Coffee Diterpenes Research Articles

Effects of 12-Week Supplementation with Coffee Diterpene Cafestol in Healthy Subjects with Increased Waist Circumference: A Randomized, Placebo-Controlled Trial.

Background: Coffee consumption is inversely associated with type 2 diabetes. Cafestol, a bioactive compound in coffee, has demonstrated glucose-lowering and insulin-secretory properties in cell and animal studies. The acute effects of cafestol on glucose metabolism in humans have only been briefly investigated, and longer-term effects have not been explored. This study aimed to assess the effects of purified cafestol on insulin sensitivity and other metabolic parameters in healthy individuals with increased waist circumference at risk of developing type 2 diabetes. Methods: A 12-week randomized, placebo-controlled, parallel trial was conducted with 40 participants. Insulin suppression tests, mixed meal tests, and MRI scans were performed before and after the intervention. Results: Administering 6 mg of cafestol twice daily did not alter insulin sensitivity or glucose tolerance but led to significant reductions in body weight (2%), visceral fat volume (5%), and gamma-glutamyl transferase levels (15%) compared to the placebo. Conclusions: Cafestol may hold promise for weight and visceral fat reduction. Cafestol did not improve insulin sensitivity or glucose tolerance in this study but might still contribute to the observed inverse association between coffee consumption and type 2 diabetes. Future research should explore higher dosages and longer treatment durations, particularly in individuals with impaired glucose metabolism and type 2 diabetes.

Two Coffee Diterpenes, Kahweol and Cafestol, Inhibit Extracellular Melanogenesis: An In Vitro Pilot Study

Hyperpigmentation skin disorders are marked by an abnormal accumulation or export of melanin pigment synthesized within melanocytes and pose a significant aesthetic concern. The search for novel natural compounds that exhibit pharmacological potential for treating pigmentation disorders is growing. In this study, kahweol (KW) and cafestol (CFS), two structural analogs of coffee diterpenes, were evaluated and compared for their effects on melanogenesis using B16F10 mouse melanoma cells and primary human melanocytes derived from Asian and African American skin. To the best of our knowledge, there are no reports of the effects of KW and CFS on melanogenesis yet. We first screened nontoxic concentrations of both compounds using an MTS assay after 72 h incubations and subsequently tested their effects on melanin synthesis and export. Cellular tyrosinase activity and cell-free mushroom tyrosinase activity were assayed to study the mechanisms of melanogenesis suppression. Human melanocytes from a moderately pigmented donor (HEMn-MP cells) and from a darkly pigmented donor (HEMn-DP cells) were next examined, and effects on cellular viability, melanin content, cellular tyrosinase activity, and melanin export (quantitated via dendricity) were similarly examined for both compounds. Our results show that KW and CFS did not significantly affect intracellular melanin content but suppressed extracellular melanin in B16F10 cells and dendritic parameters in human melanocytes, indicating their unique capacity to target extracellular melanogenesis and melanin export. Although KW showed a greater extracellular melanogenesis inhibitory capacity in B16F10 cells, in both primary melanocyte cells, CFS emerged as a potent inhibitor of melanin export compared to KW. Together, these results reveal novel modes of action of both compounds and indicate a promise to use CFS as a novel candidate for treating hyperpigmentation disorders of the human skin for clinical and cosmetic use. Additional research is necessary to shed light on the molecular pathways and the efficacy of melanogenesis inhibition by CFS in 3D human skin equivalents and in vivo studies.

On the Cholesterol Raising Effect of Coffee Diterpenes Cafestol and 16-O-Methylcafestol: Interaction with Farnesoid X Receptor.

The diterpene cafestol represents the most potent cholesterol-elevating compound known in the human diet, being responsible for more than 80% of the effect of coffee on serum lipids, with a mechanism still not fully clarified. In the present study, the interaction of cafestol and 16-O-methylcafestol with the stabilized ligand-binding domain (LBD) of the Farnesoid X Receptor was evaluated by fluorescence and circular dichroism. Fluorescence quenching was observed with both cafestol and 16-O-methylcafestol due to an interaction occurring in the close environment of the tryptophan W454 residue of the protein, as confirmed by docking and molecular dynamics. A conformational change of the protein was also observed by circular dichroism, particularly for cafestol. These results provide evidence at the molecular level of the interactions of FXR with the coffee diterpenes, confirming that cafestol can act as an agonist of FXR, causing an enhancement of the cholesterol level in blood serum.

Diterpenes of Coffea seeds show antifungal and anti-insect activities and are transferred from the endosperm to the seedling after germination

Species of the genus Coffea accumulate diterpenes of the ent-kaurane family in the endosperm of their seeds, of which cafestol and kahweol are the most abundant. The diterpenes are mainly stored in esterified form with fatty acids, mostly palmitate. In contrast to the numerous studies on their effects on human health and therapeutic applications, nothing was previously known about their biological and ecological role in planta. The antifungal and anti-insect activities of cafestol and cafestol palmitate were thus investigated in this study. Cafestol significantly affected the mycelial growth of five of the six phytopathogenic fungi tested. It also greatly reduced the percentage of pupation of larvae and the pupae and adult masses of one of the two fruit flies tested. By contrast, cafestol palmitate had no significant effect against any of the fungi and insects studied. Using confocal imaging and oil body isolation and analysis, we showed that diterpenes are localized in endosperm oil bodies, suggesting that esterification with fatty acids enables the accumulation of large amounts of diterpenes in a non-toxic form. Diterpene measurements in all organs of seedlings recovered from whole seed germination or embryos isolated from the endosperm showed that diterpenes are transferred from the endosperm to the cotyledons during seedling growth and then distributed to all organs, including the hypocotyl and the root. Collectively, our findings show that coffee diterpenes are broad-spectrum defence compounds that protect not only the seed on the mother plant and in the soil, but also the seedling after germination.

The Coffee Diterpene, Kahweol, Ameliorates Pancreatic β-Cell Function in Streptozotocin (STZ)-Treated Rat INS-1 Cells through NF-kB and p-AKT/Bcl-2 Pathways.

Kahweol is a diterpene molecule found in coffee that exhibits a wide range of biological activity, including anti-inflammatory and anticancer properties. However, the impact of kahweol on pancreatic β-cells is not known. Herein, by using clonal rat INS-1 (832/13) cells, we performed several functional experiments including; cell viability, apoptosis analysis, insulin secretion and glucose uptake measurements, reactive oxygen species (ROS) production, as well as western blotting analysis to investigate the potential role of kahweol pre-treatment on damage induced by streptozotocin (STZ) treatment. INS-1 cells pre-incubated with different concentrations of kahweol (2.5 and 5 µM) for 24 h, then exposed to STZ (3 mmol/L) for 3 h reversed the STZ-induced effect on cell viability, apoptosis, insulin content, and secretion in addition to glucose uptake and ROS production. Furthermore, Western blot analysis showed that kahweol downregulated STZ-induced nuclear factor kappa B (NF-κB), and the antioxidant proteins, Heme Oxygenase-1 (HMOX-1), and Inhibitor of DNA binding and cell differentiation (Id) proteins (ID1, ID3) while upregulated protein expression of insulin (INS), p-AKT and B-cell lymphoma 2 (BCL-2). In conclusion, our study suggested that kahweol has anti-diabetic properties on pancreatic β-cells by suppressing STZ induced apoptosis, increasing insulin secretion and glucose uptake. Targeting NF-κB, p-AKT, and BCL-2 in addition to antioxidant proteins ID1, ID3, and HMOX-1 are possible implicated mechanisms.

Anti-proliferative and anti-migratory properties of coffee diterpenes kahweol acetate and cafestol in human renal cancer cells

Despite improvements in systemic therapy options for renal cancer, it remains one of the most drug-resistant malignancies. Interestingly, reports have shown that kahweol and cafestol, natural diterpenes extracted from coffee beans, exhibit anti-cancer activity. However, the multiple potential pharmacological actions of both have yet to be fully understood. This study therefore investigated the effects of kahweol acetate and cafestol on human renal cancer ACHN and Caki-1 cells. Accordingly, the combination of kahweol acetate and cafestol administration synergistically inhibited cell proliferation and migration by inducing apoptosis and inhibiting epithelial–mesenchymal transition. Mechanistic dissection revealed that kahweol acetate and cafestol inhibited Akt and ERK phosphorylation. Moreover, kahweol acetate and cafestol downregulated the expression of not only C–C chemokine receptors 2, 5, and 6 but also programmed death-ligand 1, indicating their effects on the tumor microenvironment. Thus, kahweol acetate and cafestol may be novel therapeutic candidates for renal cancer considering that they exert multiple pharmacological effects.

16-O-Methylated diterpenes in green Coffea arabica: ultra-high-performance liquid chromatography-tandem mass spectrometry method optimization and validation.

Coffee diterpenes are the main constituents of the coffee oil unsaponifiable fraction. The three most important diterpenes are cafestol, kahweol, and 16-O-methylcafestol (16-OMC), and they are produced, except for cafestol, only by plants of the Coffea genus. Recently, in addition to these three major diterpenes, another 16-O-methylated diterpene (16-O-methylkahweol: 16-OMK) has been identified and quantified, for the first time, in Robusta coffee. For many years, 16-OMC has been considered present exclusively in Robusta, and so it has been reputed an excellent authenticity marker for the presence of Robusta in coffee products. For its quantification, nuclear magnetic resonance (NMR) has proved very useful when compared with other methods. Quite recently, the detection of very low levels of the two 16-O-methylated diterpenes (16-OMD) 16-OMC and 16-OMK in roasted Arabica was reported. This finding makes the use of NMR methods in 16-OMD quantification in Arabica coffee particularly challenging in view of both the trace amounts of 16-OMD and the impossibility to discriminate between 16-OMC and 16-OMK. The ultra-high performance liquid chromatography mass spectrometry (UHPLC-MS) method, already used to detect 16-OMC and 16-OMK in Arabica roasted coffee, is then more suitable for quantitative analyses. Up to now however, no quantification of coffee 16-OMD via ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) has been carried out; this largely stimulated the present study. For the first time, a simple procedure for the quantitative detection of 16-OMD in Arabica coffee has been developed, and as far as 16-OMC is concerned, fully validated in terms of specificity, linearity, concentration range, limit of detection (LOD), limit of quantification (LOQ), and repeatability following the criteria specified in the EU Commission Decision 2002/675/EC. This method proved to be very specific and sensitive. In order to avoid the chemical complexity generated by the roasting process, the method was optimized and validated on several green Arabica samples from different geographical origins.

Isolating valuable coffee diterpenes by using an inexpensive procedure

Cafestol and kahweol (C&K) are natural coffee diterpenes with high commercial value. There is thus interest in their synthesis although their isolation from natural products remains an alternative to recovering high quantities of C&K. The present work aimed at developing an isolation procedure to obtain C&K for both analytical purposes and in gram-scale from coffee beans. The procedure was divided into three steps: an initial saponification reaction optimized by Factorial Design, a choice of solvent for extraction of a crude C&K mixture based on Hansen Solubility Parameters, and a subsequent purification by column chromatography and preparative liquid chromatography in recycling mode to improve C&K mixture purity and for separate preparative collection of each diterpene. The first two steps are useful to maximize yield for C&K quantification, and the complete procedure allowed isolation of 1.5 g of C&K per 300 g of green coffee beans. This isolation procedure can be repeated 25 times using ca. US$1k in reagents, with an estimated value of product varying from US$227k to US$322k for a diterpene mass of 37.5 g, at the prevailing commercial rate.

Interaction of the Coffee Diterpenes Cafestol and 16-O-Methyl-Cafestol Palmitates with Serum Albumins.

The main coffee diterpenes cafestol, kahweol, and 16-O-methylcafestol, present in the bean lipid fraction, are mostly esterified with fatty acids. They are believed to induce dyslipidaemia and hypercholesterolemia when taken with certain types of coffee brews. The study of their binding to serum albumins could help explain their interactions with biologically active xenobiotics. We investigated the interactions occurring between cafestol and 16-O-methylcafestol palmitates with Bovine Serum Albumin (BSA), Human Serum Albumin (HSA), and Fatty Free Human Serum Albumin (ffHSA) by means of circular dichroism and fluorimetry. Circular Dichroism (CD) revealed a slight change (up to 3%) in the secondary structure of fatty-free human albumin in the presence of the diterpene esters, suggesting that the aliphatic chain of the palmitate partly occupies one of the fatty acid sites of the protein. A warfarin displacement experiment was performed to identify the binding site, which is probably close but not coincident with Sudlow site I, as the affinity for warfarin is enhanced. Fluorescence quenching titrations revealed a complex behaviour, with Stern–Volmer constants in the order of 103–104 Lmol−1. A model of the HSA-warfarin-cafestol palmitate complex was obtained by docking, and the most favourable solution was found with the terpene palmitate chain inside the FA4 fatty acid site and the cafestol moiety fronting warfarin at the interface with site I.

Mitochondrial Protection Promoted by the Coffee Diterpene Kahweol in Methylglyoxal-Treated Human Neuroblastoma SH-SY5Y Cells.

The coffee diterpene kahweol (KW; C20H26O3) is a cytoprotective agent exhibiting potent antioxidant actions, as demonstrated in several experimental models. In spite of the efforts to elucidate exactly how KW promotes cytoprotection, it was not previously examined whether KW would be able to protect mitochondria of human cells undergoing redox stress. In the present work, we have treated the human neuroblastoma SH-SY5Y cell line with KW at 0.1-10μM for 12h prior to a challenge with methylglyoxal (MG), a reactive dicarbonyl that impairs mitochondrial function. We have found that KW at 10μM suppressed the loss of mitochondrial membrane potential (MMP) and the bioenergetics decline (including decreased activity of the mitochondrial complexes I and V and reduced production of adenosine triphosphate, ATP) in the MG-treated SH-SY5Y cells. KW also prevented the MG-elicited generation of reactive oxygen and nitrogen species (ROS and RNS, respectively) in the SH-SY5Y cells. In this regard, KW exerted an antioxidant effect on the membranes of mitochondria obtained from the MG-treated cells. The mitochondria-related effects induced by KW were blocked by inhibition of the phosphoinositide 3-kinase (PI3K)/Akt or of the p38 mitogen-activated protein kinase (MAPK) signaling pathways. Moreover, silencing of the transcription factor nuclear factor E2-related factor 2 (Nrf2) suppressed the mitochondrial protection promoted by KW in the MG-challenged cells. Therefore, KW protected mitochondria by a mechanism associated with the PI3K/Akt and p38 MAPK/Nrf2 signaling pathways.

857 - Coffee diterpenes kahweol acetate and cafestol synergistically inhibit the proliferation and migration of prostate cancer cells

857 - Coffee diterpenes kahweol acetate and cafestol synergistically inhibit the proliferation and migration of prostate cancer cells

Coffee diterpenes kahweol acetate and cafestol synergistically inhibit the proliferation and migration of prostate cancer cells.

Coffee inhibits the progression of prostate cancer; however, the direct mechanism through which coffee acts on prostate cancer cells remains unclear. This study aimed to identify the key compounds of coffee that possess anti-cancer effects and to investigate their mechanisms of action. The anti-proliferation and anti-migration effects of six potentially active types of coffee compounds, including kahweol acetate, cafestol, caffeine, caffeic acid, chlorogenic acid, and trigonelline hydrochloride, were evaluated using LNCaP, LNCaP-SF, PC-3, and DU145 human prostate cancer cells. The synergistic effects of these compounds were also investigated. Apoptosis-related and epithelial-mesenchymal transition-related proteins, androgen receptor in whole cell and in nucleus, and chemokines were assessed. A xenograft study of SCID mice was performed to examine the in vivo effect of coffee compounds. Among the evaluated compounds, only kahweol acetate and cafestol inhibited the proliferation and migration of prostate cancer cells in a dose-dependent manner. The combination treatment involving kahweol acetate and cafestol synergistically inhibited proliferation and migration (combination index <1) with the induction of apoptosis, the inhibition of epithelial-mesenchymal transition, and decrease in androgen receptor, resulting in the reduction of nuclear androgen receptor in androgen receptor-positive cells. Moreover, kahweol acetate and cafestol downregulated CCR2 and CCR5 without an increase in their ligands, CCL2 and CCL5. The xenograft study showed that oral administration of kahweol acetate and cafestol significantly inhibited tumor growth. Kahweol acetate and cafestol synergistically inhibit the progression of prostate cancer. These coffee compounds may be novel therapeutic candidates for prostate cancer.

Coffee Diterpenes: before Harvesting the Bean to your Cup

Coffee Diterpenes: before Harvesting the Bean to your Cup

Coffee Diterpenes: before Harvesting the Bean to your Cup

Coffee Diterpenes: before Harvesting the Bean to your Cup

Diterpenes biochemical profile and transcriptional analysis of cytochrome P450s genes in leaves, roots, flowers, and during Coffea arabica L. fruit development

Lipids are among the major chemical compounds present in coffee beans, and they affect the flavor and aroma of the coffee beverage. Coffee oil is rich in kaurene diterpene compounds, mainly cafestol (CAF) and kahweol (KAH), which are related to plant defense mechanisms and to nutraceutical and sensorial beverage characteristics. Despite their importance, the final steps of coffee diterpenes biosynthesis remain unknown. To understand the molecular basis of coffee diterpenes biosynthesis, we report the content dynamics of CAF and KAH in several Coffea arabica tissues and the transcriptional analysis of cytochrome P450 genes (P450). We measured CAF and KAH concentrations in leaves, roots, flower buds, flowers and fruit tissues at seven developmental stages (30–240 days after flowering - DAF) using HPLC. Higher CAF levels were detected in flower buds and flowers when compared to fruits. In contrast, KAH concentration increased along fruit development, peaking at 120 DAF. We did not detect CAF or KAH in leaves, and higher amounts of KAH than CAF were detected in roots. Using P450 candidate genes from a coffee EST database, we performed RT-qPCR transcriptional analysis of leaves, flowers and fruits at three developmental stages (90, 120 and 150 DAF). Three P450 genes (CaCYP76C4, CaCYP82C2 and CaCYP74A1) had transcriptional patterns similar to CAF concentration and two P450 genes (CaCYP71A25 and CaCYP701A3) have transcript accumulation similar to KAH concentration. These data warrant further investigation of these P450s as potential candidate genes involved in the final stages of the CAF and KAH biosynthetic pathways.

Pressurized-fluid extraction of cafestol and kahweol diterpenes from green coffee

Abstract Cafestol and kahweol were extracted from green coffee beans ( C. arabica ) using a pressurized-fluid extraction (PFE) system. The effects of extracting solvent (i.e. water, ethanol and methanol), flow rate (0.5, 1.0, 2.0 and 3.0 mL · min − 1 ), extraction temperature (60, 80 and 100 °C) and time on extraction efficiency of coffee diterpenes were investigated. Quantification of diterpenes extracts was achieved by HPLC-UV method. The solvent flow rate was clearly the most significant process parameter. Diterpenes recovery resulted also influenced by temperature, except for the lowest flow rate, indicating solubility and mass transfer constraints. Kinetic degradation was also examined suggesting a thermolability of the diterpenes at 100 °C. The optimum extraction conditions were 100 °C and 3.0 mL · min − 1 using methanol. Under these conditions, PFE showed a remarkably higher efficiency compared with conventional extraction techniques. Industrial relevance Coffee is the world's most traded food commodity. Coffee-specific diterpenes (i.e. cafestol and kahweol) are high-value bioactive compounds, naturally occurring in the coffee lipid fraction. However, they are not easy to separate and likely to be unstable when isolated. High-pressure extraction, namely pressurized fluid extraction (PFE), appears as an attractive option as it reduces extraction time, thus avoiding degradation of thermolabile compounds, while requiring smaller quantities of solvent than traditional extraction methods (e.g. Soxhlet). PFE offers the possibility to operate with a wide range of liquid solvents well above their boiling point, but very little information can be found on the effect of operational conditions to efficiently recover coffee diterpenes. Such knowledge provides a basis for the rational design of PFE processes and, may be useful to allow industrial scale-up interpretation for coffee diterpenes extraction.

New approaches on the analyses of thermolabile coffee diterpenes by gas chromatography and its relationship with cup quality

A new gas chromatography method using pulsed split injector (PS-GC) was validated to quantify thermolabile diterpenes cafestol, kahweol and isokahweol in methanolysed Arabica coffee oils. Linearity was 0.99 from 8 to 69mgmL−1, recovery ranged from 99% to 101% and precision of less than 4% was obtained. Besides, Soxhlet extraction time was evaluated and Tukey׳s test indicated that the mass of diterpenes obtained in 4h is equivalent to a 16h period, which represents a space-time yield four times higher. The microwave assisted methanolysis proved to be efficient to quantitatively convert the natural diterpene esters in their respective alcohols and fatty acid methyl esters, accompanied by PS-GC. Also, the intact diterpene esters were analyzed by GC for the first time by the comparison between cold on-column (COC) and PS injection techniques. In all these stages, the molecular integrity of the thermolabile furokaurane diterpenes was maintained.The methanolysed oils from 13 samples of green Brazilian Arabica coffees were analyzed by PS-GC and the diterpenes composition varied from 8 to 12% w/w in oil and 0.7–1% in coffee beans. The ratio between cafestol and kahweol was successfully used to predict the quality of coffee even before the roasting and brewing processes.

Coffee Diterpenes Potentiate the Cytolytic Activity of KHYG-1 NK Leukemia Cells

Natural killer (NK) cells play a critical role in early host defense against tumor and virus-infected cells. Here, we report that two coffee diterpenes, cafestol and kahweol, potentiate the cytotoxic activity of KHYG-1, an NK leukemia cell line, against the NK-sensitive cell line K562. Western blot analysis and quantitative real-time PCR indicated that cafestol and kahweol markedly enhanced the expression of granzyme B. While both diterpenes had no or little effect on the expression of other cytotoxic effector molecules, kahweol elevated the phosphorylated forms of ATF-2, c-Jun and CREB transcription factors, which might participate in granzyme B gene expression. These results suggested that cafestol and kahweol upregulate granzyme B expression by activating transcription factors, leading to enhanced cytolytic activity of NK cells.

The Coffee Diterpene Kahweol Prevents Osteoclastogenesis via Impairment of NFATc1 Expression and Blocking of Erk Phosphorylation

Osteoclasts (OCLs) are multinucleated bone resorbing cells whose differentiation is regulated by receptor activator of nuclear factor kappa-B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF). It is known that inflammatory cytokines and oxidative stress stimulate differentiation of OCLs. Here we evaluated the effects of kahweol, a coffee-specific diterpene, which has been reported to possess anti-oxidant and anti-inflammatory properties, on the differentiation of bone marrow–derived macrophages (BMMs) or murine monocytic cell line RAW-D cells into OCLs. Kahweol dose-dependently inhibited the formation of tartrate-resistant acid phosphatase staining–positive OCLs from both BMMs and RAW-D cells. In addition, kahweol prevented the bone resorbing activity of OCLs. Kahweol completely abolished RANKL-stimulated phosphorylation of extracellular signal-regulated kinase and impaired phosphorylation of Akt. Moreover, the protein levels of nuclear factor of activated T cells cytoplasmic-1 (NFATc1), a master regulator for OCL differentiation; and OCL markers transcriptionally regulated by NFATc1 such as Src and cathepsin K were down-regulated by kahweol treatment. As one of the molecular mechanisms for the inhibitory effects of kahweol, we also showed that kahweol up-regulated heme oxygenase-1 and inhibited high mobility group box 1 release. Thus, kahweol in coffee is a useful constituent for inhibition of OCL differentiation.

Correction: Anti-Angiogenic and Anti-Inflammatory Properties of Kahweol, a Coffee Diterpene

The correct Reference 45 is: 45. Lawson ND, Weinstein BM (2002) In vivo imaging of embryonic vascular development using transgenic zebrafish. Dev Biol 248:307–318.