Response floor evaluation and CPH optimization

The experimental outcomes of the response values for moisture content material, chromatic parameters (L*, a*, and b*), TPC, TFC, and RSA obtained via completely different vacuum dehydration circumstances (temperature, drying time, and stress) beneath a BBD are summarized in Desk 2. Experiments had been carried out after randomization, and every response variable corresponds to the imply worth of three repetitions. Earlier than making use of the experimental design evaluation, the means of various experiments had been analyzed utilizing ANOVA, adopted by Tukey’s HSD check, to establish the technique of experiments which are completely different.

The outcomes confirmed that top temperatures might degrade phenol compounds³⁶, with 70 °C being the best temperature each for the cocoa pod and husk. The TPC of the CPH different from 1.63 to 13.37 mg GAE/g of pattern, whereas the RSA, as measured by the DPPH check, ranged from 0.003 to 0.12 mmol TE/g (Desk 2). The outcomes confirmed that the temperature, drying time, and vacuum stress parameters influenced the response variables. These outcomes had been additionally noticed by Vakula et al.³⁷ in the course of the vacuum drying of candy cherry.

The ANOVA outcomes are described in Desk 3. The R² worth for the fashions of response variables was 0.9. TPC (0.958), TFC (0.936), and RSA (0.950; as measured by the DPPH check) and the polynomial fashions utilized had been obtained from experimental knowledge. Conversely, the statistical significance values for the responses evaluated had been decrease than 0.05 in all responses. These p-values of < 0.05 point out that the mathematical fashions developed had been perfect for experimental knowledge. Moreover, Fisher’s check used to find out non adjustment (Fisher’s lack-of-fit check) yielded p-values > 0.05, suggesting that the quadratic mannequin correctly match the experimental knowledge.

The regression knowledge and their statistical significance for every response variable are summarized in Desk 4. The TPC exhibits 4 phrases that reveal its statistical significance: temperature (X₁), vacuum stress (X₃) with p-values equal to 0.02, whereas the interplay between temperature and dehydration time (X₁X₂) offered p-values of < 0.015. Moreover, the temperature issue confirmed a quadratic impact (X₁₁) (p < 0.011), which was discovered to be appreciable for the TPC. Related outcomes had been reported by Almeida-Trasviña et al.²⁴ for the coefficients of the linear mannequin X₁, X₂ in addition to for interactions (X₁X₂) and (X₂X₃), whereas the temperature and vacuum stress elements within the quadratic mannequin exhibited a big impact on the polyphenol content material.

As for the TFC, it may very well be noticed that 5 phrases had been discovered to be statistically vital, particularly, temperature (X₁) and vacuum stress (X₃), as their p-values had been decrease than 0.05 (p = 0.013 and p = 0.004). In the meantime, interplay was influenced by (X₁X₂) and (X₂X₃), with p-values of 0.031 and 0.046, respectively. The quadratic issue impact (X₁₁) was considerably influenced by vacuum stress, adopted by temperature, whereas the dehydration time confirmed statistically nonsignificant outcomes (p = 0.319). However, Rebollo-Hernanz et al.³⁸ reported that temperature is the primary variable contributing to the retrieval of flavanols 37.7% (p < 0.001), whereas dehydration time contributes simply 0.1% (p > 0.05). Šumić et al.²⁵ reported that the temperature (X₁) confirmed vital variations (p < 0.05) in the course of the crimson currants vacuum drying course of for the content material of flavonoids and whole polyphenols.

The RSA measured utilizing the DPPH methodology confirmed that 4 phrases are accountable for its statistical significance. Just like the TPC, elements X₁ and X₃ have a linear impact on the RSA, given their p-values had been decrease than 0.01, whereas the interplay between temperature and time (X₁X₂) yielded a p-value of 0.017 and, in quadratic phrases, temperature (X₁₁) yielded a p-value of 0.009; thus, each the interplay and the quadratic mannequin had been discovered to be statistically vital. These outcomes are much like these discovered by Rebollo-Hernanz et al.³⁸ of their examine when it comes to the temperature variables (X₁) (p < 0.001) and (X₁₁) (p < 0.01), whereas the interplay between temperature (X₁) and dehydration time (X₂) was statistically nonsignificant, contributing simply 2.5%. However, Šumić et al.²⁵ talked about that the temperature (X₁) and the vacuum stress (X₃) had a big affect on the inhibition coefficient at 50% (IC₅₀) for the novel DPPH.

The regression equations (Eqs. 2, 3, 4) that describe the response variables for dehydration utilizing vacuum drying for CPH are as follows:

The outcomes for TPC discovered on this examine are similar to the worth reported by Delgado-Ospina et al.³⁹, with a imply of 8.44 ± 0.04 mg GAE/g. One other examine reported that the TPC ranged from 46 to 57 mg GAE/g, though a low content material of it may be discovered as nicely. Campos-Vega et al.⁴⁰ reported that the content material variation relies on the geographical origin, genotype, and restoration system of bioactive compounds, along with the pattern’s dehydration methodology. Desk 2 exhibits that experiment No. 12 had the next TPC (13.37 mg GAE/g) in contrast with different experiments. To attain the utmost polyphenol content material, the dehydration course of was carried out at 60 °C for 8 h with a vacuum stress of fifty mbar. However, experiment No. 8 confirmed the bottom content material of polyphenols, with a mean of 1.63 mg GAE/g, beneath dehydration parameters of fifty °C, 8 h, and 100 mbar. Total, a rise within the dehydration temperature and time improved the polyphenol content material if the vacuum stress utilized was decreased to 50 mbar (Fig. 2a–c). Experiments No. 5, 10, and seven demonstrated that the TPC elevated to eight.80, 10.41, and 11.01 mg GAE/g when stress decreased from 100 to 50 mbar. These outcomes are much like these discovered by Šumić et al.²⁵ and Almeida-Trasviña et al.²⁴. Subsequently, the optimum vacuum drying circumstances had been the next: temperature (65 °C), drying time (8 h) and vacuum stress (75 mbar) to acquire a TPC of 11.17 mg GAE/g.

Fitted floor graphs for cocoa pod husk (CPH), displaying the mixed results of vacuum drying means of the issue interactions: temperature (°C), drying time (h), and stress (mbar) for TPC (mg GAE/g) whole polyphenols; TFC (mg CE/g) whole flavanols; RSA (mmol TE/g) by DPPH assay.

The TFC obtained within the completely different experiments confirmed much less variation in comparison with the TPC. Delgado-Ospina et al.³⁹ reported that the TFC in CPH was 2.9 ± 0.1 mg rutin equal per gram of pattern. Desk 2 exhibits that experiment No. 12 had a better content material of flavanols (3.09 mg CE/g), whereas a content material of 0.25 mg CE/g was obtained in experiment No. 8. Each TFC and TPC confirmed comparable outcomes for all experiments (1–15) (correlation coefficient = 0.9415; p = 0.0000). The impact of temperature, drying time, and decreased vacuum stress play an essential function in flavanol retrieval (Fig. 2d–f). Rebollo-Hernanz et al.³⁸ additionally reported that flavanol retrieval improved with a rise in temperature.

However, the RSA measured utilizing the DPPH methodology (Fig. 2g–i) was much like the imply reported by Delgado-Ospina et al.³⁹, with a price of ~ 0.058 mmol TE/g. When 5 classes are fashioned, the RSA is distributed as follows: in 2 experiments starting from 0.0035 to 0.0269 mmol TE/g, in 1 experiment starting from 0.0269 to 0.0503 mmol TE/g, in 1 experiment starting from 0.0503 to 0.0738 mmol TE/g, in 6 experiments starting from 0.0738 to 0.0972 mmol TE/g, and in 5 experiments starting from 0.0972 to 0.1206 mmol TE/g. The primary 4 correspond to experiments 4, 6, 8, and 13, with a imply of 0.03 mmol TE/g and a mean temperature and vacuum stress of 57.5 °C and 137.5 mbar, respectively, whereas the 6-experiment group introduced a imply of 0.09 mmol TE/g. This group was characterised by a mean temperature worth of 60 °C and a vacuum stress of 91.7 mbar. Lastly, the final group confirmed a imply of 0.11 mmol TE/g, with a temperature of 62 °C and a vacuum stress of 80 mbar. These outcomes are in line with these reported by Almeida-Trasviña et al.²⁴, who discovered that the perfect impact for RSA was achieved when the temperature was excessive, and the vacuum stress was low. The correlation of DPPH with TPC and with TFC was excessive: DPPH vs. TPC (r² = 0.9531) and DPPH vs. TFC (r² = 0.9572; Fig. 3). CPH is a superb supply of antioxidants. Indrianingsih et al.⁴¹ reported that this subproduct incorporates an essential bioactive compound, which has been discovered to have antioxidant and antidiabetic properties.

Correlation between response variables for cocoa pod husk: TPC (mg GAE/g) whole polyphenols; TFC (mg CE/g) whole flavanols; RSA (mmol TE/g) by DPPH assay.

Response floor evaluation and CBS optimization

The outcomes obtained with the BBD for the CBS (Desk 5) confirmed that the TPC different between 19.56 and 35.31 mg GAE/g of pattern, the TFC ranged from 5.07 to six.32 mg CE/g of pattern, and the AA ranged from 0.21 to 0.24 mmol TE/g. These outcomes point out that the polyphenol content material was largely influenced by temperature, drying time, and vacuum stress elements, in comparison with the TFC and RSA.

The ANOVA outcomes are summarized in Desk 6. As for the shell of cocoa beans, the variables TFC, and RSA had been statistically nonsignificant. The F-values for the variables had been 0.45, and 0.36, whereas the p-values had been 0.860, and 0.911, respectively. The R² variables for the response variables had been removed from encouraging, with TPC, TFC, and RSA values of 0.710, 0.449, and 0.393, respectively. Since TPC contribution was vital (p < 0.046), the mathematical mannequin was generated for this variable.

The regression equation (Eq. 5) describing the response variable for dehydration via vacuum drying for cocoa bean husk is as follows:

The illustration of the 3D response floor was generated for TPC (Fig. 4), whereas ANOVA outcomes (Desk 7) revealed just one issue as statistically vital for polyphenol content material—drying time (X₂) (p = 0.043). However, the flavanol content material and RSA yielded nonsignificant values (p > 0.05). As well as, the mathematical fashions generated weren’t match to foretell the responses. The truth is, the shortage of health was vital for the polyphenol content material variable (p < 0.05), whereas the p-value was 0.271 and 0.826 for the flavanol content material and RSA responses, respectively (Desk 6). The elements chosen didn’t present an incredible impact on the response variables however solely a slight affect of drying time on polyphenol content material within the linear mannequin. However, vacuum stress had no vital impact on the responses both. Rebollo-Hernanz et al.³⁸ reported a excessive affect of the temperature issue (X₁) on polyphenol content material, flavanol content material, and RSA, with contributions starting from 37 to 43%. The temperature ranged from 30 to 100 °C in the course of the examine. Moreover, it may very well be noticed that the drying time (X₂) didn’t have a big affect, with a contribution of 0.1–0.5%. The interplay between temperature and drying time (X₁X₂) for the three response variables was statistically nonsignificant. As for vacuum stress (X₃), Almeida-Trasviña et al.²⁴ reported that the impact of X₃ within the linear mannequin was nonsignificant, each for polyphenol content material and RSA.

Fitted floor graphs for cocoa bean shell (CBS), displaying the mixed results of vacuum drying means of the issue interactions: temperature (°C), drying time (h), and stress (mbar) for TPC (mg GAE/g).

The TPC in CBS was fairly much like that reported by Rojo-Poveda et al.⁴², starting from 3 to 43 mg GAE/g for CBS of various origins—even Peruvian samples had a median of ~ 34.5 mg GAE/g. Conversely, Cádiz-Gurrea et al.¹⁰ reported bean shell values between ~ 7 and ~ 22 mg GAE/g. These variations in polyphenol content material could also be related to geographic origin⁴³. Desk 5 signifies that experiment No. 14 had the next polyphenol content material, with elements (X₁ = 50 °C, X₂ = 5 h, and X₃ = 100 mbar) statistically nonsignificant in contrast with experiment No. 13 (X₁ = 60 °C, X₂ = 5 h, and X₃ = 150 mbar). Experiment No. 2 obtained the bottom polyphenol content material (19.56 mg GAE/g). Subsequently, the optimum vacuum drying circumstances had been the next: temperature (50 °C), drying time (5 h) and vacuum stress (100 mbar) to acquire a TPC of 29.61 mg GAE/g.

As for the flavanol content material, the experiments carried out confirmed little variation—the decrease and higher quartile yielded values starting from 5.12 to six.08 mg CE/g, and the distinction between the minimal and most values was 1.25 mg CE/g of pattern. The minimal and most values corresponded to experiments No. 9 (X₁ = 70 °C, X₂ = 3 h, and X₃ = 100 mbar) and 6 (X₁ = 50 °C, X₂ = 4 h, and X₃ = 50 mbar), respectively. The imply worth of the set of experiments was present in experiment No. 8 (5.68 mg CE/g) (X₁ = 70 °C, X₂ = 4 h and X₃ = 150 mbar). The flavanol content material on this examine was decrease in comparison with the outcomes discovered by Cádiz-Gurrea et al.¹⁰, who reported values from ~ 16 to ~ 36 mg of catechin equal per g of pattern. These variations could also be attributed to geographic origin⁴², pattern traits, extraction system, and analytical process¹⁰.

The RSA within the experiments different barely, with a decrease and higher quartile of 0.21 and 0.24 mmol TE/g of pattern, respectively. Delgado-Ospina et al.³⁹ reported values of ~ 0.071 mmol TE/g, whereas Rojo-Poveda et al.⁴² examined CBS samples of various geographic origins and reported a variation of ~ 0.03 to ~ 0.18 mmol TE/g. Peruvian samples confirmed a imply of ~ 0.15 mmol TE/g, whereas the RSA values reported by Barbosa-Pereira et al.⁴³ for the CBS of Creole and Trinitarian genotype, native to Venezuela, ranged from ~ 0.017 to ~ 0.026 mmol TE/g. The RSA values obtained on this examine had been barely greater than these reported by different researchers^{42, 43}.

Moisture content material of CPH and CBS

When the set of experiments was assessed, the moisture content material for CPH different from 8.59 to 44.12% (Desk 2). Nguyen et al.¹³ reported moisture values of 9.22%, 10.61%, and 11.99%, with temperatures of 60 °C, 80 °C, and 100 °C for vacuum drying. Three teams may be recognized when contemplating a histogram. The primary group, comprising experiments with a moisture vary of 5–10%, represented 26.67% of the experiments; the second group, comprising experiments with a moisture vary of 10–15%, accounted for 53.33% of the entire experiments; and the third group comprised experiments with moisture values > 15%, representing 20% of the experiments. Experiments with low moisture percentages had a imply temperature of 67.50 °C and a imply vacuum stress of 87.50 mbar, whereas these with a excessive moisture share yielded a mean drying temperature worth of 53.33 °C and a vacuum stress of 133.33 mbar. These outcomes had been much like these obtained by Šumić et al.²⁵, who reported that an elevated vacuum stress leads to gradual drying and produces samples with excessive moisture content material. In distinction, when vacuum stress decreases, the drying course of is quicker, producing samples with low moisture content material. Moreover, the drying time influenced the moisture content material—experiments with values starting from 5 to 10% yielded a mean of 11 h, whereas these with values starting from 10 to fifteen% had a imply of 8.67 h. Moisture content material additionally affected the response variables—experiments with excessive drying temperatures and low vacuum stress confirmed excessive TPC, TFC, and RSA. Conversely, experiments (4, 6, and eight; Desk 2) carried out at low temperature and excessive vacuum stress yielded decrease values of polyphenol (3.11 mg GAE/g) and flavanol (0.47 mg CE/g) contents in addition to an RSA of 0.02 mmol TE/g. Related outcomes had been reported by Almeida-Trasviña et al.²⁴, with decrease values for TPC and RSA for temperatures starting from 32 to 41 °C and a vacuum stress starting from ~ 420 to ~ 505 mbar.

Within the case of CBS, the moisture content material different between 0.74 and a couple of.0%, that are much like these printed by Delgado-Ospina et al.³⁹, who reported a imply of 1.9%. As well as, the moisture content material barely different as a operate of the experiment. Nevertheless, these between 1.0 and 1.6% accounted for 73.33% of all experiments. This group confirmed a imply of 53.33 °C, 3.67 h, and a vacuum stress of 83.33 mbar. Conversely, experiments with a decrease moisture share confirmed the next elements: X₁ = 70 °C, X₂ = 4 h, and X₃ = 50 mbar. As within the case of CPH, a lower within the vacuum stress offered pretty speedy drying kinetics. For this group of experiments, the TPC was better than that of experiments with the next moisture share. Nonetheless, the flavanol content material and RSA confirmed no main adjustments. As for AA, Šumić et al.²⁵ discovered {that a} greater temperature and vacuum stress yielded decrease values of RSA; thus, excessive temperatures consequence within the inactivation of antioxidants.

Chromatic properties of CPH and CBS

The colour parameters of the CPH utilized in completely different experiments confirmed that readability (L*) different between 33.05 and 64.18 models (Desk 2). Chromaticity coordinates, similar to a*, ranged from 9.17 to 13.55 models, whereas b* different between 8.60 and 27.40. These chromatic values discovered for the completely different experiments are fairly much like these obtained by Delgado-Ospina et al.³⁹ and Delgado-Ospina et al.⁴⁴. The variations in colour parameters may be attributed to the pattern’s stabilization course of, similar to drying by lyophilization and thermal remedy to inactivate enzymes, adopted by lyophilization, sun-drying, vacuum drying, infrared drying, and microwave drying, amongst others¹³. On this regard, the drying temperatures influenced the chromatic parameters, and experiments that yielded low moisture content material values (5–10%) confirmed clearer L* values (imply: 62.18 models), whereas experiments with values starting from 10 to fifteen% confirmed barely decrease L* values (imply: 59.13 models). Conversely, a lot decrease L* values had been present in experiments with a moisture content material exceeding 15% (imply: 36.47 models). These outcomes are similar to these reported by different authors the place the rise in moisture content material produces a lower within the L* parameter. As for the chromaticity parameter a*, no outstanding adjustments had been noticed regardless of the variation in moisture content material. Delgado-Ospina et al.⁴⁴ noticed comparable adjustments in dehydrated (4.02 models) and hydrated (4.56 models) CPH samples. As for the chromaticity coordinate b*, the completely different experiments confirmed appreciable variations—therapies with < 15% of moisture confirmed greater means (roughly 23.67 models), whereas these with > 15% of moisture yielded a imply of 12.74 models. The contribution of coordinate b* (yellowness) to the colour of CPH was extra related, most likely on account of its carotenoid content material. Pico Hernández et al.⁴⁵ reported a carotenoid content material of 64.35 mg/g, utilizing a supercritical fluid extraction system. Taking the correlation values into consideration, parameters L* and b* vs. moisture confirmed an adverse relation (L* vs. moisture: r =  − 0.9512; p = 0.0000; R² = 0.9049) and (b* vs. moisture: r =  − 0.9238; p = 0.0000; R² = 0.8535), whereas the chromaticity parameter a* vs. moisture confirmed little or no correlation (a* vs. moisture: r =  − 0.1648; p = 0.5572; R² = 0.0272).

With regard to the CBS, the readability parameter (L*) ranged from 31.43 to 39.69 models (Desk 5), whereas chromaticity parameters for a* and b* different between 8.32 and 15.08 models and 6.14 and 16.78 models, respectively. As for the L* parameter, a number of authors reported a variation from 45 to 51 models^{46, 47}. The variation within the L* parameter could also be related to the impact of temperature and drying occasions on account of the method of caramelization of carbohydrates and amino acids, inflicting their browning and making their readability lower towards the darkest aspect. The outcomes of chromaticity parameters, each a* and b*, had been barely much like the outcomes present in earlier research^{39, 46, 47}.

Methylxanthines and catechin derivatives of CPH and CBS

The outcomes obtained from the LC-DAD evaluation displaying the content material of theobromine, caffeine, and catechin derivatives are introduced in Desk 8. These compounds had been beforehand recognized in methanol extracts of the CPH and CBS^{10, 13}. The theobromine content material in CPH and CBS was 0.14 and seven.49 mg/g of pattern, respectively. In keeping with Nguyen and Nguyen⁴⁸, the theobromine focus in CPH obtained with numerous extraction methods, similar to 70% chloroform and ethanol, was ~ 0.004 and ~ 0.02 mg/g, respectively, whereas beneath optimum extraction circumstances with 70% ethanol for 90 min, it was ~ 0.07 mg/g. However, the theobromine content material in CBS was discovered to be inside the vary established for toasted CBS of various geographic origins and genotypes, with values starting from ~ 0.76 to ~ 9.03 mg/g⁴², though the values for CBS of various Mexican varieties ranged from 7.39 to 18.20 mg/g of pattern⁴⁹. The caffeine content material within the CPH and CBS pattern was 0.05 and 1.86 mg/g, respectively, that are decrease than these reported by Botella-Martínez et al.⁴⁷, who discovered that the caffeine content material in keeping with a particle measurement of 417–701 µm and < 417 µm was 11 mg/g and 6.13 mg/g, respectively⁴⁷. The full methylxantine content material in CPH and CBS was 0.19 and 9.35 mg/g per pattern, respectively. Rojo-Poveda et al.⁴² reported comparable values for CBS of Peruvian origin, with values between 8.3 and 9.7 mg/g.

Relating to the catechin content material of CPH and CBS, values starting from 1.23 to three.3 mg/g had been discovered, with the values for CPH being much like these reported by Valadez-Carmona et al.⁵⁰, who reported values ranging between ~ 0.84 and ~ 2.28 mg/g for various dehydration methods, with microwave dehydration as the perfect remedy. Though the CBS catechin content material was discovered to be inside the vary established by Hernández-Hernández et al.⁴⁹, with values between 0.55 and 4.66 mg/g, the catechin content material in toasted CBS of various geographical origins and genotypes was discovered to be decrease, starting from 0.012 to 0.18 mg/g⁴². Furthermore, Botella-Martínez et al.⁴⁷ reported a free catechin content material starting from 1.96 to 4.21 mg/g of pattern.

One other bioactive part present in related quantities was epicatechin; on this examine, CPH and CBS had epicatechin quantities of roughly 2.23 and 5.64 mg/g of pattern. Moreover, an investigation carried out to evaluate the impact of microwaves drying, hot-air drying, and lyophilization on the epicatechin content material of CPH resulted in values between 1.59 and three.69 mg/g⁵⁰. However, Rojo-Poveda et al.⁴² reported an epicatechin content material of 0.044–0.74 mg/g in toasted CBS. These values had been decrease than these reported within the current examine, though Hernández-Hernández et al.⁴⁹ obtained a lot better values, starting from 4.40 to 26.68 mg/g of pattern.