Ingeniería, Tecnología e Informática

Chemical characterization and morphology of moringa oleifera's wood as potential raw material for biorefineries

Caracterización química y morfología de la madera de Moringa oleifera como materia prima potencial para biorrefinerías

 

Julieta B. Benitez¹*, María E. Vallejos¹, María C. Area¹, Fernando E. Felissia¹

1- Programa de Celulosa y Papel - Instituto de Materiales de Misiones (CONICET-UNaM), Facultad de Ciencias Exactas Químicas y Naturales, Félix de Azara 1552 (3300). Posadas, Misiones, Argentina.

* E-mail: xulibel@yahoo.com.ar

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Resumen

La creciente demanda de biomasa forestal ha motivado el cultivo de plantaciones forestales de corta rotación en países desarrollados y en desarrollo. La Moringa oleifera es una especie de rápido crecimiento que se adapta a un amplio rango de suelos. La incorporación de esta especie como cultivo forestal en los sistemas silvo-pastoriles permitiría el uso de las semillas y hojas como forraje y productos medicinales/alimentos de bajo costo, obteniendo una mejor rentabilidad de los productores y mayor sustentabilidad de la actividad. El objetivo de este trabajo es conocer la composición química y la estructura morfológica de la madera de Moringa oleifera de dos edades diferentes (3 años y 8 años) para analizar su potencial uso como materia prima fibrosa de biorrefinerías.

Palabras clave: Composición química; Análisis microscópico; Relaciones biométricas; Biorrefinería.

Abstract

The growing demand for forest biomass has motivated the culture of forest plantations of short rotation in developed and in developing countries. Moringa oleifera is a fast-growing species that is to a wide range of soils. The incorporation of this species as forest cultures in silvopastoral systems would allow the use of seeds and leaves as forage and as low-cost medicinal products and foods, improving the profitability of producers and the greater sustainability of the activity. The aim of this study is to know the chemical composition and morphological structure of the wood of Moringa oleifera of two different ages (3 years-old and 8 years-old) to analyze its potential as raw material for biorefineries.

Keywords: Chemical composition; Microscopic analysis; Biometric Relationships; Biorefineries.

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Introduction

The growing demand of forest biomass for conventional (forest industry and as energy resource) and non-conven-tional uses (new materials and foods), has motivated the cultivation of short-rotation forest plantations in developed and developing countries (1-3). The application of sustai-nable forestry practices of fast-growing plantations for the production of woody biomass in agricultural or forest lands, fertile but degraded, enables the environmental and economic use of natural resources.

Moringa oleifera is native from the southern Himala-yas, northeast India, Pakistan, Bangladesh, and Afghanis-tan (4), and currently it is distributed in South America (Peru, Paraguay and Brazil) as an ornamental tree and as a "green" curtain (5, 6). It is a fast growing species, reaching 10-12 m in height and 20-40 cm in diameter at maturity (7), which adapts to a wide range of soils without requirements of natural fertilizers or agrochemicals. Its

cultivation brings a high amount of nutrients to the soil, in addition to protect it from external factors such as erosion, dehydration and high temperatures (8, 9).

This culture can be used in silvopastoral systems (forestation together with livestock) as an alternative to improve the profitability of the land through the diversif-cation of the production. The incorporation of this species as a forest culture would ofer the possibility of using the seeds and leaves as forage and low cost products for human consumption, improving the profitability of the producers and the sustainability of the activity. Additionally, the use of the log would generate a 100% utilization of this forest resource.

There are few precedents of the use of Moringa olei-fera wood (10, 11). Moringa oleifera's wood could be an alternative lignocellulosic material since it is characterized by its rapid growth and adaptability (10) in South America, where the leaves and seeds are already sold. In recent years, the cultivation of this species has aroused great interest in Argentina due to its nutritional and medicinal properties. The optimal conditions for its cultivation are average annual temperatures greater than 12°C and ave-rage annual rainfalls of 500 mm, which could be found in Misiones, Corrientes, Entre Ríos, Santa Fe, half the surface of the provinces of Formosa and Chaco, and part of Buenos Aires and Salta (11).

The Moringa oleifera is cultivated in many developing countries as a low cost food resource to prevent malnutri-tion and pathologies associated with shortages of essential dietary components (vitamins and minerals) (5). Its leaves and seeds can be used for the production of supplements for human food and animal forage (livestock, poultry, and pigs) due to its high nutritional value (12, 13). Its seed contains 35-40% of oil (70% of oleic acid) (14),which also makes it an important resource for biodiesel production (15, 16). The powder of seeds contains a cationic coa-gulant protein that can be used to decrease turbidity in water treatment (17). Also, the seed cake is composed of a natural and not toxic polypeptide that can be used to absorb and retain volatile substances, therefore it is a valuable additive in the fragrance industry to stabilize aromas (18), in the cleaning of vegetable oil, and in the sedimentation of fbers in the fruit juice and brewing industries (19). The extraction of natural antioxidant from leaves and stems has also been studied for potential applications in the food and pharmaceutical industries (20). There are few reports about its chemical composition, wood structure, fiber morphology, and aptitude for pulp and paper production (11, 21-24).

Cellulose (40-45%), hemicelluloses (25-40%), lignin (15-30%) and extractives are the main chemical com-ponents of wood (25, 26). Cellulose is a linear polymer (homopolysaccharide) composed of D-glucose units linked by β-1,4-glycosidic bonds, with a degree of polymerization (DP) higher than 10,000. Its structure is semi-crystalline because it is formed by amorphous and crystalline regions (25, 26). Hemicelluloses are amorphous polymers and they are composed by low molecular weight branched hetero-polysaccharide polymers (DP 50 - 500) from hexo-ses (glucose, mannose and galactose) and pentoses (xylose and arabinose), and diferent types of uronic acids (25, 26). Their proportion and composition depends on the type of species, age, variability of species, and others. Lignin is a complex three-dimensional phenolic and amorphous polymer constituted of phenyl propane units (27).The extractives are composed mainly of fatty acids, alcohols, phenols, terpenes, steroids, resinic acids, among others, and can be removed from wood by extraction with diferent solvents (26).

The Moringa oleifera is a hardwood species. The main morphological characteristic of hardwood is that they present vessels which have the function of conduction. These vessels can be seen in the cross section of the stem as "holes" called porous. Its structure is more complex than

that sofitwood due to the greater number of types of cellular elements which are highly variable in type, size, shape and arrangement (vessels, tracheids, fbers and parenchyma) (28). Unlike, softwood is formed mainly by longitudinal tracheids (approximately 90%) (29). The morphological characteristics of wood can be studied through the ob-servation of the microscopy structure of the cross, radial and tangential sections in which it is possible to see the size and distribution of porous (vessels), the fibers and parenchymal cells.

The aim of this work was to determine the chemical composition and morphological structure of Moringa oleifera's wood of two diferent ages (3 and 8 years-old) to assess its potential as a fbrous raw material for biorefneries, including paper production.

Materials and methods

Raw Materials

The three-years-old and eight-years-old stems of Moringa oleifera were extracted from experimental regional plantations. The samples without bark were milled in a laboratory knife mill, collecting the sawdust passing through an ASTM sieve N° 20 (0.840mm) and retained on ASTM sieve N° 80 (0.177mm). The fraction of the material retained on ASTM sieve N° 80 was used for the chemical determinations.

Physical and chemical characterization

Bark content was determined as percentage considering the weight of bark of a disk and the weight of the initial disk with bark. The basic density and dry density of wood were gravimetrically determined according to TAPPI T258 om-94.

Laboratory Analytical Procedures of the National Renewable Energy Laboratory (NREL - LAP) used for the chemical characterization were: "Preparation of sam-ples for Compositional Analysis" NREL/TP-510-42620, "Determination of Extractives in Biomass" NREL/TP-510-42619, "Determination of Ash in Biomass" NREL/TP510-42622, and "Determination of Structural carbohydrates and lignin in Biomass" NREL/TP-51042618. The different chemical components determined are detailed in Figure 1.

The determination of the structural carbohydrates was carried out by liquid chromatography (HPLC) using SHO-DEX SP810 and AMINEX-HPX87H (BIO-RAD) columns with detectors of refractive index and the diode array.

Starch content was determined by a colorimetric tech-nique (30). The oligomers and inorganic anionic content were quantifed by HPLC in the hot water extract using an AMINEX-HPX87H (BIO-RAD) column with a refractive index detector. Soluble inorganic were characterized by HPLC (Hamilton PRP-X-100 column) using a conductivity detector.

Holocellulose was determined by the modified acid Runkel ratio: Relationship between the fiber wall chlorite method (acetate bufer) (31) and alpha, beta, and thickness and lumen diameter (Eq. 3): gamma cellulose were quantifed by applying the TAPPI

T203 cm-99.

R = ^(Eq. 3) (Eq- 3)

Morphological characterization of wood

Histological sections (tangential, radial, and transverse cuts) were observed by an optical microscope (Zeiss) with image analyzer. The samples were disintegrated with chlo-rine dioxide and sodium carbonate (32) for the study of the morphological parameters of fbers. The average valúes of length, width and lumen of about 200 fbers were measu-red by optical microscopy and the coeficients of variation were calculated. The wall thickness was determined as the diference between the width and the lumen of the fbers.

The aptitude of Moringa oleífera for pulp and paper can be predicted from the biometric relationships calculated from the dimensions of the fibers (33). The biometric relationships are listed below.

Flexibility coefficient: Relationship between lumen width and fber width (Eq. 1).

Results and discussion

Physical and chemical characterization

Physical and chemical characteristics of Moringa oleife-ra samples are shown in Table 1. The percentage of bark was lower in the 8 years-old tree as compared with the youngest one because of its growth in diameter. The average density was lower than other species used for the pulp and paper production, which usually ranges between 0.30 and 0.60 g.cm^-3 (34). The Moringa oleifera density was almost half of eucalyptus one, which is about 0.48 g.cm^-3 in trees from the Northeast Region of Argentina (35).

The extractives content was variable depending on the age, the sample, and the solvent used. The substances ex-tracted by dichloromethane are usually waxes, fats, resins, phytosterols and non-volatile hydrocarbons, whereas the extractives in hot water are usually phenolic substances and some carbohydrates. The extractives content in ethanol was similar for both ages, although the extractive content in dichloromethane and in hot water showed a great diference between them. The sequential extraction (ethanol followed by hot water) in the 8 years-old sample has extracted a similar amount of substances than those extracted in hot water (single extraction) indicating that the components extracted with ethanol are also soluble in hot water (Table 1).The hot water extractives of the 8 years-old wood (23.89%) are almost two-folds of those of the 3 years-old, consisting of 18.86% oligomers (13.07% glucans and 5.79% xylans), and 1.89% of inorganic compounds (0.61% carbonates, 0.32% chloride, 0.75% phosphate, and 0.21% sulfate). Acetyl groups and simple sugars were not detected by HPLC. The total extractives content is very high as compared, for example, with that of eucalyptus (4.80% over dry wood) (36).

Figure 1: Methodology for the chemical characterization of Moringa oleifera.

 

Table 1: Physical and chemical characterization of 3 years-old and 8 years-old Moringa oleifera's wood.

	3 years	8 years
Bark (%)	21.68	16.16
Basic density (gcnr3)	0.21 ± 0.01	0.24 ± 0.02
Chemical composition
Hot water extractives (% otw)*	16.46 ± 1.45	23.89 ± 0,36
Total ethanol and hot water extractives(a)	16.90 ±0.34	23.88 ± 0.48
Ethanol Extractive state 1 (% otw)*	8.50 ± 0.34	8.86 ± 0.32
Hot water Extractive state 2 (% otw)*	8.40 ± 0.00	15.02 ± 0.16
Dichloromethane extractives (% otw)*	0.71 ± 0.00	1.12 ±0.01
Extractable carbohydrates (Starch) (% otw)*	0.41 ± 0.03	12.44 ± 0.31
Total Lignin	19.60 ± 0.05 (23.6)**	20.37± 0.3 (26.8)**
Soluble Lignin (% otw)*	1.30 ± 0.03 (1.56)**	1.23± 0.03 (1.62)**
Insoluble Lignin (% otw)*	18.30 ± 0.02 (22.0)**	19.14± 0.27 (25.1)**
Total structural carbohydrates	50.00 ± 0.05 (60.2)**	45.17± 0.05 (59.3)**
Glucan (% otw)*	38.40 ± 0.08 (46.2)**	34.95 ± 0.08 (45.9)**
Xylans (% otw)*	8.10 ± 0.08 (9.75)**	7.99 ± 0.08 (10.5)**
Arabinans (% otw)*	0.80 ± 0.02 (0.96)**	0.48 ±0.02 (0.63)**
Galactans (% otw)*	1.70 ± 0.10(2.04)**	1.05 ± 0.11 (1.38)**
Mannan (% otw)*	1.00 ±0.00 (1.20)**	0.70 ± 0.00 (0.92)**
Acetyl groups (% otw)*	1.70 ± 0.00 (2.05)**	2.17 ± 0.00 (2.85)**
Hemicelluloses (% otw)****	13.30 ± 0.28 (16.0)**	12.39 ± 0.29 (16.3)**
Ash at 525°C (% otw)*	10.40 ± 0.03**	3.92 ± 0.04

(a) Sequential extraction.

Percentages are expressed on dry base of: * total wood (% otw), ** extracted wood (% oew). *** Hemicelluloses were calculated based on the sum of xylans, galactans, arabinans, mannans, and acetyl groups.

Significant diferences were also found in ashes content, being much higher in the 3 years-old tree as compared to the 8 years-old one. The starch content extracted in hot water shows the opposite behavior.

The contents of alfa, beta, and gamma cellulose (holo-cellulose) of total and extracted wood are shown in Table 2.

Table 2: Analysis of the holocellulose fraction.

	Holocellulose (% oth*)	Holocellulose (% oew**)
Alpha celullose	59.93 ± 0.06	43.93
Gamma celullose	31.29 ± 0.26	22.94
Beta celullose	8.78 ± 0.32	6.43
Total	100.00	73.31

Percentages are expressed on dry base of: * total holocellulose (% oth), ** extracted wood (% oew).

Holocellulose content was 73.31% oew (± 0.25), hig-her than that reported by Khider et al. (37) for Moringa oleifera in Sudan (68.5%). The alpha cellulose amount (43.93% oew) was similar to the glucans quantified by HPLC (45.9% oew), which would indicate that glucans content corresponds mainly to cellulose. This result could be interesting to produce nanofbrilated or microfbrilated cellulose.

Morphological characterization

The images of the histological sections of the samples of 8 years-old obtained by optical microscopy are shown in Figures 2 to 4. Fibers of very thin walls and solitary vessels are observed in the cross sections (Figure 2).

Figure 2: Cross sections of Moringa oleifera's wood (objectives 4X and 10X).

Uniseriate and biseriate woody rays of 1 to 9 cells in height, solitary vessels, and multiples vessel can be obser-ved in the tangential sections (Figure 3).

 

Figure 3: Tangential sections of Moringa oleifera's wood (objectives 4X and 10X).

The radial sections show inhomogeneous woody rays and fbers widened in their central part can be observed in Figure 4. This and other peculiar forms of fbers are shown in Figure 5. A large quantity of parenchymal cells can be observed, which justify the high extractives content of the wood. The fbers are very wide at the center (a and b).Some fbers have bifurcations at the ends (f, h, i, and k). Vessels have also peculiar shapes (c, d, e, g, and j). Fibers are broad and thin walled so they are supposed to easily collapse (28).

Figure 4: Radial sections of Moringa oleifera's wood (objectives 4X and 10X). RECyT / Año 20 / N° 30 / 2018

Figure 5: Optical microscopy images of fbers (a, b) (objective 4X). Fi-ber ends (f, h, i, and k) and vessel elements (c, d, e, g, and j), (objective 20X).

Figure 6: Length distribution of pulp fbers (200 fbers for each type of pulp).

The fber length distribution for each of the pulps is presented in Figure 6. The MO 3 year-old shows a bimodal fber length distribution. The range of fber length the MO 3 year-old comprises 76% of the fbers between 700 - 1100 µm, whereas, in the MO 8 year-old 63% of the measured fbers are longer, between 900 - 1300 µm, which is unders-tandable considering the diference in age of trees.

Fiber dimensions of Moringa oleifera samples are detailed in Table 3. Other species are included for comparative purposes. The dimensions of the fbers vary between species and in the same species depend on climate, soil, and position in the stem. Thus, it is very dificult to make comparisons between fbers that, for example, do not come from the same region. However, broadly speaking, it can be said that Moringa oleifera fbers have a typical length of hardwoods, although the fibers appear to be shorter than those of Ekhuemelo and Udo (38) originating from Nigeria. On the contrary, the fbers are much wider than those of other species, according to this work and those of Olson and Carlquist (39) and Cobas and Molina Tirado (21). The values reported by Ekhuemelo and Udo (38) are atypical, considering the shape of the fbers as shown in Figure 5.a and 5.b.

Table 3: Fiber dimensions of the Moringa oleifera's samples and other wood species.

The biometric relationships of the samples and of the studied Moringa oleifera samples and those of Sugarcane bagasse and eucalyptus are shown in Table 4.

Table 4: Main biometric relationships of fibers from Moringa oleifera's wood, Sugarcane bagasse and eucalyptus.

The Flexibility coeficient (Eq.1), indicates the capacity of fbers to collapse and form a paper web. Table 4 shows that fic values for Moringa oleifera are really low consi-dering the general rule (it has to be more than 50) and also as compared with usual papermaking species. Felting index (Eq. 2). Table 4 shows that Fel which valorizes the length of the fber, presents the same behavior. Runkel Index (Eq. 3) indicates the rigidity of the fber and the strength of fber walls. It R was significantly lower than those of the other species in Table 4.

Considering the fber quality of both studied ages, typical of juvenile wood, this raw material would be better used as a source of nano or microfbrillated cellulose or as material for the manufacture of high-value chemical products.

Conclusions

Moringa oleifera wood presents higher starch content than usual commercial hardwoods.

Due to the high content of extractives, an extraction in hot water is recommended as the frst step of fractionation.

Since fbrous and biometric parameters indicate that Moringa oleifera is not suitable for paper production, it can be exploited as source of high-valuable chemicals.

The high content of alpha cellulose makes Moringa oleifera an interesting raw material for the production of nanofbrilated cellulose or microfbrilated cellulose.

Acknowledgements

The authors acknowledge the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET) for the doctoral scholarship; PROCYP, IMAM (Argentina) for the financial support. Mr. Carlos Brizuela, President of the "Moringa Includes" Cooperative is acknowledged for providing the Moringa oleifera wood.

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Recibido: 21/08/2018. Aprobado: 09/10/2018.