms associated with the different seasons of the year. The diterpene composition of 44 resin samples from seven Austrocedrus chilensis (Cupressaceae) trees, including male and female individuals, was investigated in three different seasons of the year (February, June and November). Twelve main diterpenes were isolated by chromatographic means and identified by gas chromatography-mass spectrometry and nuclear magnetic resonance (NMR). The diterpene composition was submitted to multivariate analysis to find possible associations between chemical composition and season of the year. The principal component analysis showed a clear relation between diterpene composition and season. The most characteristic compounds in resins collected in summer were Z-communic acid (9) and 12-oxo-labda-8(17),13E-dien-19 oic acid methyl ester (10) for male trees and 8(17),12,14-labdatriene (7) for female trees. For the winter samples, a clear correlation of female trees with torulosic acid (6) was observed. In spring, E-communic acid (8) and Z-communic acid (9) were correlated with female trees and 18-hydroxy isopimar-15-ene (1) with male tree resin. A comparison between percent diterpene composition and collection time showed p < 0.05 for isopimara-8(9),15-diene (2), sandaracopimaric acid (4), compound (7) and ferruginol (11). Full article

Seventeen diterpenes belonging to the labdane, abietane and isopimarane skeleton classes were isolated from the resin of the Chilean gymnosperm Austrocedrus chilensis and identified by spectroscopic and spectrometric methods. The diterpene 12-oxo-labda-8(17),13E-dien-19 oic acid is reported for the first time as a natural product and 14 diterpenes are reported for the first time for the species. Full article

An ancient organic residue was collected from the bottom of a Greco-Italian amphora found in the Adriatic Sea and investigated by direct GC and GC-MS analysis. The headspace composition was determined by HS-SPME using: (1) DVB/CAR/PDMS and (2) PDMS/DVB fibres. Higher percentages of benzene derivatives, monoterpenes and other low-molecular aliphatic compounds were obtained by method (1) in contrast to higher percentage of naphthalene and phenanthrene derivatives found by method (2). In comparison with the composition of pine resin, it is more likely that the found low-molecular aliphatic alcohols, acids, esters and carbonyls with 2-phenylethanol were trapped and preserved within the organic residue from stored wine – the amphora’s originally content. Semi-volatile diterpenes methyl dehydroabietate (33.6%) and retene (24.1%) were dominant in the residue CH₂Cl₂ solution. Other abundant compounds were 1,4-dimethoxyphenanthrene (6.8%) as well as other naphthalene and/or phenanthrene derivatives [7-(1-methylethyl)-1,4a-dimethyl-1,2,3,4,4a,9,10,10a-octahydronaphthalene, 7-(1-methylethyl)-1,4a-dimethyl-2,3,4,4a,9,10-hexahydrophenanthrene, 7-(1-methylethyl)-1,4a-dimethyl-1,2,3,4,4a,9,10,10a-octahydro-phenanthrene, 3,6-dimethylphenanthrene and 2,3,5-trimethylphenanthrene]. Possible sources and formation pathways of the major compounds in the residue were discussed. Full article