
currently brought up many criticisms (Palmade-Le Dantec & Picot, 2010; Baudelaire, 2013) : Worries have been reported in relation to the dangerousness of many solvents used for extraction (e.g., dichloromethane, acetyl acetate, and toluene), their impact on atmosphere, environment, and human health, the costs associated with the treatment of generated toxic wastes, and the impact on extracts quality and safety (Palmade-Le Dantec & Picot, 2010) .

Recently, alternation of drying and grinding process (ADG) and CDS extraction, that is, grinding and controlled differential screening, consisting in combinations of drying, grinding, and controlled sieving processes, has received increasing attention due to the raising desire to develop ecological extraction technologies of natural and active ingredients (Baudelaire, 2013; Becker et al., 2017 Becker et al., ,2016 Brewer, Kubola, Siriamornpun, Herald, & Shi, 2014; Karam, Petit, Zimmer, Djantou, & Scher, 2016; Li et al., 2015; Lucas-González, Viuda-Martos, Pérez-Álvarez, & Fernández-López, 2017; Zaiter, Becker, Baudelaire, & Dicko, 2018; Zaiter, Becker, Petit, et al., 2016) . Indeed, the competitive advantage of plant powders in comparison with conventional bioactive molecules obtained from plants by solvent extraction mainly resides in the preservation of bioactive ingredients, and more especially of their bioactivity, for human interest. Becker et al. (2017) reported that the sieving process separates plant powders by granulometric differentiation through sieves of decreasing mesh, leading to selective distribution of bioactive molecules in the different granulometric fractions. Moreover, these authors explained that various parts of the same plant, more or less hard, more or less fibrous, are thus more or less difficult to crush, and thus, different plant parts may likely lead to particle fractions presenting different chemical compositions and/or structures. The sieving process leads to the separation of particles according to their sizes, thus leading to different physicochemical properties of resulting particle size classes (GuerreroBeltrán, Jiménez-Munguía, Welti-Chanes, & Barbosa-Cánovas, 2009; Toth et al., 2005; Wang & Flores, 2000) . The fine particles issued from the combination of drying, grinding, and sieving processes enable a better release of bioactive substances owing to their high specific surface area (Rosa, Barron, Gaiani, Dufour, & Micard, 2013; Zhao et al., 2018) . Several recent studies have reported the link between particle size range of plant powders, bioactive ingredient contents, physicochemical properties, and functionalities (Becker et al., 2017; Sharma, Kadam, Chadha, Wilson, & Gupta, 2013; . With the increasing quantity and variety of powdered ingredients used in industry (Sharma et al., 2013) , production of plant powders constitutes an important step for their valorization in various industrial sectors. Plant processing into powders not only allows the production of functionally adequate products, but also ensures their preservation during an extended shelf life, while supplying bioactive molecules under an adapted form for market of food supplements.

Africa is home to some of the most important species-rich biodiversity regions in the world (Linder, 2014) . In several African countries, natural products constitute an important part of human diet and are also an excellent source of bioactive molecules. First plant of medicinal interest of the current study, Hibiscus sabdariffa, is an edible plant, for which previous studies on alcoholic and aqueous extracts from its calyx reported anti-inflammatory, antioxidant, hypolipidemic (Chung, Kong, Choi, & Kong, 2017; Medina-Carrillo et al., 2015) , and anti-hypertensive effects (Abubakar, Ukwuani, & Mande, 2015) , owing to its wealth of bioactive compounds (polyphenols, flavonoids, saponins, tannins, alkaloids, etc.) . Then, it has also been showed that the aqueous, alcoholic, and hydroalcoholic extracts of the fruits of Dichrostachys glomerata, which is the second plant of medicinal interest of the study, exhibit antioxidant (Kuate, Etoundi, Soukontoua, Ngondi, & Oben, 2010) , anti-hypertensive, hypoglycemic (Fankam, Kuete, Voukeng, Kuiate, & Pages, 2011), anti-inflammatory, and anti-hyperlipidemic activities (Kuate et al., 2013) ; these numerous biological activities were attributed to their contents in a broad range of bioactive molecules such as alkaloids, saponins, tannins, mucilage, glucocapparins, and sterols. Likewise, the seed extracts of the third plant of medicinal interest of the current study, Boscia senegalensis, are rich in saponins, tannins, anthraquinone, alkaloids, and flavonoids; it has been shown that they have anti-inflammatory, anti-hyperglycemic, and antioxidant properties (Dongmo, Dogmo, & Njintang, 2017) .

This work aims at evaluating the effect of grinding and sieving on chemical composition in macronutrients, phytochemical contents, and antioxidant activity of powders of Boscia senegalensis seeds, Dichrostachys glomerata fruits, and Hibiscus sabdariffa calyces. It intended to find the granulometric fractions presenting the highest antioxidant activity for their use in nutraceutical formulations.
| MATERIAL S AND ME THODS

| Plant material
Samples were collected in May 2015 from different localities: Sundried red calyces of Hibiscus sabdariffa were purchased from local markets in the Adamawa region of Cameroon, sun-dried fruits of Dichrostachys glomerata were bought in a market located in Yaounde (Central Region, Cameroon), and dry fruits of Boscia senegalensis were purchased from rural farmers in a local market of Oum-Madjer (Batha state, Chad). The fruits of Dichrostachys glomerata and Hibiscus sabdariffa calyces were manually separated from inorganic materials, dirt, and dust particles before grinding. On other hand, the fruits of Boscia senegalensis were manually decorticated and the obtained seeds were taken into grinding.
| Chemicals
Ascorbic acid, aluminum chloride, gallic acid, sodium hydroxide, Folin-Ciocalteu reagent, sodium carbonate, methanol, ethanol, potassium ferricyanide (K 3 Fe(CN) 6 ), potassium ferrocyanide (K 4 Fe(CN) 6 ), rutin, catechin, vanillin, sulfuric acid, 6-hydroxy-2,5,7,8-tetramethylchlorman-2-carboxylic acid (trolox), α-tocopherol, butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis 3-ethylbenzothiazoline-6-sulfonic acid (ABTS), trichloroacetic acid, and anhydrous ferric chloride were purchased from Sigma-Aldrich (SaintLouis, USA).
| Methods

| Plant grinding
An electric ultra-centrifugal mill ZM 200 (Retsch, Haan, Germany) supplied with a sieve with 24-tooth rotor of 99 mm diameter and sieve drilled with 1 mm trapezoid holes; was used to grind dry plant parts by impact and shearing size reduction principles. and local temperature increase in plant parts during grinding, as the latter is known to be enhanced at high rotor speed and lead to bioactive compounds alteration Zaiter, Becker, Petit, et al., 2016) .
| Plant powder sieving
The sieving process is based on the separation of particles from a granular material by making them pass through several sieves of decreasing mesh size (315, 212, and 180 µm in the current study).
| Plant powder sieving
Ground plants were sieved with the Analysette 3 Spartan apparatus (Fritsch, Idar-Oberstein, Germany), operating by vertical vibration.
| Plant powder sieving
Basically, 100 g ground plant sample was sieved in permanent vibratory pclark@example.net mm amplitude for 10 min. The fraction of the powder retained on each sieve was recovered and weighed for the calculation of the mass fraction of each granulometric class. A sample of unsieved plant powder was kept for comparison purposes.
| Plant powder sieving
Resulting plant powders were then put in sealed polyethylene plastic bags and stored at 10°C until analyses.
| Particle size distribution
Particle size distribution of each the powder classes and unsieved plant powders was determined by laser diffraction (Mastersizer 3000, Malvern Instruments France, Orsay, France) at ambient temperature, supplied with the Aero S dry dispersion unit that uses high-pressure air to disperse particles. The obscuration level was set lower than 2% to avoid multiple scattering by adjusting dispersion pclark@example.net bar air pressure: 30% air pressure, 30% feed rate, 2.5 mm hopper length for powder samples of Dichrostachys glomerata and Hibiscus sabdariffa; 70% air pressure, 70% feed rate, 2.5 mm hopper length for Boscia senegalensis powder samples.
| Particle size distribution
Particle sizes were expressed in terms of equivalent spherical diameters in volume. Characteristic sizes of the particle size distribution, D10, D50, and D90, were measured, where DX means that X % of the volume of particles has a diameter inferior to DX. Also, the span, a common parameter related to the width of particle size distribution, was evaluated as follows:
| Macronutrient composition
Moisture content was determined by drying 5 g plant powder in an oven at 103°C during 24 hr until reaching constant weight, according to AOAC method 925.10 (AOAC, 1990) . Total ash content was determined by incinerating from 3 to 5 g plant powder sample in a furnace at 550°C for 6 hr, then weighing the residue after cooling to room temperature in a desiccator, following the AOAC method 920.87 (AOAC, 1990) . The crude nitrogen content was obtained using the Kjeldahl method by the AOAC method 991.20 (AOAC, 1990) , and the protein content was deduced from it with a conversion factor of 6.25 (AACC, 1999) . In the procedure of fat content determination [34] , 8 g of powder samples was added to 30 ml of chloroform/methanol (2/1 [v/v] ) and mixed for 20 min.
| Macronutrient composition
The mixture was filtered under dinitrogen, and the residue was re-extracted in 20 ml of the same solvent and filtered. The extracts were then mixed and allowed to separate after the addition of 0.02 ml of NaCl pclark@example.net g/100 ml. The fat was recovered by rotary evaporation at 40°C under liquid nitrogen (Rotavapor R-144 Büchi, Flawil, Switzerland), and the fat contents were calculated by weight difference. Powder samples were hydrolyzed in 1.5 N sulfuric acid, and the available sugars were quantified by the phenol method (Dubois, Gilles, Hamilton, Ribers, & Smith, 1956 ).
| Determination of Phytochemical composition
Extraction of phenolic compounds 2 g plant powder was mixed with 20 ml methanol/water (70/30 [v/v] ).
| Determination of Phytochemical composition
The mixture was subjected to maceration by stirring at 300 rpm for 24 hr at room temperature (18 ± 2°C) and then filtered through with a Whatman filter paper (GE Healthcare companies, China) of 2-3 µm pore size. Thereafter, the supernatant was brought to 15 ml by addition of extraction solvent and stored at 4°C until analysis. This choice of extraction procedure (maceration under agitation) allowed shortening the process of extraction, thus minimizing the contact time of plant sample with solvent and better preserving the bioactivity of extracted molecules. Besides, the extraction at ambient temperature was also a compromise between extraction efficiency and limitation of thermal alteration of extracted biomolecules (Ćujíc et al., 2015) .
Determination of total phenolic content
Total phenolic content of the hydromethanolic extract of plant powders was determined using the Folin-Ciocalteu method (Wafa, Amadou, Larbi, & Héla, 2014) . Briefly, 20 µl of filtered extracts was diluted with 2,980 µl distilled water. Then, 500 µl of 10% (v/v) Folin-Ciocalteu reagent was added and the mixture was mixed. After 3 min, 400 µl of saturated solution of sodium
Determination of total phenolic content
carbonate Na 2 CO 3 (20% [w/v]) was added. After stirring, the tubes were placed at room temperature for 60 min and absorbance was measured at 760 nm using a spectrophotometer (Shimadzu UV-VIS 1605, Tokyo, Japan). A calibration curve (R 2 = 0.99) was prepared using standard solutions of gallic acid (40, 80, 120, 160, 200, 240 , and 280 g/L). Thus, total phenolic content was expressed as milligram gallic acid equivalents per gram dry weight (mg GAE/g DW) of plant powder.
Determination of flavonoid content
The total content in flavonoid compounds of the different samples was measured following the method of Dewanto, Wu, Adom, and Liu (2002) . In brief, 0.1 ml diluted and filtered hydromethanolic extract was added to 2.4 ml of distilled water and 0.15 ml of 5% Na 2 NO 2 (w/v). After 6 min, 0.3 ml of 10% aluminum chloride (AlCl 3 ·6H 2 O) (w/v) was added. The mixture was kept at room temperature for 5 min, and 1 ml NaOH (1 M) was added. Absorbance was then meas- 
Determination of condensed tannins content
The content in condensed tannins of plant powders was evaluated using concentrated sulfuric acid to depolymerize tannins, which allowed them to react with vanillin and produce red anthocyanidins that were detected at 510 nm by UV/visible spectrophotometry (Sun, Ricardo-Da-Silva, & Spranger, 2008) . 0.05 µl of diluted and filtered hydromethanolic extract was mixed with 3 ml of 4% vanillin (w/v), and 1.5 ml concentrated sulfuric acid was added. The mixtures were stirred and then kept at room temperature for 30 min.
Determination of condensed tannins content
The absorbance was measured at 500 nm by UV/visible spectrophotometry against the hydromethanolic solvent (70/30) as blank. A calibration curve (R 2 = 0.99) was prepared using standard solutions of catechin (100, 200, 300, 400, 500, and 600 µg/ml). The content in condensed tannins was expressed as milligrams catechin equivalents per gram dry weight (mg CE/g DW).
| Determination of antioxidant activity
DPPH radical scavenging activity assay
| Determination of antioxidant activity
Antioxidant activity was first evaluated by the DPPH method (Zhang & Yasumori, 2004) , in which the electron donating capacity of the extracts was measured by whitening of the purple-colored
| Determination of antioxidant activity
This assay is based on the ability of antioxidants to scavenge the DPPH cation radical. Briefly, 2 ml of 0.1 mM DPPH methanolic solution was added to 0.5 ml hydromethanolic extract of plant sample at different concentrations (0.025, 0.05, 0.1, 0.5, 1, 5, 10, 100 mg/ ml). The mixture was thoroughly stirred and incubated in the dark for 1 hr at room temperature. After that, absorbance of the mixture was measured at 517 nm by UV/visible spectrophotometry. Lower absorbance of the reaction mixture indicated higher free radical scavenging activity. The scavenging activity was estimated based on the percentage of scavenged DPPH radical using the following:
| Determination of antioxidant activity
The antioxidant activity was expressed as the concentration required to cause 50% DPPH scavenging, referred as IC 50
| Determination of antioxidant activity
(µg/ml). Ascorbic acid which was used as reference standard at the same concentrations as plant extracts showed an IC 50 value of 14.33 ± 0.58 µg/ml.
ABTS cation radical scavenging activity assay
The ABTS radical cation scavenging activity was measured according to the method described by Re et al. (1999) with slight modifications. In brief, ABTS solution was generated as follows: 6.62 mg of potassium persulfate and 38.4 mg of ABTS reagent were weighed in a glass beaker, 10 ml distilled water was added, and then the mixture 
| Statistical analysis
Results of chemical composition, phytochemical contents, and antioxidant activity were each subjected to analysis of variance (ANOVA) to determine the occurrence of statistically significant differences among them (p < 0.05). Duncan multiple range test was used to determine the degree of significance of the difference be- glomerata fruits, and Hibiscus sabdariffa calyces are displayed in Figure 1 . In addition, the results of sieve fractionation and particle size characteristics of powder fractions ( 
| Biochemical composition
The proximate composition of fractions and unsieved powders is shown in Table 2 . The moisture content of plants was ranging from 4.66% for Boscia senegalensis to 8.22% for Hibiscus sabdariffa, which makes them more stable during storage and packaging. Indeed, higher moisture content (generally over 10%) induces the development of microorganisms and product deterioration (Kaur, Kaushal, & Sandhu, 2011) . No clear trend about the influence of particle size can be drawn from moisture results. A similar observation was reported by Becker et al. (2016) : According to these authors, the heating effect of grinding, more pronounced for smaller particles and leading to moisture content decrease by heat-induced evaporation, could be compensated by the higher specific surface of small particles, facilitating the absorption of surrounding air humidity.
| Biochemical composition
The results also showed that the chemical composition depends on particle size. A significant difference (p < 0.05) was observed between the protein content of the different fractions of Boscia senegalensis: Proteins were more concentrated in finer particles (<180 µm).
| Biochemical composition
On the contrary, protein contents were similar or at least very close for the different powder fractions of the two other plants.
| Biochemical composition
It is often observed that the smallest particles are richer in minerals, because fibrous plant parts, containing fewer minerals, are harder to grind, resulting in larger particles (Becker et al., 2017 . Similar observations were made on Eucalyptus grandis powders. In this respect, fractions with smaller particle size were found to possess higher ash content and smaller fiber (hemicellulose and cellulose) content (Flávia, Edwil, & Fernando, 2016 Note. For each plant, means ± SD followed by the same superscripted letter were not significantly different (p < 0.05). It can be noted that the content in phenolic components considerably varied between unsieved plant powders and powder fractions of the same plant. The <180 µm granulometric fraction of Boscia senegalensis presented a higher total phenolic content. This can be related to the highest total protein and lipid contents of this particle size fraction. Indeed, large particles, richer in carbohydrates and probably in fibers (Oghbaei & Prakash, 2016) , are expected to contain less bioactive compounds . of Hibiscus sabdariffa (Hakimeh, Shahryar, & Majid, 2016) and some other plants such as Halimium halimifolium (Rebaya et al., 2014) and
| Polyphenol composition
Thymus kotschyanus (Baharfar, Azimi, & Mohseni, 2015) . Strong amounts of bioactive contents are responsible for their more significant antioxidant activity.
| In vitro antioxidant activity

| DPPH radical scavenging activity
The results of DPPH radical scavenging activity presented in Table 4 were expressed in terms of concentration required to cause 50% DPPH inhibition (IC 50 ). The lower the IC50 value, the more important the antioxidant activity of the studied plant powder. From the DPPH assay, the maximal antioxidant activity (lowest IC respectively. This suggests that phenolic compounds significantly contributed to the antioxidant activity of investigated plant powders. This is consistent with the fact that antioxidant activity of plant products is generally attributed to radical scavenging activity of phenolic compounds such as flavonoids, polyphenols, and tannins (Rahman & Moon, 2007) . The antioxidant activity of phenolic compounds is mainly due to their redox properties, which can play an important role in adsorbing and neutralizing free radicals, quenching singlet and triplet oxygen, or decomposing peroxides (Hasan et al., 2008) . The high contents in bioactive compounds of studied plant powders are likely responsible for their significant antioxidant activity. 
| ABTS radical scavenging activity

| Principal Component Analysis of physicochemical and phytochemical properties of studied plant powders
In order to compare all powder fractions of the three plants on the basis of their physicochemical, phytochemical, and antioxidant properties, measured parameters were organized into principal components by the principal component analysis (PCA) approach. The principal components 1 (PC1) and 2 (PC2), respectively, explained 61.9% and 28.3% variations among powder properties, allowing PCA to explain a total variation of 90.2%. Figure 2 gives the representation of analyzed powder parameters on the PC1 and PC2 axes. All powder properties highly contributed to the PC1xPC2 axes with the exception of D50 which contributed well to the PC3. The variables in PC1 that contribute much to discriminate the powders were DPPH and ABTS IC50, followed by fat, total phenol, flavonoid, and protein contents. Highly significant correlations (r > 0.95; p < 0.001) were found between phenols, flavonoids, and tannins, as attested by their proximity in the correlation circle of the PCA (Figure 2) . Similarly, the concentration for 50% scavenging (IC50) of DPPH and ABTS (r = 0.98; p < 0.001) was almost superimposed in the correlation circle, as well as the fat and protein contents (r = 0.91; p < 0.001). We found negative linear correlation although lower (DPPH, r = −0.77; ABTS, r = −075; all p < 0.01), between the IC50 scavenging activities and the total phenol content.
| Principal Component Analysis of physicochemical and phytochemical properties of studied plant powders
In addition, the D50 was significantly correlated negatively with protein (r = 0.61; p < 0.01) and fat (r = −0.76; p < 0.001) meaning that as the particle size increased, the protein and fat contents decreased.
| Principal Component Analysis of physicochemical and phytochemical properties of studied plant powders
We can also observe in Figure 2 the opposition between the total carbohydrate and the phenol contents. In this respect, antioxidant activities of the powders were increased by the total phenol, flavonoid, TA B L E 3 Contents in total phenols, flavonoids, and condensed tannins in hydromethanolic extracts of the different granulometric fractions and unsieved powders of Boscia senegalensis seeds, Dichrostachys glomerata fruits, and Hibiscus sabdariffa calyces and tannin contents while the protein and fat contents tended to alter them. These observations were in concordance with previous findings (Marhuenda et al., 2016; Singh, Singh, Ashish, & Salim, 2013) . 
| CON CLUS ION
The aim of this study was to determine the granulometric class of The Extrapole consortium, funded by the former Lorraine region (France), and more especially its project leader Elie DJANTOU BAUDELAIRE are also thanked for the initiative of the study presented in this manuscript.
CO N FLI C T O F I NTE R E S T
The authors declare no conflict of interest.
AUTH O R CO NTR I B UTI O N
NYN, EBN, and JS have made substantial contributions to conception and design, while MD, TJNM, and JP contributed to acquisition, analysis, and interpretation of data, and drafting the manuscript. All the authors critically revised and approved the final submitted version of the manuscript. Prior to submitting the article, all authors agreed on the order in which their names are listed in the manuscript.
E TH I C A L S TATEM ENT
This study does not involve any human nor animal testing.
O RCI D
Nicolas Njintang Yanou https://orcid.org/0000-0002-8949-7150

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