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Article

Cassia grandis L.f. Pods as a Source of High-Value-Added Biomolecules: Optimization of Extraction Conditions and Extract Rheology

by
Filipe M. M. Cordeiro
1,
Salomé G. Bedoya
1,
Daniel A. P. Santos
1,
Rebeca S. Santos
1,
Thomas V. M. Bacelar
1,
Filipe S. Buarque
2,*,
George Simonelli
1,
Ana C. M. Silva
1 and
Álvaro S. Lima
1,*
1
Programa de Pós-Graduação em Engenharia Química, Universidade Federal da Bahia, Rua Aristides Novis, Salvador 40210-630, Brazil
2
Biochemical Engineering Department, School of Chemistry, Federal University of Rio de Janeiro, Av. Athos da Silveira Ramos, 149. Ilha do Fundão, Rio de Janeiro 21941-909, Brazil
*
Authors to whom correspondence should be addressed.
Biomass 2025, 5(2), 24; https://doi.org/10.3390/biomass5020024
Submission received: 10 March 2025 / Revised: 11 April 2025 / Accepted: 21 April 2025 / Published: 25 April 2025
(This article belongs to the Topic Recovery and Use of Bioactive Materials and Biomass)

Abstract

:
High-value-added biomolecules such as phenolic compounds and flavonoids from secondary metabolism and macromolecules such as sugars are the main constituents of several plants. Thus, this work aims to optimize the extraction of these biomolecules present in the pods of Cassia grandis L.f. Initially, the effect of choline-based ionic liquids—ILs (choline chloride [Ch]Cl, dihydrogen citrate [Ch][DHC], and bitartrate [Ch][BIT]) as extracting agents for phenolic compounds and flavonoids was evaluated based on their efficiency and selectivity. Then, a 23 full factorial design with six central points was performed using the IL concentration, the solid–liquid ratio, and the temperature as independent variables. The extract obtained in the best condition was subjected to pervaporation, after which the concentrates and the crude extract were used to determine the physical properties (density, viscosity, and refractive index). The hydrophobic–hydrophilic balance of the extracting agent and the biomolecules are the extraction process’s driving force. The best extraction condition was formed by [Ch][DHC] at 15 wt%, with a solid–liquid ratio of 1:15, at 45 °C for 30 min, resulting in 153.71 ± 5.81 mg·g−1 of reducing sugars; 483.51 ± 13.10 mg·g−1 of total sugars; 11.79 ± 0.54 mg·g−1 of flavonoids; and 38.10 ± 2.90 mg·g−1 of total phenolic compounds. All the physical properties of the biomolecules are temperature-dependent; for density and refractive index, a linear correlation is observed, while for viscosity, the correlation is exponential. Increasing the temperature decreases all properties, and the extract concentration for 8× presents the highest values of density (1.283 g·cm−3), viscosity (9224 mPa·s), and refractive index (1.467).

1. Introduction

The genus Cassia comprises more than 500 species worldwide and belongs to the Fabaceae family. The tree, which belongs to the Fabaceae family, has a height between 15 and 30 m. Their leaves are alternately formed by 15 to 20 pairs of opposite leaflets, and the in-florescence has more than 15 pink flowers [1]. The fruits (pods) reach up to 75 cm long and are rigid. Inside, lobes are found which are separated by transverse partitions, which contain seeds surrounded by a dark red flesh with a thick consistency, pungent odor, sweet taste, and high water solubility [2]. Due to its wide geographical distribution, rich phytochemical profile, and abundance of bioactive compounds, this plant species has become a promising target for pharmacological, medical, and industrial use [3,4].
The occurrence of C. grandis L.f. across tropical regions of Asia (India and China), East Africa, and America (Brazil, Colombia, Mexico, Cuba, El Salvador, Nicaragua, Costa Rica, and Honduras) suggests a high adaptability to diverse ecosystems, which can influence the production of secondary metabolites. Preliminary studies have already identified the presence of cardiac glycosides, flavonoids (105 mg quercetin equivalent per gram), alkaloids (45 mg RU·g−1), phenolic compounds (169.73 mg gallic acid·g−1), stilbenes, and anthraquinones, substances recognized for their biological activity [5]. This chemical diversity gives C. grandis L.f. multiple therapeutic applications, including antiparasitic, antimicrobial, and anti-inflammatory properties and cytotoxicity against cancer cell lines. Additionally, the aqueous extract of its sweet-tasting pulp is traditionally used to treat respiratory diseases, cardiac glycosides, gastric disorders, and skin infections, allowing us to call it a multipurpose plant [6]. Despite these empirical uses, there remains a lack of systematic studies that explore its rheological properties and thermodynamic stability.
Syrups rich in phenolic compounds and sugars from C. grandis L.f. are produced through evaporation and are used in the above-mentioned health applications. To the best of our knowledge, there are no existing studies on the behavior of syrups. However, the rheological behavior of syrups affects sugar refining, provides information on the physical properties of foods, and helps us understand the heat transfer mechanism for their production [7]. The wide variety of high-value compounds resulting from secondary metabolism, combined with macromolecules such as sugars, and their diverse application in treating various diseases put pressure on the scientific community to develop methodologies for extracting these compounds. Nevertheless, research in this field is still quite scarce [8,9].
The literature has been extensive in presenting different techniques for the extraction of biomolecules from plant matrices, highlighting traditional methods such as solid–liquid extraction, liquid–liquid extraction, and maceration, or unconventional techniques such as supercritical extraction, ultrasound-assisted extraction, and microwaves [10]. Conventional methods are simple to perform. However, they are not selective; in most cases, they use volatile solvents and require long processing times [11]. On the other hand, unconventional techniques require more sophisticated and expensive equipment. New approaches, such as ionic liquids, have been proposed as alternative, more sustainable solvents [12].
Ionic liquids (ILs) are salts in the liquid stage below 100 °C formed by organic cations (ammonium, sulfonium, imidazolium, etc.) combined with organic or inorganic anions (chloride, tosylate, acetate, dicyanamide, etc.) through relatively stronger ionic interactions between symmetrically packed ions [13]. The various combination possibilities allow the designation of ionic liquids as designer solvents since their physicochemical properties can be tuned for specific tasks [14]. The interesting properties of ILs include non-volatility, low vapor pressure, non-flammability, wide electrochemical windows, high thermal stability, and good electrical conductivity [15]. Choline (2-hydroxyethyltrimethylammonium)-based ionic liquids belong to the quaternary ammonium salts associated with different anions. Choline salts are biodegradable, relatively inexpensive, and water-soluble [16,17]. According to Ventura et al. [18], choline is a complex B vitamin widely used as a food additive. It presents ecotoxicity to Vibrio fischeri marine bacteria, which is practically harmless and moderately toxic. The use of Aliivibrio fischeri as a bioindicator is advantageous due to its natural bioluminescence, which enables rapid, sensitive, and cost-effective screening of bioactive compounds. Inhibition or modulation of its luminescence by plant extracts may indicate toxic, antimicrobial, or antioxidant properties [8,19].
This study aims to optimize the extraction of high-value-added biomolecules, including phenolic compounds, flavonoids, and sugars (reducing and total) from Cassia grandis L.f. pods using choline-based ionic liquids as sustainable and selective extracting agents. The extraction process was optimized through a full factorial experimental design, evaluating the effects of ionic liquid concentration, solid–liquid ratio, and temperature on the yield of the target compounds. Additionally, the physical and rheological properties of the extracts and their concentrated forms were characterized to assess their potential applications.

2. Materials and Methods

2.1. Materials

C. grandis L.f. pods were collected from different trees (5 at least) in public gardens in Salvador (13°00′08.4″ S and 38°51′81.1″ W). These trees are not part of a collection; however, they were confirmed by a botanist through a plaque informing the genus and species Cassia grandis L.f. The biomass was transported to the Federal University of Bahia laboratory, where the dirt was removed, and the material was sanitized using 20 mg L−1 sodium hypochlorite. The pods were opened manually with a saw, and the internal contents (seeds, molasses-like material that coats the seeds and some of the woody material of the pod) were milled in a knife mill (Usifer, Farropilha, Brazil) and dried in a drying oven (Nova Ética, Vargem Grande Paulista, Brazil) at 55 °C. The particle size distribution profile was determined using a set of sieves of varying pore sizes (Figure S1 of the Supporting Information). Particle sizes between 0.25 and 0.85 mm were homogenized to perform the extraction processes.
The extraction was carried out using different ionic liquids based on choline, such as choline chloride—[Ch]Cl (≥98 wt%), choline bitartrate—[Ch][BIT] (≥98 wt%), and choline dihydrogen citrate—[Ch][DHC] (≥98 wt%), all purchased from Merck. The biomolecules determined in the aqueous extract of the extraction process were total phenolic compounds—TPCs (standard—gallic acid—≥98 wt%), flavonoids—FLA (standard—catechin—≥98 wt%), and total sugars (TSs) and reducing sugars (RSs) (standard—glucose—≥99 wt%), all purchased from Merck. All other chemicals were of analytical grade. The chemical structure of ionic liquids based on choline, as well as the standards, is shown in Figure 1.

2.2. Extraction Protocol

Initially, preliminary extractions were performed to determine the best extraction agent, using ionic liquids based on choline ([Ch]Cl, [Ch][BIT], and [Ch][DHC] under specified conditions such as 10 wt% ionic liquid, a solid–liquid ratio of 1:10, and a temperature of 35 °C, under constant magnetic agitation (Tecnal TE-085, Piracicaba, Brazil) at 100 rpm for 30 min. After choosing the extracting agent, optimization experiments for biomolecule extraction from C. grandis L.f. pods were performed using a full factorial design (FFD). The procedure consisted of 8 experimental runs and six central points. The variables were extracting agent concentration (X1, wt%), solid–liquid ratio (X2—g mL−1), and temperature (X3—°C). The values of the variables were established based on previous works of our research group [20,21].
The dried biomass was mixed with the extracting agent (test volume 10 mL) under experimental conditions, placed in 50 mL Erlenmeyer flasks, and conditioned in a bath (Quimis Q226M1, Diadema, Brazil) at the desired temperature for 30 min. The extract was centrifuged (Centrifuge Q 222T108, Quimis, Diadema, Brazil) at 2000 rpm for 10 min, and the biomass was discarded. Quantification of total phenolic compounds, flavonoids, and total and reducing sugars was immediately performed. The kinetics of the extraction process was determined under the optimized condition by varying the extraction time at intervals of 1, 5, 15, 30, 60, and 90 min.

2.3. Analytical Determination

Total phenolic compound (TPC) content was determined according to the method described by Swain and Hillis [20,22]. The absorbance was measured at 660 nm using a spectrophotometer (Shimadzu UV-3600 plus, Shimadzu, Kyoto, Japan). The TPC content was calculated using a standard curve (Equation (1)) based on gallic acid (0.25–0.50 mg mL−1), and the results were expressed as the mean and standard deviation of mg gallic acid equivalent (GAE) g−1 of biomass.
A b s = 0.0075 × C G A E 0.1242 , R 2 = 0.998
where Abs corresponds to the absorbance at 660 nm, CGAE is the concentration of gallic acid, and R2 is the correlation coefficient.
Flavonoid (FLA) content was determined by the methodology described by Sheng et al. [23], which used a standard curve (Equation (2)) prepared from an aqueous solution of catechin (5–60 mg L−1) as a standard to calculate the absorbance at 510 nm and, consequently, the FLA content. The results were expressed as the mean and standard deviation of mg catechin equivalents (CE) g−1 of biomass.
A b s = 0.0012 × C F L V 0.0039 , R 2 = 0.991
where Abs corresponds to the absorbance at 510 nm, CFLV is the concentration of catechin, and R2 is the correlation coefficient.
Reducing sugar (RS) was quantified using the 3,5-dinitrosalicylic acid (DNS) method proposed by Miller [24]. The absorbance was measured at 540 nm, and the results were calculated using a standard curve prepared with an aqueous solution of glucose (0.1–1.0 mg mL−1), and the RS concentration was expressed as the mean and standard deviation of mg glucose equivalents (mg g−1 of biomass). Total sugar (TS) was determined by acid hydrolysis (HCl 2.0 M) followed by neutralization with NaOH (2.0 M), and the glucose equivalent was determined as previously described.

2.4. Physical Properties

The C. grandis L.f. extracts (1x—brut extract) obtained under the best process conditions were roto-evaporated (Longen Scientific LSRE52CS-1, São Paulo, Brazil) under the conditions of 5 kPa at 45 °C until obtaining 2×, 4×, and 8× concentrated extracts and subjected to determinations of properties (density, viscosity, and refractive index).
Density (ρ) and dynamic viscosity ( η ) data were simultaneously analyzed using Anton Paar SVM 3000 (Anton Paar, Graz, Austria) equipment based on an adapted Couette measuring principle. A sample of 5 mL was injected into the equipment using a glass syringe. Experimental measurements were performed at a temperature between 20 and 55 °C under atmospheric pressure (101.3 kPa). The density and viscosity measurements had a combined uncertainty of ±5 × 10−5 g·cm−3 and 0.01· η , respectively. Refractive indices (nD) were also measured using an Anton Paar RXA170 (Anton Paar, Graz, Austria) refractometer at a temperature range from 20 to 55 °C under atmospheric pressure (101.3 kPa), with a combined uncertainty of ±2 × 10−5. The measurement was carried out in triplicate.

2.5. Statistical Analyses

All analytical determinations were conducted at least in triplicate. One-way ANOVA was used to analyze the experimental data, and the means were compared by the Tukey test using the SAS 9.0 software (SAS, Cary, NC, USA). Differences of p ≥ 0.05 were considered statistically significant.
Experimental data from a three-factor full factorial design (23) were analyzed using the software Statistica 7.0 (StatSoft Inc., Tulsa, OK, USA). The experimental condition of the central point was repeated six times to determine the pure error. Linear and interaction effects of the three variables considered on the response variables were calculated. The independent variables (factors) were the ionic liquid concentration (X1), solid–liquid ratio (X2), and temperature (X3). The dependent variables were total phenolic compounds (TPCs), flavonoids (FLA), total sugars (TSs), and reducing sugar (RSs) (Equation (3)). Pareto diagrams were constructed to evaluate the effects of the factors on each of the responses. Factors that were not statistically significant in the Pareto diagram were not considered in the regression model.
Y = β 0 + i = 1 k β i X i + i j = 1 k β i j X i X j
where Y is the response variable; Xi, and Xj are the independent variables; and k is the number of tested variables (k = 3). The regression coefficient is defined as β0 for intercept, βI for linear, and βij for interaction.

3. Results

3.1. Extraction of Biocompounds from C. grandis L.f.

Initially, the effect of the extracting agent was evaluated by comparing choline-based ionic liquids (at a concentration of 10 wt%, a solid–liquid ratio of 1:10, and a temperature of 35 °C for 30 min) with water as a conventional solvent (Figure 2 and Table S1 of Supporting Information).
Ionic liquids based on cholines have little influence on the extraction of macromolecules formed by carbohydrates (reducing and total sugars) when compared with values obtained for water. These molecules are known to be hydrophilic compounds and are quite soluble in water. Examples of sugars such as sucrose (disaccharide) and glucose (monosaccharide) have pronounced solubilities in water of 66.7% and 47.7% [25].
The decreasing order for the raw averages of sugar is water > [Ch]Cl (log Kow = −3.70) > [Ch][BIT] (log Kow = −1.43 > [Ch][DHC] (log Kow = −1.32), which corroborates the decrease in the logarithm of the octanol–water coefficient (log Kow); that is, more hydrophilic ionic liquids can remove a greater quantity of sugars. The statistical analysis of the averages using the Tukey test shows that the extraction using water, [Ch]Cl, and [Ch][Bit] as the extracting agent are statistically equal (p ≥ 0.05).
On the other hand, in extracting high-added value biomolecules such as flavonoids and total phenolic compounds, the order is reversed, and the most hydrophobic compounds are the best extracting agents. Phenolic compounds, including flavonoids, are formed by one or more aromatic rings linked to a hydroxyl group. Generally, low-molecular-weight phenolic compounds are water-soluble, while high-molecular-weight ones are insoluble [11]. This observation corroborates the extraction results; water-soluble compounds are extracted by water, while most water-insoluble ones are extracted by choline-based ionic liquids. The increase in hydrophobicity allowed greater extraction of phenolic compounds and flavonoids. Flavonoids and many phenolic compounds are commonly found in glycosylated forms, which means that they are chemically linked to one or more sugar fractions. This glycosylation significantly influences the polarity and solubility of the molecule. The glycosidic bond generally increases these compounds’ hydrophilicity (water solubility) compared to their aglycone (non-glycosylated) forms, which are more hydrophobic [26,27]. This amphiphilic nature (hydrophobic/hydrophilic balance) justifies the extraction results observed. The values differ statistically from each other (p ≥ 0.05). [Ch][DHC] was chosen to continue studies to determine the best conditions for extracting biomolecules from C. grandis L.f.
Table 1 depicts the experimental results of 14 runs for RS, TS, FLA, and TPC and the coded levels of the three independent process variables (X1—[Ch][DHC]; X2—solid–liquid ratio; and X3—temperature). This variable’s effect and interactions were studied using a 23 full factorial design with six repetitions and a central point. The values of independent variables were fixed according to conditions of extraction found in the preliminary experiments for different biomass carried out in our laboratory.
The biomolecule concentration ranged from 127.46 to 157.06 mg g−1 (RS); 401.44–504.40 mg g−1 (TS); 6.34–14.04 mg g−1 (FLA); and 12.95–38.10 mg g−1 (TPC). Overall, the increase in the values of the independent variables promotes the increase in the values of high-value-added biomolecules such as FLA and TPH. However, this observation is not valid for extracting RS and TS. This variation depends on the effects of positive or negative interaction between the variables aiming to extract biomolecules. The experimental results are shown in the response surface in Figure 3.
The increase in temperature during the extraction process enhances the permeability of cell membranes, thereby facilitating the diffusion of biomolecules into the extracting liquid. Furthermore, the solubility of these biomolecules is improved, which explains the higher extraction yields observed at elevated temperatures [28].
The significance of each variable was determined by F-test and p-values and is presented through the Pareto diagram (Figure 4) and ANOVA Tables S2–S5 of the Supporting Information). According to Atkinson and Donev [29], the independent variables are significant when they reflect higher F and smaller p-values. Therefore, each experimental set has different significant variables such as RS (CIL; T and the interaction between CiL vs. T); TS (T and interaction between RS/L vs. T); FLA (RR/L and T), and TPC (CIL, RS/L, T, and interaction between CIL vs. RS/L, and RS/L vs. T). The results suggested that the independent variables were well chosen for the study of the extraction process since they are statistically significant (p ≥ 0.05). The curvature of curves was significant (p ≤ 0.05) for the extraction of TS and FLA.
According to the 23 full factorial design, the data could be expressed by the predicted linear equation. The equations are presented as coded values as shown in Table 2. The predictive equations show good agreement due to the high regression coefficients (R2), which range from 0.9502 to 0.9915, which are statistically acceptable at the 95% significance level (p ≥ 0.05).
According to Patel et al. [30], predictive models permit the theoretical determination of the optimal conditions. However, the predate model found in our work did not present an optimal point. In this case, the alternative would be to search for the optimal path and then use a new experimental matrix. However, the limitations of the independent variables prevent this procedure. An increase in the concentration of [Ch][DHC] raises process costs due to the market value of this extracting agent; an increase in the solid–liquid ratio makes it difficult to concentrate the biomolecule due to the elimination of water, which will require the use of rotary evaporation. Finally, phenolic compounds are thermolabile, and temperatures above 50 °C result in the loss of biological activity of these biomolecules [20,31]. Therefore, determining the optimal path and using axial points are not feasible. The maximum concentrations were observed at 5 wt% [Ch][DHC], 1:5 (RS/L, g mL−1), and 45 °C for RS (157.06 mg g−1) and TS (504.40 mg g−1); 15 wt% [Ch][DHC], 1:15 (RS/L, g mL−1), and 25 °C for FLA (14.04 mg g−1); and 15 wt% [Ch][DHC], 1:5 (RS/L, g mL−1), and 45 °C for TPC (38.10 mg g−1).
Although the optimal conditions for each biomolecule are obtained separately, it is important to highlight that the extraction process is unique, and the concentrations of the biomolecules are determined together in each experiment. Therefore, the decision-making process is to choose the extraction conditions with the best performance. The extraction kinetics of high-value-added compounds was studied using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C (between 1 and 90 min (Figure 5 and Table S6 of Supporting Information). The extraction of high-value-added biomolecules reached a steady state within 30 min, except for TS, which peaked at 1 min. After this time, the values of each biomolecule’s concentration were statistically similar at 95% confidence.

3.2. Physical Properties of C. grandis L.f. Extract and Its Concentrates

After determining the best conditions for extracting biomolecules from C. grandis L.f. ([Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C for 30 min), the extract was concentrated by rotary evaporation under 5 kPa at 45 °C, in order to obtain different concentrated extracts (2×, 4×, and 8× concentrated). The physical properties were measured for each extract. Figure 6 Tables S7, S8, and S9 of Supporting Information) depicts the physical properties of the C. grandis L.f. extract and its concentrates obtained using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C.
Determining physical properties plays an important role in understanding the design and operation of processes. Therefore, the properties (density, viscosity, and refraction index) were measured between 20 and 60 °C and at 0.1 MPa due to the thermal sensitivity of the biomolecules in the extract.
The correlation between the density of the C. grandis L.f. extract and its concentrates with temperature is linear. Increasing temperature decreases the density from 1.283 g·cm−3 (20 °C for the 8× concentrated extract) to 1.252 g cm−3 (20 °C for the crude extract), which is due to the higher kinetic energies of biomolecules present in the extract [32]. The linear correlation is associated with Equation (4).
ρ = a b · T
where ρ is the viscosity (g cm−3), T is the temperature (K), and the indices a and b are the linear and angular coefficients.
The viscosity of the C. grandis L.f. extract and its concentrates follows a non-linear relationship. Increasing temperature leads to an exponential decrease in viscosity. Viscosity values range from 0.811 mPa·s (60 °C for the crude extract) to 9224.8 mPa·s (20 °C for the 8× concentrated extract). The behavior is expressed by Equation (5).
η = η 0 · e E η R T
where η is the viscosity (mPa·s), η0 is the initial viscosity (mPa·s), Eη (Kj.mol−1) is the activation energy, R is the ideal gas constant (J mol−1K−1), and T is the temperature (K).
The viscosity of the extracts is lower than that observed with pure water (100 mPa·s at 25 °C), except for the values found for the 8× concentrated extracts at all temperatures. Gabsi et al. [7] attributes this to the progressive disentanglement of long-chain molecule arrangements that helps overcome intermolecular resistance to flow.
The increase in temperature also leads to a decrease in the refractive index since this increase in temperature increases the internal energy of the solvent [33,34]. This observation was also reported by Tan et al. [35] in aqueous solutions with different solutes. The values varied from 1.3535 (60 °C for the crude extract) to 1.4672 (20 °C for the 8× concentrated extract). The trend follows Equation (6).
n D = c + d · T
where n D is the (dimensionless) refractive index, T is the temperature, and c and d are linear regression parameters that correspond to the linear and angular coefficients, respectively.
Table 3 presents the values of the adjustment parameters for the physical properties’ models. These models fit the experimental data well since the correlation coefficients are higher than 0.9861.

4. Conclusions

Experimental data for the high-value-added biomolecules (flavonoids and total phenolic compounds) and macromolecules (reducing and total sugars) have been reported, and 23 full factorial designs evaluated the best condition. The results showed that best extraction of reducing sugars (153.71 ± 5.81 mg·g−1), total sugars (483.51 ± 13.10 mg·g−1), flavonoids (11.79 ± 0.54 mg·g−1), and total phenolic compounds (38.10 ± 2.90 mg·g−1) was achieved using [Ch][DHC] at 15 wt%, with a solid–liquid ratio of 1:15, at 45 °C for 30 min. The interaction between the extracting agent and biomolecules expressed by their hydrophobic–hydrophilic characteristics plays an important role in the extraction process. The kinetics of extraction reached the steady state at 30 min. The crude and concentrate extracts present physical properties depending on the temperature. A linear correlation was observed for density and refraction index, while viscosity had an exponential dependence. Increasing the temperature implies decreased physical properties for all the studied extract concentrations. Thus, extracts from Cassia grandis L.f. pods are promising sources of bioactive compounds with potential applications in pharmaceutical, cosmetic, and food products, adding value to a widely available yet underexplored biomass. Moreover, the selection of this plant species is justified not only by its broad distribution across tropical regions of Latin America but also by its traditional use in folk medicine, where it is recognized for its laxative, antimicrobial, and antioxidant properties. Additionally, the pods were chosen because they represent biomass often discarded after the plant’s reproductive cycle, making them a sustainable and low-cost alternative for obtaining bioactive compounds.

Supplementary Materials

The following supporting information can be downloaded at https://www.mdpi.com/article/10.3390/biomass5020024/s1, Table S1. Effect of solvent on the extraction of biomolecules from C. grandis L using 10 wt% choline-based ionic liquid, with a 1:10 solid–liquid ratio, at 35 °C for 30 min.; Table S2. ANOVA table for linear models by reducing sugar extraction of C. grandis L.f.; Table S3. ANOVA table for linear models by total sugar extraction of C. grandis L.f.; Table S4. ANOVA table for linear models by flavonoid extraction of C. grandis L.f.; Table S5. ANOVA table for linear models by total phenolic compound extraction of C. grandis L.f.; Table S6. Kinetics of biomolecules (RS, TS, FLA, and TPC) extraction using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C; Table S7. Density (g cm−3) variation with temperature for the C. grandis L.f. extract and its concentrates obtained using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C for 30 min.; Table S8. Viscosity (mPa·s) variation with temperature for the C. grandis L.f. extract and its concentrates obtained using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C for 30 min.; Table S9. Refraction index variation with temperature for the C. grandis L.f. extract and its concentrates obtained using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C for 30 min.; Figure S1. Distribution particle size of C. grandis L.f. pods after milled in a knife mill and dried in an oven at 55 °C.

Author Contributions

Conceptualization: A.C.M.S. and Á.S.L.; methodology: F.M.M.C., S.G.B., and D.A.P.S.; formal analysis: F.S.B. and G.S.; data curation: R.S.S., T.V.M.B. and F.S.B.; writing—original draft preparation: F.M.M.C. and S.G.B.; supervision: G.S., A.C.M.S. and Á.S.L.; funding acquisition: A.C.M.S. and Á.S.L. All authors have read and agreed to the published version of the manuscript.

Funding

We thank the funding agency Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—CAPES (0001) and Conselho Nacional de Desenvolvimento Científico e Tecnológico—CNPq (0001) for the scholarship awarded to A.S. Lima (Grant number 306073/2023-4). Buarque, F.S. acknowledge the scholarship grant from FAPERJ: E-26/204.344/2021.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Data will be made available on reasonable request.

Conflicts of Interest

The authors declare no conflicts of interest.

Abbreviations

Choline chloride—[Ch]Cl; choline dihydrogen citrate—[Ch][DHC]; choline bitartrate—[Ch][BIT]; Cassia grandis L.f.—C. grandis L.f.; ionic liquids—ILs; total phenolic compounds—TPCs; flavonoids—FLA; total sugars—TSs; reducing sugars—RSs; full factorial design—FFD; Abs—absorbance; gallic acid equivalent—GAE; 3,5-dinitrosalicylic acid—DNS; logarithm of the octanol–water coefficient—log Kow.

References

  1. Lodha, S.R.; Joshi, S.V.; Vyas, B.A.; Upadhye, M.C.; Kirve, M.S.; Salunke, S.S.; Kadu, S.K.; Rogye, M.V. Assessment of the Antidiabetic Potential of Cassia grandis Using an in Vivo Model. J. Adv. Pharm. Technol. Res. 2010, 1, 330–333. [Google Scholar] [CrossRef] [PubMed]
  2. Fuentes, J.A.M.; Fernández, I.M.; Fernández, H.Z.; Sánchez, J.L.; Alemán, R.S.; Navarro-Alarcon, M.; Borrás-Linares, I.; Maldonado, S.A.S. Quantification of Bioactive Molecules, Minerals and Bromatological Analysis in Carao (Cassia grandis). J. Agric. Sci. 2020, 12, 88. [Google Scholar] [CrossRef]
  3. Luciméri, P.M.M.; Bárbara, A.R.; Márcia, V.d.S.; Maria, T.d.S.C.; Kêsia, X.F.R.d.S.; Túlio, D.d.S.; André, d.L.A.; Mônica, C.P.d.A.A.; Gardênia, C.G.M.; Teresinha, G.d.S.; et al. Antibacterial, Cytotoxic, and Schistosomicidal Activities of the Methanolic Extract from Cassia grandis L.f. (Fabaceae) Stem Bark and Its Fractions. J. Med. Plants Res. 2020, 14, 265–282. [Google Scholar] [CrossRef]
  4. Souza, A.A.; Ribeiro, K.A.; Seixas, J.R.P.C.; Silva Neto, J.C.; Santiago, M.G.P.F.; Aragão-Neto, A.C.; Lima-Ribeiro, M.H.M.; Borba, E.F.O.; Silva, T.G.; Kennedy, J.F.; et al. Effects Including Photobiomodulation of Galactomannan Gel from Cassia grandis Seeds in the Healing Process of Second-Degree Burns. Int. J. Biol. Macromol. 2023, 251, 126213. [Google Scholar] [CrossRef]
  5. Adimurti Kusumaningtyas Jenderal Achmad, V.; dewi Juliawaty, L.; Syah, Y.M.; Adimurti Kusumaningtyas, V.; Maolana Syah, Y.; Dewi Juliawaty, L. Two Stilbenes from Indonesian Cassia grandis and their antibacterial activities. Res. J. Chem. Environ. 2020, 24, 61–63. [Google Scholar]
  6. Harika, K.; Rao, B.G.; Ramadevi, D. Qualitative physicochemical, phytochemical analysis and quantitative estimation of total phenols, flavonoids and alkaloids of Cassia grandis. J. Glob. Trends Pharm. Sci. 2017, 8, 3860–3867. [Google Scholar]
  7. Gabsi, K.; Trigui, M.; Barrington, S.; Helal, A.N.; Taherian, A.R. Evaluation of Rheological Properties of Date Syrup. J. Food Eng. 2013, 117, 165–172. [Google Scholar] [CrossRef]
  8. Ferreira, K.; Cardoso, K.; Brandão-Costa, R.; Martins, J.T.; Botelho, C.; Neves, A.; Nascimento, T.; Batista, J.; Ferreira, É.; Damasceno, F.; et al. Physicochemical Properties of a Bioactive Polysaccharide Film from Cassia grandis with Immobilized Collagenase from Streptomyces Parvulus (DPUA/1573). Cosmetics 2024, 11, 86. [Google Scholar] [CrossRef]
  9. Medina, L.; Aleman, R.S.; Cedillos, R.; Aryana, K.; Olson, D.W.; Marcia, J.; Boeneke, C. Effects of Carao (Cassia grandis L.) on Physico-Chemical, Microbiological and Rheological Characteristics of Yogurt. LWT 2023, 183, 114891. [Google Scholar] [CrossRef]
  10. Osorio-Tobón, J.F. Recent Advances and Comparisons of Conventional and Alternative Extraction Techniques of Phenolic Compounds. J. Food Sci. Technol. 2020, 57, 4299–4315. [Google Scholar] [CrossRef]
  11. Alara, O.R.; Abdurahman, N.H.; Ukaegbu, C.I. Extraction of Phenolic Compounds: A Review. Curr. Res. Food Sci. 2021, 4, 200–214. [Google Scholar] [CrossRef]
  12. Ventura, S.P.M.; E Silva, F.A.; Quental, M.V.; Mondal, D.; Freire, M.G.; Coutinho, J.A.P. Ionic-Liquid-Mediated Extraction and Separation Processes for Bioactive Compounds: Past, Present, and Future Trends. Chem. Rev. 2017, 117, 6984–7052. [Google Scholar] [CrossRef]
  13. Kaur, G.; Kumar, H.; Singla, M. Diverse Applications of Ionic Liquids: A Comprehensive Review. J. Mol. Liq. 2022, 351, 118556. [Google Scholar] [CrossRef]
  14. Greer, A.J.; Jacquemin, J.; Hardacre, C. Industrial Applications of Ionic Liquids. Molecules 2020, 25, 5207. [Google Scholar] [CrossRef]
  15. Singh, S.K.; Savoy, A.W. Ionic Liquids Synthesis and Applications: An Overview. J. Mol. Liq. 2020, 297, 112038. [Google Scholar] [CrossRef]
  16. Gadilohar, B.L.; Shankarling, G.S. Choline Based Ionic Liquids and Their Applications in Organic Transformation. J. Mol. Liq. 2017, 227, 234–261. [Google Scholar] [CrossRef]
  17. Buarque, F.S.; Monteiro e Silva, S.A.; Ribeiro, B.D. Choline Chloride-Based Deep Eutectic Solvent as an Inhibitor of Metalloproteases (Collagenase and Elastase) in Cosmetic Formulation. 3 Biotech. 2023, 13, 219. [Google Scholar] [CrossRef]
  18. Ventura, S.P.M.; e Silva, F.A.; Gonçalves, A.M.M.; Pereira, J.L.; Gonçalves, F.; Coutinho, J.A.P. Ecotoxicity Analysis of Cholinium-Based Ionic Liquids to Vibrio Fischeri Marine Bacteria. Ecotoxicol. Environ. Saf. 2014, 102, 48–54. [Google Scholar] [CrossRef]
  19. Lafourcade Prada, A.; Achod, L.D.R.; Keita, H.; Carvalho, J.C.T.; de Souza, T.P.; Rodríguez Amado, J.R. Development, Pharmacological and Toxicological Evaluation of a New Tablet Formulation Based on Cassia grandis Fruit Extract. Sustain. Chem. Pharm. 2020, 16, 100244. [Google Scholar] [CrossRef]
  20. Inocêncio, E.S.; Buarque, F.S.; Ferreira, L.F.R.; Soares, C.M.F.; Lima, Á.S.; Souza, R.L.d. Exploring the Potential of Licuri (Syagrus Coronata) Using Sustainable Techniques and Solvents for Extracting Bioactive Compounds. Sustainability 2025, 17, 1507. [Google Scholar] [CrossRef]
  21. Lima, T.S.P.; Borges, M.M.; Buarque, F.S.; de Souza, R.L.; Soares, C.M.F.; Lima, Á.S. Purification of Vitamins from Tomatoes (Solanum Lycopersicum) Using Ethanolic Two-Phases Systems Based on Ionic Liquids and Polypropylene Glycol. Fluid Phase Equilibria 2022, 557, 113434. [Google Scholar] [CrossRef]
  22. Hillis, W.E.; Swain, T. The Phenolic Constituents of Prunus Domestica. II.—The Analysis of Tissues of the Victoria Plum Tree. J. Sci. Food Agric. 1959, 10, 135–144. [Google Scholar] [CrossRef]
  23. Sheng, Z.L.; Wan, P.F.; Dong, C.L.; Li, Y.H. Optimization of Total Flavonoids Content Extracted from Flos Populi Using Response Surface Methodology. Ind. Crops Prod. 2013, 43, 778–786. [Google Scholar] [CrossRef]
  24. Buarque, F.S.; Lima, T.S.P.; Carniel, A.; Ribeiro, B.D.; Coelho, M.A.Z.; Souza, R.L.; Soares, C.M.F.; Pereira, M.M.; Lima, Á.S. Hormones Concentration in an Aqueous Two-Phase System: Experimental and Computational Analysis. Chem. Eng. Technol. 2024, 47, 716–721. [Google Scholar] [CrossRef]
  25. Docoslis, A.; Giese, R.F.; Van Oss, C.J. Influence of the water–air interface on the apparent surface tension of aqueous solutions of hydrophilic solutes. Colloids Surf. B Biointerfaces 2000, 19, 147–162. [Google Scholar] [CrossRef]
  26. Ismail, B.B.; Yusuf, H.L.; Pu, Y.; Zhao, H.; Guo, M.; Liu, D. Ultrasound-Assisted Adsorption/Desorption for the Enrichment and Purification of Flavonoids from Baobab (Adansonia digitata) Fruit Pulp. Sonochemistry 2020, 65, 104980. [Google Scholar] [CrossRef]
  27. Mahindrakar, K.V.; Rathod, V.K. Ultrasonic Assisted Aqueous Extraction of Catechin and Gallic Acid from Syzygium Cumini Seed Kernel and Evaluation of Total Phenolic, Flavonoid Contents and Antioxidant Activity. Chem. Eng. Process.-Process Intensif. 2020, 149, 107841. [Google Scholar] [CrossRef]
  28. Mané, C.; Souquet, J.M.; Ollé, D.; Verriés, C.; Véran, F.; Mazerolles, G.; Cheynier, V.; Fulcrand, H. Optimization of Simultaneous Flavanol, Phenolic Acid, and Anthocyanin Extraction from Grapes Using an Experimental Design: Application to the Characterization of Champagne Grape Varieties. J. Agric. Food Chem. 2007, 55, 7224–7233. [Google Scholar] [CrossRef]
  29. Sousa, K.M.; Maciel, G.E.L.O.; Buarque, F.S.; Santos, A.J.; Marques, M.N.; Cavalcanti, E.B.; Soares, C.M.F.; Lima, Á.S. Novel Phase Diagrams of Aqueous Two-Phase Systems Based on Tetrahydrofuran + Carbohydrates + Water: Equilibrium Data and Partitioning Experiments. Fluid Phase Equilib. 2017, 433, 1–9. [Google Scholar] [CrossRef]
  30. Patel, S.B.; Attar, U.A.; Sakate, D.M.; Ghane, S.G. Efficient Extraction of Cucurbitacins from Diplocyclos palmatus (L.) C. Jeffrey: Optimization Using Response Surface Methodology, Extraction Methods and Study of Some Important Bioactivities. Sci. Rep. 2020, 10, 2109. [Google Scholar] [CrossRef]
  31. Xu, B.J.; Chang, S.K.C. A Comparative Study on Phenolic Profiles and Antioxidant Activities of Legumes as Affected by Extraction Solvents. J. Food Sci. 2007, 72, S159–S166. [Google Scholar] [CrossRef]
  32. Velho, P.; Sousa, E.; Macedo, E.A. Binary Aqueous Solutions of Choline Salts: Determination and Modelling of Liquid Density (298.15 or 313.15 K) and Vapour Pressure Osmometry (313.15 K). Fluid Phase Equilibria 2024, 587, 114197. [Google Scholar] [CrossRef]
  33. Rathnasamy, S.K.; sri Rajendran, D.; Balaraman, H.B.; Viswanathan, G. Functional Deep Eutectic Solvent-Based Chaotic Extraction of Phycobiliprotein Using Microwave-Assisted Liquid-Liquid Micro-Extraction from Spirulina (Arthrospira platensis) and Its Biological Activity Determination. Algal Res. 2019, 44, 101709. [Google Scholar] [CrossRef]
  34. Buarque, F.S.; Camêlo, L.C.A.; Soares, C.M.F.; Mattedi, S.; Ferreira, A.F.B.; Feitosa, F.X.; Souza, R.L.; de Sant’Ana, H.B.; Lima, Á.S. Binary Mixture of Double Protic Ionic Liquid: Density, Viscosity, Refractive Index, Surface Tension, and Derivative Properties. J. Chem. Eng. Data 2021, 66, 4309–4325. [Google Scholar] [CrossRef]
  35. Tan, C.Y.; Huang, Y.X. Dependence of Refractive Index on Concentration and Temperature in Electrolyte Solution, Polar Solution, Nonpolar Solution, and Protein Solution. J. Chem. Eng. Data 2015, 60, 2827–2833. [Google Scholar] [CrossRef]
Figure 1. Chemical structure of solvents and standard compounds.
Figure 1. Chemical structure of solvents and standard compounds.
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Figure 2. Effect of the extracting agent formed by choline-based ionic liquids (10 wt%) on the extraction of reducing sugars—RSs (A), total sugars—TSs (B), flavonoids—FLA (C), and total phenolic compounds—TPCs (D) with a solid–liquid ratio of 1:10 and at 35 °C for 30 min. In each graph, means followed by the same letters do not differ statistically by the Tukey test (p ≥ 0.05).
Figure 2. Effect of the extracting agent formed by choline-based ionic liquids (10 wt%) on the extraction of reducing sugars—RSs (A), total sugars—TSs (B), flavonoids—FLA (C), and total phenolic compounds—TPCs (D) with a solid–liquid ratio of 1:10 and at 35 °C for 30 min. In each graph, means followed by the same letters do not differ statistically by the Tukey test (p ≥ 0.05).
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Figure 3. Response surface of the extraction of RS (a), TS (b), FLA (c), and TFC (d) from C. grandis L.f. using [Ch][DHC] for 30 min.
Figure 3. Response surface of the extraction of RS (a), TS (b), FLA (c), and TFC (d) from C. grandis L.f. using [Ch][DHC] for 30 min.
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Figure 4. Pareto chart representation of the main effects. Influence of the variables—extracting agent concentration (X1, wt%), solid–liquid ratio (X2—g mL−1), and temperature (X3—°C)—using [Ch][DHC] as the extracting agent on the response variables RS (a), TS (b), FLA (c), and TPC (d).
Figure 4. Pareto chart representation of the main effects. Influence of the variables—extracting agent concentration (X1, wt%), solid–liquid ratio (X2—g mL−1), and temperature (X3—°C)—using [Ch][DHC] as the extracting agent on the response variables RS (a), TS (b), FLA (c), and TPC (d).
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Figure 5. Kinetics of biomolecule (RS, TS, FLA, and TPC) extraction using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C. (a) reducing sugars—RSs, (b) total sugars—TSs, (c) flavonoids—FLA, and (d) total phenolic compounds—TPCs. In each graph, means followed by the same letters do not differ statistically by the Tukey test (p ≥ 0.05).
Figure 5. Kinetics of biomolecule (RS, TS, FLA, and TPC) extraction using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C. (a) reducing sugars—RSs, (b) total sugars—TSs, (c) flavonoids—FLA, and (d) total phenolic compounds—TPCs. In each graph, means followed by the same letters do not differ statistically by the Tukey test (p ≥ 0.05).
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Figure 6. Experimental data of density (ρ), viscosity (η), and refraction index (nD) as a function of temperature for aqueous extracts () of C. grandis L.f. using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C for 30 min, and their concentrates (—2×, —4×, —8×).
Figure 6. Experimental data of density (ρ), viscosity (η), and refraction index (nD) as a function of temperature for aqueous extracts () of C. grandis L.f. using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C for 30 min, and their concentrates (—2×, —4×, —8×).
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Table 1. Extraction of reducing and total sugars, flavonoids, and total phenolic compounds from C. grandis L.f. by [Ch]Cl for 30 min using the full factorial design (FFD).
Table 1. Extraction of reducing and total sugars, flavonoids, and total phenolic compounds from C. grandis L.f. by [Ch]Cl for 30 min using the full factorial design (FFD).
RunX1 (CIL, wt%)X2 (RS/L, g mL−1)X3 (T, °C)RS (mg g−1)TS (mg g−1)FLA (mg g−1)TPC (mg g−1)
1−1 (5)−1 (1:5)−1 (25)149.55401.446.9612.95
2+1 (15)−1 (1:5)−1 (25)127.46427.7311.6722.57
3−1 (5)+1 (1:15)−1 (25)151.94447.447.1815.40
4+1(15)+1 (1:15)−1 (25)136.69459.4414.0430.93
5−1 (5)−1 (1:5)+1 (45)157.06504.408.7416.48
6+1 (15)−1 (1:5)+1 (45)146.25499.8413.5428.22
7−1 (5)+1 (1:15)+1 (45)153.34457.4910.8924.05
8+1 (15)+1 (1:15)+1 (45)153.71483.5115.7938.10
90 (10)0 (1:10)0 (35)142.57481.266.6722.80
100 (10)0 (1:10)0 (35)139.78490.168.0023.42
110 (10)0 (1:10)0 (35)143.35479.046.3422.99
120 (10)0 (1:10)0 (35)147.73491.078.3123.57
130 (10)0 (1:10)0 (35)143.29464.446.8223.22
140 (10)0 (1:10)0 (35)145.51473.317.1523.34
Table 2. Predicted equations and regression coefficients for the dependent variables obtained from the 23 full factorial design (coded value).
Table 2. Predicted equations and regression coefficients for the dependent variables obtained from the 23 full factorial design (coded value).
VariableEquationR2
Reducing Sugar—RSRS = 147.00 − 5.97X1 + 1.92X2 + 5.59X3 + 2.25X1X2 + 3.36X1X3 − 0.99X2X30.9502
Total Sugar—TSTS = 460.16 + 7.47X1 + 1.81X2 + 26.15X3 + 2.04X1X2 − 2.10X1X3 − 17.62X2X30.9276
Flavonoids—FLAFLA = 11.10 + 2.66X1 + 0.87X2 + 1.14X3 + 0.28X1X2 − 0.23X1X3 + 0.23X2X30.9726
Total Phenolic Compounds—TPCsTPC = 23.59 + 6.37X1 + 3.53X2 + 3.13X3 + 1.03X1X2 + 0.08X1X3 + 0.83X2X30.9915
Table 3. Regression parameters for the physical properties of the C. grandis L.f. extracts and their concentrates obtained using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C for 30 min.
Table 3. Regression parameters for the physical properties of the C. grandis L.f. extracts and their concentrates obtained using [Ch][DHC] 15 wt%, solid–liquid ratio 1:15 at 45 °C for 30 min.
Density ρ
SampleabR2
Crude1.230 ± 0.0086.0 × 10−4 ± 2.0 × 10−50.9863
Concentrated 2×1.283 ± 0.0056.0 × 10−4 ± 1.5 × 10−50.9957
Concentrated 4×1.386 ± 0.0037.0 × 10−4 ± 1.0 × 10−50.9983
Concentrated 8×1.510 ± 0.0038.0 × 10−4 ± 1.1 × 10−50.9987
Viscosity—η
Sampleη0EηR2
Crude1.1 × 10−4 ± 8.8 × 10−51.8 × 105 ± 2.1 × 1020.9991
Concentrated 2×9.0 × 10−4 ± 1.0 × 10−42.0 × 105 ± 3.9 × 1020.9977
Concentrated 4×1.0 × 10−4 ± 2.5 × 10−53.1 × 105 ± 4.8 × 1020.9987
Concentrated 8×7.6 × 10−7 ± 8.8 × 10−85.6 × 105 ± 2.8 × 1020.9999
Refraction Index—nD
SamplecdR2
Crude1.394 ± 0.0021.0 × 10−4 ± 5.4 × 10−60.9861
Concentrated 2×1.451 ± 0.0012.0 × 10−4 ± 3.9 × 10−60.9980
Concentrated 4×1.498 ± 0.0023.0 × 10−4 ± 8.0 × 10−60.9933
Concentrated 8×1.548 ± 0.0013.0 × 10−4 ± 1.5 × 10−60.9998
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Cordeiro, F.M.M.; Bedoya, S.G.; Santos, D.A.P.; Santos, R.S.; Bacelar, T.V.M.; Buarque, F.S.; Simonelli, G.; Silva, A.C.M.; Lima, Á.S. Cassia grandis L.f. Pods as a Source of High-Value-Added Biomolecules: Optimization of Extraction Conditions and Extract Rheology. Biomass 2025, 5, 24. https://doi.org/10.3390/biomass5020024

AMA Style

Cordeiro FMM, Bedoya SG, Santos DAP, Santos RS, Bacelar TVM, Buarque FS, Simonelli G, Silva ACM, Lima ÁS. Cassia grandis L.f. Pods as a Source of High-Value-Added Biomolecules: Optimization of Extraction Conditions and Extract Rheology. Biomass. 2025; 5(2):24. https://doi.org/10.3390/biomass5020024

Chicago/Turabian Style

Cordeiro, Filipe M. M., Salomé G. Bedoya, Daniel A. P. Santos, Rebeca S. Santos, Thomas V. M. Bacelar, Filipe S. Buarque, George Simonelli, Ana C. M. Silva, and Álvaro S. Lima. 2025. "Cassia grandis L.f. Pods as a Source of High-Value-Added Biomolecules: Optimization of Extraction Conditions and Extract Rheology" Biomass 5, no. 2: 24. https://doi.org/10.3390/biomass5020024

APA Style

Cordeiro, F. M. M., Bedoya, S. G., Santos, D. A. P., Santos, R. S., Bacelar, T. V. M., Buarque, F. S., Simonelli, G., Silva, A. C. M., & Lima, Á. S. (2025). Cassia grandis L.f. Pods as a Source of High-Value-Added Biomolecules: Optimization of Extraction Conditions and Extract Rheology. Biomass, 5(2), 24. https://doi.org/10.3390/biomass5020024

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