Beneficial Effects of Cocoa Flavanols on Microvascular Responses in Young Men May Be Dependent on Ethnicity and Lifestyle.

Cocoa flavan-3-ols affect endothelium-dependent responses in resistance vessels and microcirculation has received little attention. We tested the effects of dark chocolate consumption (396 mg total flavanols/day for 3 days) in two Groups of 10 men (18-25 years; non-smokers) each comprising equal numbers of White European (WE) and South Asian (SA) ethnicity. In Group 1, dark chocolate did not affect reactive hyperaemia in forearm muscle, but augmented muscle dilatation evoked by acute mental stress, and reactive hyperaemia and acetylcholine (ACh)-evoked dilatation in cutaneous microcirculation. Conversely, in Group 2, chocolate did not affect cutaneous reactive hyperaemia or ACh-evoked dilatation, but these responses were blunted in Group 1 relative to Group 2. Further, when Groups 1 and 2 were combined, responses were blunted in SAs relative to WEs, augmented by chocolate in SAs only. In Group 2 individuals whose ACh-evoked dilatation was attenuated by nitric oxide synthase (NOS) inhibition, ACh-evoked dilatation was not altered after chocolate, but the attenuating effect of NOS inhibition was lost. Conversely, in Group 2 individuals whose ACh-evoked dilatation was enhanced by NOS inhibition, ACh-evoked dilatation was also augmented by chocolate. We propose that in resistance and microvessels of young men, cocoa flavan-3-ols preferentially augment endothelium-dependent dilator responses whose responses are depressed by familial and lifestyle factors more prevalent in SAs than Wes. Flavan-3-ols may facilitate the NOS pathway but also influence other endothelium-dependent dilators.

Unveiling the cocoa-carob flavour gap in dark chocolates via instrumental and descriptive sensory analyses.

Roasted carob pulp (Ceratonia siliqua) is a cocoa substitute known for its faint cocoa-like resemblance. However, the cocoa-carob flavour gap remains poorly uncharacterised. This study aimed to elucidate the sensory and molecular aspects of this flavour gap in a 70 % dark chocolate formulation via a two-pronged instrumental-sensorial approach. Descriptive Sensory Analysis (DSA) revealed carob-based chocolate was significantly sweeter, less sour and astringent than conventional dark chocolate due to the high total sugar content (45-50 % DM; HPLC/RID), low titratable acidity and tannin content, respectively. As for aroma, a distinct, albeit weak, cocoa-like aroma was present in carob-based chocolate. HS-SPME-GC-MS/FID revealed this was attributed to branched-chain Strecker aldehyde generation during roasting (2-methylbutanal, 1.17 µg/g; 3-methylbutanal, 2.89 µg/g). Notably, there was a distinct lack of alkylpyrazines. Additionally, a distinct woody, tree bark-like odour was uniquely associated with carob-based chocolates. This was due to furfural generation during roasting (2.33 µg/g). In conclusion, the aroma and taste gap between cocoa and carob was successfully characterised in this study. These findings substantiate the potential of carob application in chocolate manufacturing, thus empowering confectioners to make evidence-based decisions when evaluating cocoa substitutes.

Cocoa and Carob Supplementation, Alone or in Combination with Metformin, Protects against Hepatorenal Injury in Zucker Diabetic Fatty Rats.

The liver and kidneys are crucial for glucose homeostasis and are seriously damaged in diabetes. Cocoa and carob possess antidiabetic activity, but their hepatorenal protective effects, especially when combined with antidiabetic drugs, are unknown. The aim of this study is to investigate the effects of a cocoa-carob-supplemented diet (CC), either alone or in combination with metformin, on liver and kidney damage in Zucker diabetic fatty (ZDF) rats, a type 2 diabetes model. Male ZDF animals received a control or CC-supplemented diet, with or without metformin, and Zucker lean rats were fed the control diet. The CC-supplemented diet improved glucose tolerance and insulin resistance and alleviated functional and structural alterations in the diabetic liver and renal cortex. The CC-supplemented diet also ameliorated oxidative stress, downregulated apoptosis, and improved insulin signalling and glucose homeostasis. The combination of CC and metformin boosted several benefits as certain parameters related to morphological and structural alterations, apoptosis, oxidative stress, glucose homeostasis, and insulin resistance, were improved in comparison to animals receiving the CC-supplemented diet or metformin alone; these include the following: apoptotic index, Bax, hepatic insulin receptor or glutathione content, among others. These results demonstrate that the CC-supplemented diet alleviates the hepatorenal damage in type 2 diabetic ZDF rats, highlighting its potential alone or as an adjuvant therapy.

Molecular Changes during Germination of Cocoa Beans, Part 1.

Some germination is known to occur during the process of fermentation in cocoa beans. The impact of this biological process on the course of cocoa fermentation is not known and was thus investigated. In order to determine the impact of germination at the molecular level as well as on flavor, an untargeted metabolomics approach using Ultra Performance Liquid Chromatography-Electrospray Ionization-Time of Flight-Mass Spectrometry (UPLC-ESI-ToF-MS) with simultaneous acquisition of low- and high-collision energy mass spectra (MSe) was performed. Extracts of raw and germinated cocoa beans of the same origin were measured and compared for characteristic differences by unsupervised principal component analysis. OPLS-DA revealed 12-hydroxyjasmonic acid (HOJA) sulfate, (+)-catechin and (-)-epicatechin as most down-regulated compounds as well as two hydroxymethylglutaryl (HMG) glucosides A and B among others as decisive up-regulated compounds in the germinated material. Additionally, further HMG glucosides and 12-hydroxyjasmonic acid could be identified in cocoa for the first time by coelution with isolated and synthesized reference compounds. HOJA sulfate, which has been postulated in cocoa, and HOJA were revealed to impart bitter and astringent taste qualities.

An untargeted metabolomics approach applied to the study of the bioavailability and metabolism of three different bioactive plant extracts in human blood samples.

Advances in the understanding of bioavailability and metabolism of bioactive compounds have been achieved primarily through targeted or semi-targeted metabolomics approaches using the hypothesis of potential metabolized compounds. The recent development of untargeted metabolomics approaches can present great advantages in this field, such as in the discovery of new metabolized compounds or to study the metabolism of compounds from multiple matrices simultaneously. Thus, this study proposes the use of an untargeted metabolomics strategy based on HPLC-ESI-QTOF-MS for the study of bioavailability and metabolism of bioactive compounds from different vegetal sources. Specifically, this study has been applied to plasma samples collected in an acute human intervention study using three matrices (Hibiscus sabdariffa, Silybum marianum and Theobroma cacao). This approach allowed the selection of those significant variables associated with exogenous metabolites derived from the consumption of bioactive compounds for their subsequent identification. As a result, 14, 25 and 3 potential metabolites associated with supplement intake were significantly detected in the plasma samples from volunteers who ingested the H. sabdariffa (HS), S. marianum (SM) and T. cacao (TC) extracts. Furthermore, Tmax values have been computed for each detected compound. The results highlight the potential of untargeted metabolomics for rapid and comprehensive analysis when working with a wide range of exogenous metabolites from different plant sources in biological samples.

Flavan-3-ols and Vascular Health: Clinical Evidence and Mechanisms of Action.

Cardiovascular diseases (CVDs) are one of the main causes of mortality and morbidity worldwide. A healthy diet rich in plant-derived compounds such as (poly)phenols appears to have a key role in improving cardiovascular health. Flavan-3-ols represent a subclass of (poly)phenols of great interest for their possible health benefits. In this review, we summarized the results of clinical studies on vascular outcomes of flavan-3-ol supplementation and we focused on the role of the microbiota in CVD. Clinical trials included in this review showed that supplementation with flavan-3-ols mostly derived from cocoa products significantly reduces blood pressure and improves endothelial function. Studies on catechins from green tea demonstrated better results when involving healthy individuals. From a mechanistic point of view, emerging evidence suggests that microbial metabolites may play a role in the observed effects. Their function extends beyond the previous belief of ROS scavenging activity and encompasses a direct impact on gene expression and protein function. Although flavan-3-ols appear to have effects on cardiovascular health, further studies are needed to clarify and confirm these potential benefits and the rising evidence of the potential involvement of the microbiota.

Exploring volatile compounds and microbial dynamics: Kluyveromyces marxianus and Hanseniaspora opuntiae reduce Forastero cocoa fermentation time.

Traditional cocoa bean fermentation is a spontaneous process and can result in heterogeneous sensory quality. For this reason, yeast-integrated starter cultures may be an option for creating consistent organoleptic profiles. This study proposes the mixture of Hanseniaspora opuntiae and Kluyveromyces marxianus (from non-cocoa fermentation) as starter culture candidates. The microorganisms and volatile compounds were analyzed during the cocoa fermentation process, and the most abundant were correlated with predominant microorganisms. Results showed that Kluyveromyces marxianus, isolated from mezcal fermentation, was identified as the dominant yeast by high-throughput DNA sequencing. A total of 63 volatile compounds identified by HS-SPME-GC-MS were correlated with the more abundant bacteria and yeast using Principal Component Analysis and Agglomerative Hierarchical Clustering. This study demonstrates that yeasts from other fermentative processes can be used as starter cultures in cocoa fermentation and lead to the formation of more aromatic esters, decrease the acetic acid content.

Conformational preferences of cocoa oligomeric proanthocyanidins and their influence on polarity.

Proanthocyanidins (OPACs) are the second largest class of plant metabolites after lignans. Although knowledge of their 3D conformations would add greatly to our understanding of their biological properties, very little has been published on the conformations of OPACs with a degree of polymerization (DP) above 4. We investigated the conformations of the linear epicatechin oligomers, prominent representatives of OPACs prevalent in apples and cocoa, where the epicatechin units are interconnected through the 4ß-8 bonds. For DP-2 to DP-10 oligomers, conformational preferences reflected in the arrangement of consecutive flavan-3-ol units, are characterized by the φ torsion. For dimers, there are two energy wells corresponding to two preferred φ torsions, designated as compact and extended form. This behaviour is preserved in OPACs with higher DPs, but the most energetically favoured conformations are a combination of both, with compact-only or extended-only conformations being very unlikely. Thus, oligomers with DP ≥ 7 tend to assume an overall conformation approximating a spherical shape. This shape has a significant influence on the polarity of the OPAC oligomers expressed as 3D polar surface area, calculated using Spartan software for geometry-optimized 3D models, and possibly on other physicochemical properties. The results of polarity calculations provide a molecular-level rationale for the polarity-based chromatographic separation of the cocoa B-type procyanidins with DP range 4 to 10. In our experiments, using centrifugal partition chromatography (CPC) (a solvent system consisting of EtOAc-EtOH-water (6:1:5) v/v/v with aqueous phase stationary and upper phase mobile) we found that an enriched mixture of proanthocyanidins eluted first DP-1 (epicatechin) followed by consecutive elution of the DP-2 to DP-10 in the linear 4ß-8 form. We demonstrated that such separation would not be possible if compact-only or extended-only conformations were present in solution. However, for the energy-favoured, spherically shaped conformations, the observed CPC elution order is fully justified.

Removal of Rhodamine-B dye from Aqueous Solutions Using Alkaline-Modified Activated Carbon from Cocoa Pod Husk.

Rhodamine-B (RhB) dye in wastewater poses health and environmental risks due to respiratory and eye infections, neurotoxicity, and carcinogenicity, necessitating proper disposal for risk mitigation. This study investigates RhB removal from water using NaOH-modified activated carbon derived from cocoa pod husk (CPHAC). Employing a face-centered central composite design, operational variables were optimized to achieve maximum RhB dye removal efficiency. The study reveals a removal efficiency of 98.87 ± 0.84% under optimized conditions: adsorbent dose of 1.34 g, contact time of 71.59 min, and an initial RhB concentration of 6.61 ppm. The Freundlich isotherm model demonstrated a good fit, suggesting that RhB removal is governed by heterogeneity and multilayer adsorption. Kinetic experiments revealed that adsorption follows a pseudo-second-order model, indicating likely irreversible adsorption with dye molecules forming chemical bonds on CPHAC's surface. Overall, this study demonstrates the effectiveness of CPHAC as an efficient adsorbent for RhB removal from water.

Cocoa extract induces browning of white adipocytes and improves glucose intolerance in mice fed a high-fat diet.

Cocoa extract (CE) offers several health benefits, such as antiobesity and improved glucose intolerance. However, the mechanisms remain unclear. Adipose tissue includes white adipose tissue (WAT) and brown adipose tissue. Brown adipose tissue leads to body fat reduction by metabolizing lipids to heat via uncoupling protein 1 (UCP1). The conversion of white adipocytes into brown-like adipocytes (beige adipocytes) is called browning, and it contributes to the anti-obesity effect and improved glucose tolerance. This study aimed to evaluate the effect of CE on glucose tolerance in terms of browning. We found that dietary supplementation with CE improved glucose intolerance in mice fed a high-fat diet, and it increased the expression levels of Ucp1 and browning-associated gene in inguinal WAT. Furthermore, in primary adipocytes of mice, CE induced Ucp1 expression through ß3-adrenergic receptor stimulation. These results suggest that dietary CE improves glucose intolerance by inducing browning in WAT.

Correlation of rheology and oral tribology with sensory perception of commercial hazelnut and cocoa-based spreads.

This study examined the effects of spread formulation and the structural/lubricant properties of six different commercial hazelnut and cocoa spreads on sensory perception. Rheology, tribology, and quantitative descriptive analysis (QDA) was assessed by also evaluating the correlation coefficients between the quality descriptor and the rheological and textural parameters. The viscosity was evaluated at different temperatures to better simulate conditions before and after ingestion. Tribological analysis was executed at 37°C to mimic the human oral cavity. The effect of saliva presence and the number of runs on tribological behaviors was investigated. Moreover, textural, calorimetric, and particle size distribution measurements were performed to reinforce the correlation between structural/thermal parameters (e.g., firmness, stickiness, sugar melting point) and sensory aspects. "Visual viscosity," defined as a sensory attribute evaluated prior to consumption, negatively correlated with apparent viscosity measured at 20°C and 10 s-1, whereas "body," defined during oral processing and related to creaminess, positively correlated with apparent viscosity measured at 37°C and 50 s-1. These attributes were mainly influenced by particulate microstructure and solid volume fraction within the formulation. Textural stickiness positively correlated with sensory "adhesiveness" and was related to fat composition and milk powder addition, while "sweetness" was related to sucrose content and sugar melting enthalpy. Tribological data provided meaningful information related to particle-derived attributes, as well as after-coating perception (fattiness/oiliness), thus better predicting food evolution during oral consumption.

Diet enriched with high-phenolic cocoa potentiates hippocampal brain-derived neurotrophic factor expression and neurogenesis in healthy adult micewith subtle effects on memory.

Cocoa is widely known for its health benefits, but its neurocognitive impact remains underexplored. This preclinical study aimed to investigate the effects of cocoa and cocoa polyphenols on hippocampal neuroplasticity, cognitive function and emotional behavior. Seventy young-adult C57BL/6JRj male and female mice were fed either a standard diet (CTR) or a diet enriched with 10% high-phenolic content cocoa (HPC) or low-phenolic content cocoa (LPC) for at least four weeks. In a first experiment, behavioral tests assessing exploratory behavior, emotional responses and hippocampal-dependent memory were conducted four weeks into the diet, followed by animal sacrifice a week later. Adult hippocampal neurogenesis and brain-derived neurotrophic factor (BDNF) expression in the hippocampus and prefrontal cortex were evaluated using immunohistochemistry and western blot. In a different experiment, hippocampal synaptic response, long-term potentiation and presynaptic-dependent short-term plasticity were studied by electrophysiology. Cocoa-enriched diets had minimal effects on exploratory activity and anxiety-like behavior, except for reduced locomotion in the LPC group. Only the HPC diet enhanced object recognition memory, while place recognition memory and spatial navigation remained unaffected. The HPC diet also increased adult hippocampal neurogenesis, boosting the proliferation, survival and number of young adult-born neurons. However, both cocoa-enriched diets increased immobility in the forced swimming test and hippocampal BDNF expression. Hippocampal electrophysiology revealed no alterations in neuroplasticity among diets. The results were mostly unaffected by sex. Overall, the HPC diet demonstrated greater potential regarding cognitive and neuroplastic benefits, suggesting a key role of cocoa flavanols in dietary interventions aimed at enhancing brain health.

Optimization of subcritical water extraction for pectin extraction from cocoa pod husks using the response surface methodology.

This study optimized subcritical water extraction (SWE) conditions to maximize pectin yield from cocoa pod husk (CPH) and compared the characteristics of CPH pectin extracted through SWE with those of CPH pectin obtained through conventional extraction (CE) with citric acid. The Box-Behnken experimental design was employed to optimize SWE and examine the influence of process parameters, including temperature (100 °C-120 °C), extraction time (10-30 min), and solid:liquid ratio (SLR) (1:30-2:30 g/mL), on pectin yield. The maximum pectin yield of 6.58% was obtained under the optimal extraction conditions of 120 °C for 10 min with 1:15 g/mL SLR and closely corresponded with the predicted value of 7.29%. Compared with CE, SWE generated a higher yield and resulted in a higher degree of esterification, methoxyl content, and anhydrouronic acid value but a lower equivalent weight. The extracted pectin was pure, had low-methoxyl content, and similar melting and degradation temperatures.

Molecular Changes during Germination of Cocoa Beans, Part 2.

A recently published untargeted metabolomics approach toward marker compounds of cocoa germination revealed and identified 12-hydroxyjasmonic acid sulfate, (+)-catechin, and (-)-epicatechin as the most downregulated compounds and two hydroxymethylglutaryl glucosides (HMG gluc) A and B, among others, as the decisive upregulated compounds in the germinated material. These findings were quantitatively evaluated using ultrahigh-performance liquid chromatography-tandem mass spectrometry not only in previously examined sample material but also in a vastly expanded array of cocoa samples of different provenience and process and in cocoa products such as cocoa liquor and chocolate. Hereby, yields of newly identified HMG gluc derivatives could be determined in raw, fermented, germinated, and alternatively processed cocoa, and isomers of HMG gluc A and B could be established as key process indicators. Based on unsupervised clustering and supervised classification, models could identify germinated samples in testing sets consisting of raw, fermented, and germinated samples.

Inclusion of cocoa bran in the diet of lambs and its effect on reproductive parameters.

The objective of the study was to evaluate the effect of the inclusion of cocoa bran in the diet of lambs and its effect on reproductive parameters. For this, 40 lambs were randomly assigned to four treatments, and including 0, 10, 20 and 30% levels of cocoa bran in the concentrate. Blood was collected to measure cholesterol and testosterone and semen for physical and morphological evaluation; testicular biometry and morphometry were also evaluated. There was significant difference (P < 0.05) in body weight and tubulosomatic index between the lambs in the control treatment and those in the 30% cocoa bran treatment. There was no difference in testicular biometry, physical and morphological parameters of fresh semen, testicular morphometry, and volumetric ratio between lambs in all the treatments (P < 0.05). In addition, there was no difference in plasma cholesterol or testosterone concentration (P > 0.05). Thus, it is possible to include up to 30% of cocoa bran in diet without affecting the reproductive parameters of lambs.

In Vitro Protective Effect of Pea-Derived Peptides (PPs) via the Keap1/Nrf2 Signaling Pathway on Alpha-Gliadin-Sensitizing Peptide Induced Cacao-2 Cells.

SCOPE: Celiac disease (CD) is an allergic intestinal disease caused mainly by gliadin in wheat, which is widespread in the population and currently lacks effective treatment. α-Gliadin peptides cause cellular damage by substantially increasing cellular reactive oxygen species (ROS) levels. METHODS AND RESULTS: This study investigates the protective effect of 11 pea-derived peptides (PPs) on ɑ-gliadin peptide (P31-43) treated Caco-2 cells. Results show that cells treated with PP2, PP5, and PP6 peptides significantly reduce the cell mortality caused by P31-43. Three PPs significantly reduce the P31-43-induced decrease in ROS levels to control levels, and there is no difference between them and the vitamin C (Vc) group. The results in terms of antioxidant-related enzymes show that PPs significantly decrease superoxide dismutase activity (SOD), glutathione reductases (GR), and glutathione (GSH)/oxidized glutathione (GSSG) levels, thus significantly enhancing the antioxidant level of cells. By studying the key proteins of the Kelch-like ECH-associated protein 1 (Keap1)/NF-E2-related factor 2 (Nrf2) pathway, it is found that PPs activate the Keap1/Nrf2 signaling pathway. CONCLUSION: The study finds that peptides from peas can effectively alleviate ɑ-gliadin peptide-induced cell damage. The discovery of these food-derived peptides provides novel potential solutions for the prevention and treatment of CD.

Flow properties of single origin chocolates: Effect of product formulation and particle size.

The objective of this research was to evaluate changes in flow behavior of chocolate during chocolate grinding using a stone grinder as affected by chocolate formulation. Three different types of chocolates were evaluated. Two chocolates without milk added (70% chocolate) and two chocolates with milk added and with different amounts of cocoa nibs (30% chocolate and 14% chocolate) were tested. For the 70% chocolates, nibs of two different origins were used; therefore, a total of four samples were evaluated. Chocolates were processed in a stone grinder, and samples were taken as a function of grinding time. For each timepoint, the flow behavior of the samples was measured using a rotational rheometer and fitted to the Casson model. Particle size was measured using a laser scattering instrument. Results showed that yield stress increased linearly while the Casson plastic viscosity decreased exponentially with grinding time (smaller particles). Particle size distribution of the chocolates showed a prominent bimodal distribution for short grinding times (∼9 h) with small (∼15 µm) and large (∼100 µm) particles; with longer grinding time, the population of larger particles decreased. Yield stress values were higher for the 70% chocolate, but they were not very different between the two milk chocolates tested. The Casson plastic viscosity was greatest for the 70% chocolate, followed by the 30% chocolate. The 14% chocolate had the lowest Casson plastic viscosity. Changes of Casson plastic viscosity with particle size were more evident for the dark chocolates compared to the milk ones. These results are helpful to small chocolate producers who need better understanding of how the formulation and grinding of chocolate affect its flow behavior, which will ultimately affect chocolate handling during production.

Chemical Composition of Fat Bloom on Chocolate Products Determined by Combining NMR and HPLC-MS.

To reduce unwanted fat bloom in the manufacturing and storage of chocolates, detailed knowledge of the chemical composition and molecular mobility of the oils and fats contained is required. Although the formation of fat bloom on chocolate products has been studied for many decades with regard to its prevention and reduction, questions on the molecular level still remain to be answered. Chocolate products with nut-based fillings are especially prone to undesirable fat bloom. The chemical composition of fat bloom is thought to be dominated by the triacylglycerides of the chocolate matrix, which migrate to the chocolate's surface and recrystallize there. Migration of oils from the fillings into the chocolate as driving force for fat bloom formation is an additional factor in the discussion. In this work, the migration was studied and confirmed by MRI, while the chemical composition of the fat bloom was measured by NMR spectroscopy and HPLC-MS, revealing the most important triacylglycerides in the fat bloom. The combination of HPLC-MS with NMR spectroscopy at 800 MHz allows for detailed chemical structure determination. A rapid routine was developed combining the two modalities, which was then applied to investigate the aging, the impact of chocolate composition, and the influence of hazelnut fillings processing parameters, such as the degree of roasting and grinding of the nuts or the mixing time, on fat bloom formation.

Optimization of HS-SPME/GC-MS Method for Determining Volatile Organic Compounds and Sensory Profile in Cocoa Honey from Different Cocoa Varieties (Theobroma cacao L.).

This study aimed to develop an analytical method using HS-SPME/GC-MS to determine the volatile organic compound (VOC) profiles and evaluate the sensory attributes of cocoa honey from four cocoa varieties (CCN51, PS1319, SJ02, and Parazinho). Using a multivariate factorial experimental design, the HS-SPME/GC-MS method was optimized to determine the VOC profiles. Twenty previously trained tasters participated in the ranking descriptive analysis, while 108 consumers participated in the acceptance and purchase intention tests. A total of 84 volatile organic compounds were identified from various chemical classes, including acids, alcohols, aldehydes, esters, ketones, monoterpenes, oxygenated monoterpenoids, sesquiterpenes, and oxygenated sesquiterpenoids. Palmitic acid was the compound found in the highest concentration in all varieties (5.13-13.10%). Multivariate analysis tools identified key compounds for differentiation and grouping of the samples. The results revealed that the variety significantly influenced both the VOCs' concentrations and sensory profiles. The CCN51, PS1319, and SJ02 varieties exhibited the highest diversity of VOCs and sensory attributes. Notably, the SJ02 and CCN51 varieties demonstrated superior acceptability and purchase intention, with means ranging from 7.21 and 7.08 to 3.71 and 3.56, respectively. These results indicate their potential as promising sources of cocoa honey for the food industry.

Investigating Key Volatile Compound Diffusion in Cocoa Beans during Yeast Fermentation-like Incubation.

An experimental setup was devised to investigate the permeability of cocoa bean seed coat and pulp to key volatile compounds during fermentation. Four labeled compounds (ethyl acetate-d3, ethyl octanoate-d15, 2-phenylethanol-d5, linalool-d5) and 2 unlabeled (beta-damascenone, delta-decalactone) were chosen for the investigation. The beans (cotyledons), depulped beans, or pulped beans were immersed separately in a concentrated solution of these volatile compounds at 36 or 46 °C for durations ranging from 3 to 120 h. The imbibed beans were dissected, and the cotyledons were analyzed by SPME-GC/MS. The diffusion of volatile compounds from the external solution to the seed was categorized into three groups: (1) not diffusible (ethyl octanoate-d15); (2) semidiffusible (ethyl acetate); and (3) totally diffusible (2-phenylethanol-d5, linalool-d5, beta-damascenone, delta-decalactone). The impact of the yeast on volatile compound diffusion was also investigated by immerging the pulped beans into the same concentrated solution with a yeast starter. Results highlighted the positive role of yeast in the diffusion of volatile compounds. The starter positively contributed to volatile compound diffusion after a transition phase occurring at approximately 48 h of fermentation, enriching the cocoa beans with key aromatic volatile compounds.