Alkali-Catalyzed Organosolv Treatment of Oat Bran for Enhanced Release of Hydroxycinnamate Antioxidants: Comparison of 1-and 2-Propanol

: Oat bran (OB) may be regarded as food industry by-product, with a high perspective as a bioresource in the production of high-value polyphenolic substances. On this basis, the investigation presented herein aimed at (i) using 1-and 2-propanol organosolv treatment and studying the effect of an alkali (sodium hydroxide) catalyst on releasing bound polyphenols, (ii) establishing models of polyphenol recovery by employing severity and response surface methodology, and (iii) investigating the polyphenolic proﬁle of the extracts produced. Yield in total polyphenols as a function of treatment severity was satisfactorily described by linear models, indicating that for both 1-and 2-propanol treatments, temperature and time could be used interchangeably to adjust severity. Furthermore, the 1-propanol process was found to be more efﬁcient at lower severity compared to the 2-propanol process. The optimization using response surface methodology revealed that, under identical condition settings ( t = 300 min, T = 90 ◦ C), the 1-propanol treatment afforded a total polyphenol yield of 17.15 ± 0.51 mg ferulic acid equivalents per g − 1 dry mass, whereas the 2-propanol treatment gave a yield of 14.78 ± 1.11 mg ferulic acid equivalents per g − 1 dry mass. Liquid chromatography–tandem mass spectrometry analyses showed that the extract produced from the 1-propanol treatment was signiﬁcantly more enriched in ferulic acid and p -coumaric acid compared to the extract generated with the 2-propanol treatment. Moreover, the antioxidant activity of the extracts was in concurrence with the polyphenolic composition. Based on the evidence, the alkali-catalyzed, 1-propanol organosolv treatment of OB is proposed as a sustainable and efﬁcient methodology to recover multipurpose natural antioxidants.


Introduction
Population growth around the world has put an unprecedented strain on how bioresources are used, leading to non-regulated overexploitation, ecosystem degradation, and serious environmental aggravation. The production of extraordinary amounts of waste is a matter of utmost importance in relation to agriculture and food production. This leftover biomass is composed of biological tissues with a high organic content; as a result, inappropriate handling and excessive disposal in landfills may pose immediate concerns to ecosystem conservation, public health, and environmental contamination [1,2]. Based on the aforementioned factors, linear economy models have, up to this point, been widely acknowledged to be both inefficient and environmentally harmful. As a result, circular economy aspects are becoming universally accepted by embracing sustainable routes of agricultural development and food manufacturing. Valorization of food wastes to produce

Oat Bran
Commercially available oat bran (OB) was purchased from a local grocery store (Chania, Greece), and upon receipt, it was milled in a table blender. The milled material was sieved, and the powder, with an average particle diameter < 0.5 mm, was collected and stored in air-tight plastic vials at 4 • C for no longer than 7 days. This material was used for all organosolv treatments tested.

Solvent Assay and Acid-Catalyzed Organosolv Treatment
The solvents considered for the organosolv treatments were 1-and 2-propanol. These solvents were initially tested for their efficacy in polyphenol extraction from OB. For this purpose, an exact mass of 1 g of milled OB was combined with 10 mL of pure 1-or 2-propanol, or 20, 40, 60, and 80 (v/v) aqueous mixtures. For all extractions, a hotplate (Witeg, Wertheim, Germany) at a stirring speed of 500 rpm was used at a resident time of 210 min and a constant temperature of 70 • C [29]. Upon completion of the extraction, centrifugation at 10,000× g was employed to separate the solid residue, and the clear extract was used for any further analysis.
To accomplish the organosolv treatments, a previously published protocol was implemented [27]. Thus, mixtures with the optimum 1-or 2-propanol/water proportion were spiked with concentrated sodium hydroxide solution to provide a final sodium hydroxide concentration (C SoHy ) of 1.5% (w/v). Following this, 10 mL of each of these solvent systems was mixed with 1 g OB, and treatments were carried out under continuous stirring at 500 rpm and 90 • C for 60, 180, and 300 min. After each treatment, cell debris was removed by centrifugation at 10,000× g.

Severity Determination
Taking into consideration the treatment temperature and time, severity may be appraised and used as a tool to compare the harshness of the processes, as shown below [30,31]: SF is termed as the severity factor, R o is considered to represent the severity, and the value 100 • C is regarded as the reference temperature. The empirical factor 14.75 is related to the treatment temperature and activation energy. The combined severity factor (CSF) is Environments 2023, 10, 118 4 of 14 a more integrated form of SF, and takes into account the pH of the treatment, which may be a critical parameter that affects biomatrix (OB) disintegration [32]: It has also been proposed that an alternative expression, termed as CSF , may represent a fairer comparison of treatment severities at widely different pHs [32]:

Experimental Design and Treatment Optimization
Utilizing the response surface approach, the organosolv treatment was optimized. Treatment temperature, T, and residence time, t, were used as independent (process) variables, and the yield in total polyphenols (Y TP ), was used as the response. An 11-point central composite with three center points was used as the experimental design. Both independent variables (t, T) were codified in three levels: −1, 0, and 1, following a procedure described elsewhere [33]. Table 1 displays the actual and codified levels of each variable. Table 1. Information on the ranges of the process variables used and their actual and coded levels.

Process Variables
Codes The ranges for each variable evaluated were chosen based on recent literature as well as preliminary research [27]. The temperature of 90 • C was selected as a safe upper limit to avoid high vapor pressure when approaching solvent boiling point, since the solvent systems used were aqueous 1-and 2-propanol mixtures. The statistical significance of each of the resulting model's individual coefficients as well as the overall statistical significance of the models (R 2 , p) were evaluated using the analysis of variance (ANOVA) and lack-of-fit tests, both with a minimum 95% significance level.

Determination of Total Polyphenols and Antioxidant Properties
Total polyphenols were determined following a previously published Folin-Ciocalteu method [34]. Results were given as ferulic acid equivalents using a standard curve (50-700 mg L −1 , R 2 = 0.9987). The antiradical activity (A AR ) and the ferric-reducing power (P R ) were measured using well-established methodologies, as previously described [35].

Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS)
To identify and quantify the principal polyphenols found in OB, a TSQ Quantum Access MS/MS detector (Thermo Scientific, Waltham, MA, USA) was used in conjunction with an ACQUITY Binary Solvent management and ACQUITY Sample management (Waters, Milford, MA, USA). Separations were accomplished on a Fortis SpeedCore C18, 100 mm × 2.1 mm (2.6 µm), at 40 • C, and using a 5 µL injection volume and a flow rate of 0.3 mL min −1 . A linear gradient elution program was employed with the mobile phases (A) water (1% acetic acid), and (B) methanol (1% acetic acid), by applying a linear gradient elution program as follows: 0 min, 0% B; 20 min, 100% B. Negative ionization mode was employed for mass spectra acquisition, with a sheath gas pressure of 30 (arbitrary units), capillary temperature set at 300 • C, collision pressure at 1.5 mTorr, and auxiliary gas pressure at 15 (arbitrary units). p-coumaric acid and ferulic acid quantitation was carried out by external standard calibration curves using the highest intensity fragment that was produced during the collision-induced dissociation (CID) of the selected precursor molecular ion.

Statistical Analysis
JMP™ Pro 16 (SAS, Cary, NC, USA) was employed to perform distribution analysis, set up response surface experimental designs, and calculate attendant statistics (ANOVA, lack of goodness of fit). SigmaPlot™ 15.0 (Systat Software Inc., San Jose, CA, USA) was used for all linear and nonlinear regressions with a significance level of at least 95%. Organosolv treatments were performed at least twice, and analytical measurements were performed in triplicate. The displayed values are the mean ± standard deviation (SD).

Solvent Testing
The initial stage in the organosolv process development was to identify a water/alcohol proportion that would enable increased recovery of oat polyphenols. For this purpose, a screening was performed using water/alcohol mixtures with variable compositions (Figure 1). For the mixtures of 1-propanol/water, a 40% (v/v) solution was shown to provide the highest Y TP , which was significantly different (p < 0.05). On the other hand, the most efficient polyphenol recovery was accomplished with 2-propanol/water mixtures of 60% (v/v), whereas the 40% mixture was of significantly lower efficacy. Based on this outcome, 40% solutions of 1-propanol and 60% solutions of 2-propanol were selected for further testing. elution program as follows: 0 min, 0% B; 20 min, 100% B. Negative ionization mode was employed for mass spectra acquisition, with a sheath gas pressure of 30 (arbitrary units), capillary temperature set at 300 °C, collision pressure at 1.5 mTorr, and auxiliary gas pressure at 15 (arbitrary units). p-coumaric acid and ferulic acid quantitation was carried out by external standard calibration curves using the highest intensity fragment that was produced during the collision-induced dissociation (CID) of the selected precursor molecular ion.

Statistical Analysis
JMP™ Pro 16 (SAS, Cary, NC, USA) was employed to perform distribution analysis, set up response surface experimental designs, and calculate attendant statistics (ANOVA, lack of goodness of fit). SigmaPlot™ 15.0 (Systat Software Inc., San Jose, CA, USA) was used for all linear and nonlinear regressions with a significance level of at least 95%. Organosolv treatments were performed at least twice, and analytical measurements were performed in triplicate. The displayed values are the mean ± standard deviation (SD).

Solvent Testing
The initial stage in the organosolv process development was to identify a water/alcohol proportion that would enable increased recovery of oat polyphenols. For this purpose, a screening was performed using water/alcohol mixtures with variable compositions (Figure 1). For the mixtures of 1-propanol/water, a 40% (v/v) solution was shown to provide the highest YTP, which was significantly different (p < 0.05). On the other hand, the most efficient polyphenol recovery was accomplished with 2-propanol/water mixtures of 60% (v/v), whereas the 40% mixture was of significantly lower efficacy. Based on this outcome, 40% solutions of 1-propanol and 60% solutions of 2-propanol were selected for further testing.

Assessment of Treatment Severity
A previous recent study clearly demonstrated the high effectiveness of the alkalicatalyzed ethanol organosolv treatment in the recovery of polyphenols from wheat bran [27]. In that study, exceptionally increased yields of total polyphenols were attained using solvent systems that contained 1.5% sodium hydroxide as a catalyst. On such a basis, solvent systems composed of 40% 1-propanol/1.5% sodium hydroxide and 60% 2-propanol/1.5% sodium hydroxide were employed to carry out organosolv treatments Environments 2023, 10, 118 6 of 14 by testing various combinations of time and temperature, which corresponded to various severity levels ( Table 2). For the treatment accomplished with 1-propanol/sodium hydroxide, significantly higher Y TP (p < 0.05) could be obtained at CSF higher than −10.64 and CSF higher than 7.19. Likewise, the treatment with 2-propanol/sodium hydroxide afforded a significantly higher Y TP at CSF and CSF levels that corresponded to −10.49 and 7.41. For both treatments, the CSF and CSF levels required for maximum Y TP were very similar to −10.85 and 7.63, respectively, which were determined for the alkali-catalyzed ethanol organosolv treatment of wheat bran [27]. Table 2. The values determined for the combined severity factor (CSF), the alternative combined severity factor (CSF ), and yield in total polyphenols (Y TP ) as a function of the temperature and time settings. As a following step in the examination of the effect of treatment severity on the polyphenol recovery performance, linear regressions were attempted between Y TP and either CSF or CSF values to identify any possible correlation pattern. Figure 2 visualizes the outcome of the linear regressions established, which were of high statistical significance, as shown below from the linear models derived: The use of Equation (6) enabled the determination of the theoretically maximum Y TP that could be achieved, which was 17.25 mg FAE g −1 DM. Similarly, the use of Equation (7) gave a theoretical maximum Y TP of 17.28 mg FAE g −1 DM. Both values were virtually identical to the experimental value of 17.14 mg FAE g −1 DM (Table 2). In the same manner, the theoretical maximum values of Y TP computed from the Equations (8) and (9) were 14.57 and 14.63 mg FAE g −1 DM, respectively, which perfectly matched the experimental value of 14.50 mg FAE g −1 DM. These findings strongly indicated that, using the empirical models established, reliable predictions on Y TP could be generated based on any combination of T and t within the ranges tested. Environments 2023, 10, x FOR PEER REVIEW 7 of 15 of 14.50 mg FAE g −1 DM. These findings strongly indicated that, using the empirical models established, reliable predictions on YTP could be generated based on any combination of T and t within the ranges tested.

Response Surface Treatment Optimization
Considering the evidence that emerged from the linear models encompassing severity, 1-and 2-propanol/sodium hydroxide treatments exhibited differentiated effects on polyphenol recovery from OB. Therefore, response surface methodology was deployed to fully assess the effect of both solvents and provide alternative means of process modeling. For this purpose, temperature, T, and time, t, were used as the process variables. With such an approach, the effects of T and t on treatment performance could be further evaluated and possible cross (synergistic) functions could be revealed. Overall model appraisal and optimization of the treatments included analysis of variance (ANOVA) and lack-of-fit

Response Surface Treatment Optimization
Considering the evidence that emerged from the linear models encompassing severity, 1-and 2-propanol/sodium hydroxide treatments exhibited differentiated effects on polyphenol recovery from OB. Therefore, response surface methodology was deployed to fully assess the effect of both solvents and provide alternative means of process modeling. For this purpose, temperature, T, and time, t, were used as the process variables. With such an approach, the effects of T and t on treatment performance could be further evaluated and possible cross (synergistic) functions could be revealed. Overall model appraisal and optimization of the treatments included analysis of variance (ANOVA) and lack-of-fit tests (Tables 3-5), taking into account the closeness of the measured and predicted values ( Table 6).   To deliver the equations representing the models derived, non-significant terms were excluded; hence, the models (equations) contained only statistically significant terms ( Table 4). The square correlations coefficients (R 2 ) gave a measure of the total variability around the mean, as this was predicted by the models: Y TP(2-Prop) = 11.72 + 1.47X 1 + 1.84X 2 (11) Both models were shown to have R 2 values equal to or higher than 0.97 ( Figures S1 and S2, Supplementary Material), and statistically significant p values for lack-of-fit (<0.05), considering a confidence interval of at least 95%. These findings dictated that the models derived from the response surface methodology exhibited outstanding fitting to the experimental (measured) data.
On the basis of the models, the three-dimensional plots were constructed to portray the influence of the process variables on the response (Y TP ), and to visualize the overall effect of the two solvents tested (Figure 3). For both the treatments with 1-and 2-propanol/sodium hydroxide, only t (X 1 ) and T (X 2 ) were significant, whereas no cases of significant cross or quadratic effects were observed. Therefore, the models described by Equations (10) and (11) represented linear correlations, where Y TP was directly proportional to both t and T. This finding was in absolute agreement with the models established based on severity, and clearly demonstrated that increases in both process variables within the ranges tested could lead to attaining higher polyphenol recovery.
With the aid of the desirability function ( Figures S1 and S2, Supplementary Material), the determination of the theoretically optimal values for both t and T, as well as the maximum predicted Y TP for each treatment, could be performed. For the treatment with 1-propanol/sodium hydroxide, maximum Y TP (17.15 ± 0.51 mg FAE g −1 DM) could be achieved at t = 300 min and T = 90 • C. In the case of 2-propanol/sodium hydroxide treatment, the maximum Y TP was determined to be 14.78 ± 1.11 mg FAE g −1 DM under the same set of conditions. On the grounds of the above data, it was clear that the organosolv treatment with 1-propanol/sodium hydroxide was almost 14% more efficient than that with 2-propanol/sodium hydroxide when identical regimes of process time and temperature were employed. The crucial role of sodium hydroxide as an alkali catalyst was revealed when treatments were carried out in its absence. The incorporation of sodium hydroxide at the optimal concentration resulted in a 132% increase in Y TP in the 1-propanol-based treatment, while the corresponding 2-propanol-based treatment gave a performance in-crease of 373% when sodium hydroxide was used (Figure 4). This outcome underlined the undisputed importance of the alkali catalyst in recovering OB polyphenols. disputed importance of the alkali catalyst in recovering OB polyphenols.
To have a critical assessment of the efficiency of the 1-propanol/sodium hydroxide treatment and to better illustrate the prospect of the process developed, the YTP levels attained were compared to those reported in the relevant literature. It has been shown that enzyme-assisted extraction of OB polyphenols using a mixture of carbohydrases could only afford a YTP of 4.80 mg gallic acid equivalents g −1 DM [36]. Integration of alkali-catalyzed hydrolysis did not boost polyphenol recovery to higher levels [36,37]. In another study on enzyme-catalyzed release of OB polyphenols, the yields determined were just under 2 mg gallic acid equivalents g −1 DM [18].  To have a critical assessment of the efficiency of the 1-propanol/sodium hydroxide treatment and to better illustrate the prospect of the process developed, the Y TP levels attained were compared to those reported in the relevant literature. It has been shown that enzyme-assisted extraction of OB polyphenols using a mixture of carbohydrases could only afford a Y TP of 4.80 mg gallic acid equivalents g −1 DM [36]. Integration of alkali-catalyzed hydrolysis did not boost polyphenol recovery to higher levels [36,37]. In another study on enzyme-catalyzed release of OB polyphenols, the yields determined were just under 2 mg gallic acid equivalents g −1 DM [18].
Conventional solvent extraction of various milling fractions of oat did not provide yields higher than 1 mg gallic acid equivalents g −1 DM [38], while extraction with supercritical fluids was of even lower efficacy, giving a maximum yield of as high as 0.89 mg gallic acid equivalents g −1 DM [39]. Comparable levels of 0.84 mg gallic acid equivalents g −1 DM were also achieved through solid-state fermentation of OB [40]. Therefore, to the best of the authors' knowledge, the Y TP achieved by the 1-propanol/sodium hydroxide treatment represents the highest level reported.
Conventional solvent extraction of various milling fractions of oat did not provide yields higher than 1 mg gallic acid equivalents g −1 DM [38], while extraction with supercritical fluids was of even lower efficacy, giving a maximum yield of as high as 0.89 mg gallic acid equivalents g −1 DM [39]. Comparable levels of 0.84 mg gallic acid equivalents g −1 DM were also achieved through solid-state fermentation of OB [40]. Therefore, to the best of the authors' knowledge, the YTP achieved by the 1-propanol/sodium hydroxide treatment represents the highest level reported.

Polyphenolic Profile and Antioxidant Properties
In addition to the total yield in polyphenols, the composition of the extracts generated with 1-and 2-propanol-based treatments was also examined to ascertain whether the different treatments affected the polyphenolic profile and the relative amounts of individual constituents. The extracts from both treatments, produced under optimal conditions, were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) to perform a tentative identification and quantification of the main compounds. The diagnostic fragments at m/z = 178.1 and 134.1, as well as the molecular ion at m/z = 193, were used to identify ferulic acid. Similarly, p-coumaric acid was recognized by the corresponding ions at m/z = 163 and 119.3 ( Figures S3, S4, Supplementary Material).
Irrespective of the alcohol used to perform the organosolv treatment, the role of sodium hydroxide as a catalyst was of paramount importance. Thus, while simple extraction with either 1-or 2-propanol afforded ferulic acid levels that did not exceed 5.3 µg g −1 DM, the use of sodium hydroxide boosted recoveries to 481.55 and 291.05, respectively (Table  7). Likewise, p-coumaric acid occurred at undetectable levels in extracts produced only with alcohols, but in the extracts obtained with sodium hydroxide catalysis, small amounts of p-coumaric acid accompanied ferulic acid. The organosolv treatment with 1propanol was proven to be far more effective than that with 2-propanol, giving a ferulic acid recovery that was 91% higher. The corresponding difference for p-coumaric acid was almost 51%. In total, the 1-propanol treatment yielded a 65% higher amount of polyphenol recovery.

Polyphenolic Profile and Antioxidant Properties
In addition to the total yield in polyphenols, the composition of the extracts generated with 1-and 2-propanol-based treatments was also examined to ascertain whether the different treatments affected the polyphenolic profile and the relative amounts of individual constituents. The extracts from both treatments, produced under optimal conditions, were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) to perform a tentative identification and quantification of the main compounds. The diagnostic fragments at m/z = 178.1 and 134.1, as well as the molecular ion at m/z = 193, were used to identify ferulic acid. Similarly, p-coumaric acid was recognized by the corresponding ions at m/z = 163 and 119.3 ( Figures S3 and S4, Supplementary Material).
Irrespective of the alcohol used to perform the organosolv treatment, the role of sodium hydroxide as a catalyst was of paramount importance. Thus, while simple extraction with either 1-or 2-propanol afforded ferulic acid levels that did not exceed 5.3 µg g −1 DM, the use of sodium hydroxide boosted recoveries to 481.55 and 291.05, respectively (Table 7). Likewise, p-coumaric acid occurred at undetectable levels in extracts produced only with alcohols, but in the extracts obtained with sodium hydroxide catalysis, small amounts of p-coumaric acid accompanied ferulic acid. The organosolv treatment with 1-propanol was proven to be far more effective than that with 2-propanol, giving a ferulic acid recovery that was 91% higher. The corresponding difference for p-coumaric acid was almost 51%. In total, the 1-propanol treatment yielded a 65% higher amount of polyphenol recovery.
Earlier investigations showed the ferulic acid and p-coumaric acid content in OB to vary from 54.7 to 123.3 µg g −1 DM and 14.3 to 20.2 µg g −1 DM, respectively [38]. Other authors reported much lower values of 19.3 to 43.3 µg g −1 DM and 6.3 to 9.8 µg g −1 DM, respectively [39], but levels of 5.7-49.4 and 247.2-1153.6 µg g −1 DM have also been found upon alkaline hydrolysis [19]. In another study, alkaline hydrolysis of OB gave a ferulic acid level of 1769.94 µg g −1 DM [37]. On the other hand, treatment with various hydrolytic enzymes was demonstrated to provide outstanding yields of 2578.6 and 55.6 µg g −1 DM, respectively [36]. In contrast, treatment of OB with cellulase afforded ferulic acid and p-coumaric acid yields that were up to 124.03 and 11.35 µg g −1 DM, respectively [18]. Table 7. Data illustrating the composition of major polyphenols and the antioxidant activity of the extracts produced with both 1-and 2-propanol treatments under optimized conditions. Data on extracts obtained from treatments without alkali catalysts are also given for comparison. Assignments: FA, ferulic acid; p-CouA, p-coumaric acid.

Sample
Polyphenolic The extracts produced by employing treatments with 1-propanol/sodium hydroxide and 2-propanol/sodium hydroxide under optimized conditions were tested for both antiradical activity (A AR ) and ferric-reducing power (P R ). Based on A AR , the extract obtained with 1-propanol/sodium hydroxide displayed outstanding performance, which greatly surpassed that of the extract generated with 2-propanol/sodium hydroxide ( Table 7). The results drawn from the P R test were not so pronounced, yet the 1-propanol/sodium hydroxide extract was proven once again to be the most active. Taking into account the polyphenolic composition of the extracts, as portrayed in Table 4, it would appear that the large difference in ferulic acid concentration between the two extracts was reflected in the antioxidant activity.
Such a result would not come as a surprise, considering that ferulic acid was shown to have a key role in the manifestation of antioxidant activity, being the major antioxidant contributor [26,41]. This argument was also concurred by hydrolysis studies, which illustrated that ferulic acid-enriched extracts produced with alkaline hydrolysis exhibited powerful antiradical activity [25].

Conclusions
The efficient recovery of oat bran (OB) polyphenols is a challenging prospect that involves deploying extraction technologies that provide a high recovery yield, purity, and stability of the target phytochemicals, as well as minimal environmental impact and the highest economic feasibility. This study demonstrated that the alkali-catalyzed organosolv treatment developed using 1-propanol may be a highly effective means of obtaining polyphenolic antioxidants from oat bran. Severity and response surface optimization modeling illustrated that the 1-propanol-based treatment was significantly more efficacious compared to 2-propanol, and equally severe. The extract produced under optimized conditions showed ferulic acid to be the major polyphenolic constituent, whereas p-coumaric acid occurred at significantly lower levels. On the basis of the data generated, it could be supported that alkali-catalyzed 1-propanol organosolv treatment could have an important perspective in the recovery of high value compounds from OB, such as ferulic acid. Based on the outcome of this investigation, it could be proposed that 1-propanol organosolv treatment of oat bran, integrated by alkaline catalysis, could be a very effective route for releasing and recovering pure ferulic acid, which may be used in the pharmaceutical, food, and cosmetics industries as a bioactive ingredient. Similar processes could be incorporated in biorefinery approaches to shape strategies that would contribute towards the establishment of wider circular economy policies.
Supplementary Materials: The following supporting information can be downloaded at: https://www. mdpi.com/article/10.3390/environments10070118/s1, Figure S1: Data associated with the response surface methodology, implanted to optimize the 1-propanol organosolv treatment. A, desirability function; B, correlation between actual (measured) and predicted values. Number designated with different color in the inset tables denote statistically different values; Figure S2: Data associated with the response surface methodology, implanted to optimize the 2-propanol organosolv treatment. A, desirability function; B, correlation between actual (measured) and predicted values. Number designated with different color in the inset tables denote statistically different values; Figure S3: A representative total ion chromatogram of 1-propanol extract obtained under optimized conditions: RT 6.16 min, p-coumaric acid; RT 7.14 min, ferulic acid); Figure S4: Fragmentation patterns of p-coumaric and ferulic acids.

Data Availability Statement:
The data is available from the corresponding authors.

Conflicts of Interest:
The authors declare no conflict of interest.