Chemical composition, antitermite, antifungal properties, and wood protection systems from Canarium schweinfurthii (Aiele) resin from Gabon

This work evaluates the antifungal and antitermite properties, as well as the contribution of three resin fractions, i.e., raw resin (RR), essential oil (EO) and purified resin (PR) of Canarium schweinfurthii (Aiele) from Gabon, to the durability of wood material. To this end, chemical analysis using gas chromatography coupled with mass spectrometry (GC/MS) of the three fractions was first carried out, followed by a study of their antifungal and antitermite activities, and finally a study of their effects on the protection of wooden blocks subjected to degrading agents (termites and fungi). The results of the chemical analyses show that EO is made up of monoterpenes and monoterpenoids, while RR and PR consist of a mixture of monoterpenes, monoterpenoids and triterpenes. Biological tests of these fractions against Rhodonia placenta (RP), Coniophora puteana (CP), Trametes versicolor (TV) and Pycnoporus sanguineus (PS) demonstrated moderate antifungal activity but strong antitermite activity. These fractions improve the durability of impregnated wood blocks against fungi, and provide remarkable resistance to termites. However, their leachability properties are clearly demonstrated.

Biobased materials offer an exellent solution for reducing greenhouse gas emissions and combating global warming. For several years now, the use of wood as a construction material for buildings and structures of high environmental quality has been steadily increasing. Wood has a several technical (low thermal conductivity, mechanical strength, low density, etc.) and economic advantages. It also benefits from its image as an ‘ecofreindly’ material of natural origin, derived from a renewable resource that requires little energy to process. However, due to its natural formation, wood is sensitive to environmental factors and biological agents. Consequently, it is essential to use protective systems to ensure that exterior woodwork lasts as long as expected.

Improving the durability of wood against biological degradation agents for outdoor applications in use classes 3 or 4 is traditionally achieved by deep impregnation with synthetic preservatives to prolong the wood lifespan. Creosote, lindane, dieldrin, pentachlorophenol or copper chromium arsenate (CCA) have been largely used in the past to protect wood and other lignocellulosic materials from termite and wood-destroying fungi attacks. More recently, other less toxic products such as pyrethrums, triazoles or carbamates have been proposed. However, many of these products have been banned or severely restricted because of their high toxicity to humans and the environment [1,2,3].

Among the possibilities explored in new research, exploiting the extractives naturally present in wood (or in exudates) species with high natural durability is an interesting option. Numerous studies have focused on alternative active ingredients derived from natural and renewable raw materials [4,5,6,7,8]. These studies emphasized wood extractives, some of which pocess properties comparable to those of synthetic preservatives [9]. Certain wood species are naturally resistant to termite attack due to their high content of extractive compounds, which form part of their natural defence systems [4, 10,11,12,13,14]. Recent studies have highlighted the feasibility of adding value to wood species with high natural durability by extracting the active extractive components responsible for durability, including resins or essential oils [10,11,12,13,14,15,16,17].

The literature reveals that certain tree species produce or exude resin to protect themselves against microorganisms, including the Burseraceae family. Among this family, the most common species in Gabon include Aucoumea klaineana Pierre, Canarium schweinfurthii, Dacryodes buettneri and Dacryodes edulis. These trees produce abundant oleoresins and contain high proportions of compounds with in vitro antioxidant, antibacterial and antimicrobial properties in their essential oils [18, 19]. Several authors have shown that the essential oils of various plants possess antitermite and antifungal properties [15, 20,21,22,23,24,25,26,27].

For this reason, the present study focusses on Canarium schweinfurthii. Belonging to the genus Canarium and of the species schweinfurthii, it is commonly known as Aiele, the official commercial name. Its scientific appelation is Canarium schweinfurthii Engl and the synonyms are C. occidentale A. Chev. and C. khiala A. Chev. This member of Burseraceae family has a very wide geographical range in Africa [28,29,30,31]. Aiele has been reported in densely wooded areas of Guinea Forestery and is scattered throughout Gabon's secondary forest [32]. The tree is a large species with compound leaves and numerous predominant lateral veins. Its fruit is edible. This oleaginous plant holds a rich cultural heritage and produces a greenish-white, turpentine-scented resin. The architecture of the tree presents slight buttresses, the young branches are covered in down, and the leaves are compound, pinnately arranges, and form rosettes at the tips of the branches. Its small, greenish flowers give way to drupaceous and ovoid fruits resembling small plums with a purplish tinge when ripe. The bark is greyish and heavily cracked. According to Tchouamo et al. (2001) [30] and to Tchiegang et al. (2001) [31] the wood of C. schweinfurthii is pinkish-white and very tender. The fruits, which contain 30–50% oil, are of particular interest. They are eaten softened or as an accompaniment to starchy foods [33]. The oil is used to make shampoos, shoe polish or as biofuel [31, 34,35,36].

In Gabon, according to Aubreville (1959) [37], its resin is more often used to make ‘indigenous torches’. According to Burkill (1994) [38], the resin serves as a fumigant to repel mosquitoes. Traditionnal medicine employs the resin for treateing various illness as wounds and microbial infestations, as weel as for its emollient, stimulationg and diuretic properties [39]. In addition, its wood is cut into planks or used to make pirogues by local communities [40].

Although, many studies have demonstrated the effectiveness of plant extracts and essential oils against various micro-organisms and insects, the lack of scientific data on the termicidal and fungicidal properties of Canarium schweinfurthii resin from Gabon, particularly regarding its durability, motivated the present research.

The general aim of this work is to exploit the essential oils extracted from oleoresins and their crude resins for their potential use in formulating eco-friendly and biodegradable wood preservatives to protect the treated material against termites and wood-destroying fungi. For this purpose, essential oil was separated from oleoresin using steam distillation process with a Clevenger apparatus, and oleoresin was purified using different solvents. Each fraction was analysed using GC–MS and subjected to various biological tests to evaluate their termicidal and fungicidal properties.

Biologic materials

The oleoresins of Canarium schweinfurthii Engl was collected by tapping in Gabon, in August at the Sibang arboretum, located in the center of Libreville city.

A single C. schweinfurthii tree was selected and wounded at 1.30 m above the ground using a clean machete. Three days later, the collected raw resins (RR) were stored at 4 °C in a refrigerator.

Two types of wood destroying fungi, including four strains were used:

• Two white rot: Trametes versicolor (TV) [(Linneus) L. Quélet strain CTB 863 A] supplied by the Interactions Trees-Microorganisms Laboratory (IAM), UMR-INRA 1136, University of Lorraine and Pycnoporus sanguineus [(L.) Murrill, 1904] provided by the CIRAD mycobank.

• Two brown rot: Coniophora puteana (CP) [(Schumacher ex Fries) Karsten, strain BAM Ebw.15] and Rhodonia placenta (RP) [(Fries) Cooke sensu J. Eriksson, strain FPRL 280], both provided by the CIRAD mycobank. The European subterrean termite species Reticulitermes flavipes (ex santonensis de Feytaud), collected from Oleron Island, France (Lat. 45° 49′ 5.9′′ N; Long. -1° 13′ 47.8′′ W), were used for this study.

Mini blocks [dimensions: 25 ×ばつ 15 ×ばつ 5 mm³ (L, R, T)] of Scots pine sapwood (Pinus sylvestris L.) and beech (Fagus sylvatica) were employed for all biological tests.

Chemical materials

Ethanol (absolute anhydrous grade) and acetone (analysis grade) were provided by Carlo Erba Company (Val-de-Reuil, France) and Merck company (Darmstadt, Germany) companies, respectively. Malt extract and agar powder for microbiological tests, were sourced from Sigma Life Science (St Quentin Fallavier, France). Industrial-grade tebuconazole was provided by PPG Industries (Rueil-Malmaison, France).

Extraction and purification of the different resin’s fractions

Essential oil production

Essential oil (EO) was extracted from the raw resin (RR) using a hydrodistillation process with a Clevenger type apparatus for 4 h, similarly than thise used by Bruneton (2016) [41] and as described by Clevenger (1928) [42]. Figure 1 illustrates the laboratory setup used for EO production. RR was immersed directly in distilled water in a double-neck flask equipped with the Clevenger apparatus. To reduce viscosity, the sticky RR was pre-heated in a breaker containing hot water before being transferred to the flask. The condensate containing EO was collected in the separating funnel, and the two phases were separated immediately by decantation. The EO (upper phase) was recovered in an Erlenmeyer, dried using magnesium sulfate and then filtered through sintered glass.

The EO yield was calculated as follows [43]:

Resin purification

To obtain the purified resin (PR), we added the raw resin (RR) in ethanol, then centrifuged the mixture to separate the resin and impurities (such as sand, bark, etc.). Finally, using a rotary evaporator, the ethanol was evaporated completely, leaving only the purified resin (PR).

More details on the studied resins can be found in the previous study carry out by Bedounguindzi et al. (2020) [15].

Termite species and wood rots

Biological assays were conducted with a European termite species, Reticulitermes flavipes (ex. santonensis). Termites were collected from Oleron Island, (Lat. 45° 49′ 5.9′′ N; Long. -1° 13′ 47.8′′ W). The colony was reared in a climatic room at 27 ± 2 °C and relative humidity superior at 75%.

Different types of fungi were used: two white rot, Trametes versicolor and Pycnoporus sanguineus, and two brown rot, Coniophora puteana and Rhodonia placenta. The Trametes versicolor strain (Tv 110.102 MA AD) was supplied in Petri dishes by the laboratory Interactions Trees-Micro-organisms (IAM) UMR–INRA 1136 of the University of Lorraine. The strain Pycnoporus sanguineus (Ps/1969/1) was supplied in Petri dishes by the Center for International Cooperation in Agronomic Research for Development (CIRAD) in Montpellier. The other strains were part of the mycothèque of the Laboratory of Studies and Research on Wood Material (LERMAB). The strains were subcultured on malt–agar medium in the Petri dishes and maintained at 22 ± 2 °C with a relative humidity (RH) of 70 ± 5%.

Chemical analysis

Essential oil and resins were analyzed using a Perkin Elmer Clarus 680 Gas Chromatograph (GC) equipped with a fused silica DB-5MS [(diméthyl-/diphényl-polysiloxane, 95:5] column (30 m, 0.25 mm, 0.25 μm). The GC was coupled with a Perkin Elmer Clarus SQ8 Mass Spectrometer (MS) and monitored by Turbo Mass v.6.1 software.

Before injection into the GC–MS system, essential oil and resin sample was derivatizated to improve the detection of all chemical compounds. For this illation derivatization process, 2 mg of essential oil was solubilized in glass tube with 50 μL of BSTFA + 1% TMCS solution (Bistrimethylsilyltrifluoroacetamide + Trimethylchlorosilane). The glass tube was sealed and incubated in an oven at 70 °C for 120 min. Afterward, the tube was opened to allow BSTFA evaporation. The derivatized sample was dissolved in 1 mL of pure ethyl acetate (C₄H₈O₂). A 1-μL aliquot of this solution was injected in the Gas Chromatograph at an inlet temperature of 250 °C in splitless mode. Helium was used as carrier gas.

The typical temperature program was: 80 °C (2 min), increase by 10 °C.min⁻¹ to 190 °C, then increased by 15 °C.min⁻¹ to 280 °C (held for 10 min), and increased by 10 °C.min⁻¹ to 300 °C 15 °C.min⁻¹ (held for 14 min).

For some samples, the program was slightly modified to improve peak resolution, resulting in variable retention times for some compounds. A helium flow rate of 1 mL/min was used as the mobile phase. After separation, compounds were transferred to the mass spectrometer via a transfer line heated to 250 °C. Ionization was achieved using the electron impact method (70 eV ionization energy).

Chemical compounds were identified based on their mass spectra by comparison with the NIST Library (2005) using the NIST MS Search 2.0 software (2011). Identification was considered reliable when match and reverse match coefficients exceeded 900.

Biological assays

Antifungal activity tests

Fungal tests involved direct contact of fungal mycelium with agar medium infused with EO and resin fractions at various concentrations. The mycelium was grown in 8.5 cm petri dishes filled of malt–agar medium (16 mg malt + 10 mg agar + 374 mL distilled water). Essential oil or resin fractions were introduced into the sterilized medium (steam pressure sterilization process carried out at 120° C -1 bar, for 25 min).

The essential oil was solubilized in 1 mL ethanol, and the resin fractions were dissolved in 2 mL ethanol. Three concentrations were tested into the agar medium: 500 ppm, 1000 ppm, 1500 ppm. Two controls were prepared: the malt–agar medium without ethanol (Ts eth) to establish baseline growth, and malt–agar medium with ethanol (Ta eth) to assess the inhibitory effect of the essential oil or resins. Plates were inoculated centrally using a small portion of a freshly grown fungal colony on malt–agar agar. The cultures were maintained at 22 ± 2 °C and 70 ± 5% RH. Growth was evaluated daily by measuring the diameter of the fungal colony, estimated as the average of two perpendicular diameters, and expressed as a percentage of the available surface area (i.e., the diameter of the petri dish). For each concentration, three replicates were prepared to minimize the experimental error.

Antitermite activity test

Six concentrations (two of the essential oils, two of the raw resins and two of the purified resins) of C. schweinfurthii were tested against termites using screening tests: mass ratios of 50:50 and 25:75 (EO: acetone; PR: acetone; and RR: acetone). For essential oils, 70 μL of solutions were impregnated onto Whatman filter papers (WhatmanTM, CAT n° 1001-325—Grade 1, United Kingdom) before being exposed to termites (Reticulitermes flavipes). For purified resin (PR) and raw resin (RR), 90 μL of solutions were impregnated onto Whatman filter papers before being exposed to termites.

The papers impregnated with the various test solutions were dried either in the open air [20 °C–65% relative humidity (RH); 2 h] or in the oven (103 °C; 1 h). After drying, each sample was weighed before being placed in contact with the termites.

The tests were carried out in Petri dishes (5.5 cm in diameter), where 15 g of wet sand (1 volume of distilled water for 4 volumes of sand) was placed on the periphery. The treated Whatman papers were placed on a plastic grid (to avoid moisture) in the middle of the Petri dish (containing wet sand) and 20 worker termites were added to each test device. The Petri dishes were placed in the dark at 27° ± 2 °C, RH ≥ 75%. Three types of controls were also tested under the same conditions: water-soaked paper, acetone-soaked paper and filter paper-free boxes to determine the end of the test. Every 2 days, each test setup was observed to check sand humidity and to keep track of termite behaviour and activity. Water was added when needed. When all termites contained in the diet control setups had died, the test was stopped (maximum duration of 21 days). At the end, termite survival rates (TSR %) were determined, the paper samples were cleaned and air-dried or oven dried, according to the same procedure as after impregnation. The dried mass of the cellulose papers was then measured and the Weight Losses (WLterm. %) due to termite degradation were calculated.

Wood blocks impregnation

The impregnation process was adapted from the guidelines specified in the NF X 41-568 (2014) standard [44], with some adjustments concerning the wood sample size (dimensions: 25 ×ばつ 15 ×ばつ 5 mm³ [L, R, T]) [45]. Beech wood and Scots pine sapwood samples were first dried at 103 ± 2 °C until their mass stabilized and then weighed (m₀). These weight measurements were performed using a high precision balance (linearity of 0.2 mg, reproducibility of 0.1 mg) [Kern ABT 220 4 M, Germany]. For each formulation, 30 dried wood samples (12 specimens of beech and 18 specimens of Scots pine) were then vacuum-treated at 5 mbar for 30 min before impregnation with the different treatment solutions, based on the use of EO, RR and PR. The samples were kept immersed in the solution for 1 h at atmospheric pressure. After that, the wood samples were dried at 103 ± 2 °C for 48 h and weighed (m₁).

For each wood species, three series of solutions containing raw resin (RR), essential oil (EO) and purified resin (PR), with different concentrations prepared in ethanol, were tested (% m/m):

• 4 solutions of RR, diluted in ethanol: RR (1%); RR (5%); RR (10%); RR (20%).

• 1 solutions of EO, diluted in ethanol: EO (5%).

• 4 solutions of PR, diluted in ethanol: PR (1%); PR (5%); PR (10%); PR (20%).

The Weight Percentage Gain (WPG) was determined according to Eq. 2, as follows:

where m₀ is the dried mass of the wood block before impregnation, and m₁ is the dried mass of the wood block after impregnation.

Leaching

Leaching process was adapted from the guidelines of the NF X 41-568 (2014) standard [44]. Samples were immersed in distilled water (1 volume of wood for 5 volumes of water) and subjected to two series of three leaching periods of increasing duration under continuous shaking at 20 ± 2 °C. For each formulation, a total of 6 samples of treated beech wood and 9 samples of treated scots pine wood were separately leached. Untreated Beech and Scots pine, as control samples were also submitted to similar leaching process, in separated leaching flasks.

Water was replaced by fresh water after each leaching period. A first cycle of 3 leaching periods of 1, 2 and 4 h was performed. Samples were then kept air-drying at room conditions for 16 h. Leaching was then performed for 3 additional periods of 8, 16 and 48 h, with a change of water between each period. The leached samples were then dried at 103 ± 2 °C and weighed (m₂). The percentage of leaching and the weight percentage gain after leaching were calculated following Eqs. 3 and 4, respectively:

where m₀ is the dried mass of the wood block before impregnation, m₁ is the dried mass of the wood block before leaching and m₂ is the dried mass of the wood block after leaching.

Wood treatment

Fungal resistance

The effects of different fractions of canarium schweinfurthii resins on the decay resistance of wood blocks, before and after leaching, were assessed using mini-block screening tests, such as described by Bravery (1978) [45] and adapted from the guidelines of the EN 113-2 (2023) standard [46]. Sterile culture medium was prepared from malt (40 g) [Millipore, purified and clarified malt extract, suitable for microbiology, Sigma Life Science (St Quentin Fallavier, France)] and agar (20 g) [Millipore, BSM Agar, suitable for microbiology, NutriSelect® Basic, Sigma Life Science (St Quentin Fallavier, France)] in distilled water (1 L), and placed in a culture Pétri dish inoculated with a small piece of mycelium of a freshly grown pure culture and incubated for 2 weeks at 22 ± 2 °C and 70 ± 5% RH to allow full colonization of the medium by the mycelium.

Three blocks (two treated and one control, Scots pine for Coniophora puteana and beech for Trametes versicolor) were placed in each Petri dish. Each experiment was tri-plicated in a separate way, both for unleached samples and for leached samples (i.e., 6 samples per modality). Virulence control samples were also performed on twelve specimens of untreated Scots pine sapwood and beech wood. Incubation was carried out for 12 weeks at 22 ± 2 °C, 70 ± 5% in a climatic room. Once the fungal exposure was completed, the mycelium was removed from the wood blocks, and the specimens were dried at 103 ± 2 °C and their final weight recorded (m₃). Weight loss (WL) due to the fungal degradation of the sample was determined as a percentage of the initial anhydrous mass of the unleached and leached samples according to Eqs. 5 and 6, respectively:

where m₁ is the dried mass of the wood block before leaching, m₂ is the dried mass of the wood block after leaching and m₃ is the dried mass of the wood block after fungal decay exposure.

Resistance to termites

The termite resistance was determined using a non-choice screening test adapted from the guidelines of the EN 117 (2023) standard [47]. The protocol for these tests is also detailed by Bedounguindzi et al. (2020) [15]. For each product combination and for untreated control samples, three replicates were tested against Reticulitermes flavipes.

Each sample was placed in a 9 cm diameter Petri dish containing 40 g of Fontainebleau sand (4 volumes of sand for 1 volume of deionized water). The samples were then placed on a plastic mesh to avoid waterlogging. One block was placed per Petri dish. A total of 50 termite workers, one nymph and one soldier were introduced into each Petri dish. The Petri dishes were placed in the dark in a climatic chamber at 27 ± 2 °C, RH ≥ 75%, for 4 weeks. Visual observations were conducted on a weekly basis to add needed water and check termite behavior.

At the end of the termite exposure, the samples were removed, cleaned of sand, and the survival rate of the termites (TSR) was calculated according to Eq. 7:

The samples were given a visual rating according to the guidelines of the EN 117 (2023) standard [47], with some adjustments regarding the sample’s sizes. The visual rating was expressed as follows: 0 = no attack, 1 = attack attempt, 2 = slight attack, 3 = medium attack, 4 = severe attack.

The samples were then oven-dried at 103 ± 2 °C and weighed (m₄). Finally, the mass losses of unleached and leached samples were calculated according to Eqs. 8 and 9, respectively:

where m₁ is the dried mass of the wood block before leaching, m₂ is the dried mass of the wood block after leaching, and m₄ is the dried mass of the wood block after exposure to termites.

Extraction rates

Extraction rate of the essential oil (EO)

The extraction rate of the essential oil is calculated as a function of the mass of essential oil obtained relative to the quantity of oleoresin harvested, as shown in Table 1.

The extraction rate for the essential oil of C. schweinfurthii is 13.24%. This extraction rate is higher than those proposed in the literature: 4.48% and 6.92% [19, 48]. However, these resins yield a higher content of essential oils compared to other plant organs.

Extraction rate of purified resin (PR)

The extraction rate of the purified resin, calculated from the raw resin harvested and dissolved in three different solvents, is shown in Table 2.

Extraction rates vary depending on the solvents used. Methanol generally leads to lower extraction rates than ethanol and acetone. There are few differences between these solvents. Ethanol is the only potentially bio-sourced solvent, which is why chosen for our tests. The extraction rate for C. schweinfurthii resin was 99.63%. A large quantity of solvent was required to dissolve the resin adequately, which may influence the properties or even modify the chemical composition of the pure resin being tested. As this work has not been reported by other authors, it has not been referenced.

Chemical analysis of the three fractions of C. schweinfurthii resin using gas chromatography–mass spectrometry

Chemical analysis of the raw resin

Chromatographic analyses of the RR of C. schweinfurthii identified 12 compounds that account for a significant proportion of the compounds present. As specified in Table 3, the majority are: α-amyrin (40.1%), cis-β-terpineol (13.4%), terpinen-4-ol (10.4%), β-amyrin (7.7%) and 3-epi-α-amyrin (6%), accompanied by other constituents at relatively lower levels: o-cymene (4.3%), lanosterol (4.1%), terpinolene (3.8%), sabinene (3.2%), epi-β-amyrin (3%), γ-terpinene (2.4%) and betulin (1.4%). RR is also rich in triterpens and monoterpenes.

Chemical analysis of the essential oil (EO)

According to Tables 4, 16 constituents were identified in the EO of C. schweinfurthii, with an overall content of 99.9%. This essential oil is made up of four main compounds, including α-phellandrene (30.1%), o-cymene (20.5%), γ-terpinene (15.7%) and α-terpinene (8.7%), as well as other compounds at moderate or even low levels: 2-menthene (5.6%), cis-4-thujanol (4.3%) and α-pinene (2.2%). These results do not corroborate those of several authors: first, the EO of C. schweinfurthii from Bangui (Central Africa) yielded 17 compounds, with three dominant ones: octyl acetate (60.0%), nerolidol (14.0%) and octanol (9.5%) [19]. Next, the EO of C. schweinfurthii from Libreville (Gabon) consisted of 55 components, with four main ones: limonene (52.1%), sabinene (19.0%), α-pinene (10.7%) and ρ-cymene (4.3%) [48]. Finally, the EO of C. schweinfurthii from Uganda yielded 32 compounds, with the main components being γ-terpinene (32.4%), α-phellandrene (17.9%), α-thujene (14.0%), β-phellandrene (12.9%) and p-cymene (8.5%) [26].

Chemical analysis of the purified resin (PR)

In Table 5, it can be seen that the PR of C. schweinfurthii contains 8 compounds that account for 99.9% of the total resin. It is composed of: α-amyrin (41.1%), cis-β-terpineol (14.5%), terpinen-4-ol (13.8%), epi-α-amyrin (9.6%) and β-amyrin (9.4%). These results are in agreement with those already described in numerous studies on other resin species belonging to the Burseraceae family [27, 49,50,51,52,53,54].

Biological tests

Antifungal tests

Antifungal activity with RR

The tests were carried out with three concentrations: 500, 1000 and 1500 ppm on the four wood-destroying fungi. Preliminary tests showed that at 200 ppm there was no effect and that the available quantities were sufficient to conduct experiments at higher concentrations.

According to Table 6, RR of C. schweinfurthii at 500 ppm showed low antifungal activity against the four fungal strains, with growth inhibitions not exceeding 59.8%. At 1000 ppm, RR of C. schweinfurthii also showed moderate antifungal activity, with growth inhibitions ranging from 43.1% to 66.6%. At the highest concentration of 1500 ppm, growth inhibition reached 79.6%. Therefore, the RR of C. schweinfurthii demonstrated good antifungal activity. In fact, C. puteana and T. versicolor were the most sensitive fungi to the RR from C. schweinfurthii, with growth inhibitions comprised between 74.1% and 79.6%.

Antifungal activity of EO

According to Table 7, at 500 ppm, T. versicolor, P. sanguineus, C. puteana and R. placenta showed sensitivity to C. schweinfurthii EO, with fungal growth inhibitions ranging from 59.2% to 84.4%. The EO of C. schweinfurthii also showed very strong activity against the fungal strains studied from 1000 ppm, with fungal growth inhibitions equal to 100%. Figure 2 provides further visual confirmation.

Antifungal activity of PR

Table 8 shows that PR from C. schweinfurthii exhibited low antifungal activity against the four fungal strains studied at 500 ppm, with fungal growth inhibitions ranging from 27.0% to 39.1%. At 1000 ppm, the growth inhibitions ranged from 28.9% to 58.0% for all the fungal strains tested. At 1500 ppm of the RR of C. schweinfurthii, the fungal inhibition rate varied from 40.7% to 74.8% for Trametes versicolor and R. placenta, respectively.

Antitermite tests

Screening test with raw resin (RR)

The results presented in Table 9 highlight that the control paper impregnated only with water or acetone has no effect on termite behavior, as shown by the recorded mass losses (WLterm˃ 18.7%) and termite survival rate (TSR˃ 65.0%) at the end of the trial (Fig. 3). Essentially, all treatments involving RRs demonstrated a protective effect on Whatman paper. For filter paper treated with the ‘RR/Ac = 1/3 (mm/m)’ concentration, mass losses were 4.4% at 20 °C for 2 h and 6.2% at 103 °C for 1 h, with survival rates of 5% at both 20 °C and 103 °C. According to these results, RR showed strong antitermite activity.

Screening test using paper soaked in essential oil (EO)

Table 10 shows that papers impregnated with C. schweinfurthii EO are highly resistant to termites. It can be seen that all the termites are dead (TSR = 0%) at the end of the test, and that the Whatman papers impregnated with EO are almost not degraded by termites (WLterm = 0.7%) at the highest concentration (1/1). At 1/3, i.e. the lowest concentration, the Whatman papers suffered very little degradation by termites. The EO of C. schweinfurthii therefore has strong antitermite activity.

Screening test using paper soaked in purified resin (PR)

The results in Table 11 highlight that all treatments involving PR showed a protective effect for the impregnated Whatman paper. The best results were obtained with the concentrated acetone solution (1/1), leading to mass losses of 0% and a survival rate of 0% when the papers were air-dried at 20 °C.

In fact, for cellulosic paper treated with the lowest concentration [PR/Ac = 1/3(m/m)], mass losses were 4.7% with an air-drying (20 °C–2 h) and 8.4% with an oven-drying (103 °C–1 h), while termite survival rates were 0.0% at 20 °C and 6.7% at 103 °C. This means that PR from C. schweinfurthii possesses strong antitermite activity.

The effect of drying temperature was clearly highlighted: exposure to a high temperature for 1 h reduces the efficiency of treated papers. PR from C. schweinfurthii showed efficacy even after drying at 103 °C, at the highest concentration (1/1).

Test on impregnated wood

Weight gain

Table 12 highlights that, before leaching, the mass gains of wood samples impregnated with RR of C. schweinfurthii were 19.8% and 13.4% for scots pine and beech woods, respectively. After leaching, the mass gains were reduced to 17.4% and to 11.3% for scots pine and beech woods, respectively.

For wood samples treated with the EO of C. schweinfurthii, mass gains before leaching were 2.4% for scots pine and 1.6% for beech. After leaching, the mass gains of the wood samples treated with C. schweinfurthii essential oil were equal to 0% for both scots pine and beech.

For wood samples treated with RP of C. schweinfurthii, the mass gains before leaching were 18.4% for scots pine and 10.6% for beech. After leaching, the mass gains of the wood samples treated with C. schweinfurthii essential oil were reduce to 16.1% and to 8.8%, respectively.

It can, therefore, be observed that the EOs impregnate very slightly better in scote pine wood than in beech wood, but only by a very small amount, and that they are not resistant to leaching, or even to oven-drying after impregnation.

Table 12 also shows that PR impregnates much better in scote pine than in beech woods, and that these treatments are fairly resistant to leaching process.

The differences observed between the impregnation rates of scots pine and beech are probably linked to the different anatomy characteristics of these two wood species. As scots pine is a resinous tree, the penetration of solutions is certainly facilitated by the resin canals and the path taken during the exudation of resins in their tree of origin, whereas beech has much wider vessels. PRs are resistant to leaching, certainly due to their chemical profiles and above all their textures, as they are sticky to the touch.

Resistance to fungi

Treatment with RR

Table 13 shows that the mass losses of the treated wood blocks with the RR of C. schweinfurthii against the four fungi at low concentrations (1%, 5% and 10%) are lower than the mass loss of the control wood blocks. For example, at the concentration of 10% on T. versicolor, the mass loss of the treated specimens was 26.6% compared to a mass loss of 30.9% for the control specimens; on P. sanguineus, the treated specimens showed a mass loss of 7.7%, while the control specimens showed a loss of 39.5%; on C. puteana, the treated specimens showed a loss of 37.6%, compared to 46.0% for the control specimens; and on R. placenta, the treated specimens showed a loss of 35.1%, compared to 48.8% for the control specimens. These results clearly show that these concentrations provided a slight improvement in the resistance of the wood to fungi.

Treatment with EO

The results presented in Table 14 highlight that mass loss of wood blocks impregnated by C. schweinfurthii EO, following the exposition to C. puteana was less than 1.1%, much lower than the mass loss of control wood blocks, clearly demonstrating that the EO has greatly improved the resistance of impregnated wood blocks, providing total protection against this fungus.

It can also be observed that the mass losses of the treated wood blocks exposed to P. sanguineus and T. versicolor were less than 6.0% and 18.0%, respectively, which indicates that the C. schweinfurthii EO significantly improved the durability of the impregnated wood blocks against the fungal strains tested, with the exception for the sample exposed to R. placenta.

Treatment with PR

Table 15 shows that the mass losses of the wood blocks treated with PR of C. schweinfurthii exposed to the four fungal strains studied were greater than 9.2% for the low solution concentrations (1%, 5% and 10%) and that these were lower than the mass losses of the control specimens. Thus, it can be stated that these concentrations increased the conferred durability of the impregnated wood blocks.

For treatments involving 20% PR solutions, the mass losses of the wood blocks impregnated against T. versicolor, P. sanguineus and C. puteana are less than 13.6%, much lower than the mass losses registered with the control wood blocks. This result indicates that those PR fraction greatly increased the durability of the impregnated wood blocks. Two interesting mass loss values of 3.4% and 5.9% were obtained against C. puteana and P. sanguineus, respectively, using 20% PR. R. placenta remained the most virulent fungus compared with the other fungal strains tested against PR-impregnated wood blocks.

Resistance to termites

Test with RR

Results from Table 16 shows that the control wood specimens (i.e., wood samples impregnated with ethanol and not impregnated) with ethanol and not impregnated) were not resistant to termite attack, with mass losses exceeding 11.0% for the untreated control scots pine samples and over 14.0% for ethanol-treated control samples, not to mention termite survival rates ranging from 71.6% to 76.3%, and a visual rating of 4 for all the tested control samples.

Before leaching, the scots pine specimens impregnated with solutions containing 5% or more of RR showed a marked improvement in the resistance to termites, with very low mass losses (WLterm < 2.7%), almost complete termite mortality (TSR ≤ less than or equal to 2.0%) and a visual rating of 1.

After leaching, the improvement in the durability of treated specimens with RR fraction with respect to termites decreased slightly but remains significant, compare to the untreated wood.

Indeed, the RR fraction from C. schweinfurthii performed well in improving the durability of wood blocks at a concentration of 20%, with a mass loss of 2.5% due to termite attack, a termite survival rate of 8.7% and a visual rating of 2.

Test with EO

According to Table 17, before leaching, scots pine specimens treated with C. schweinfurthii EO showed very high resistance to termites, with virtually no weight loss (WLterm = 1.5%), complete termite mortality (TSR = 0%) and a visual rating of 0 for the single concentration of impregnation solution.

After leaching, the effect provided by the EO decreased completely, with a mass loss of 7.7% due to termite attack, a termite survival rate of 58.0%, and a visual rating of treated samples of 4.

Test with PR

Table 18 shows that, before leaching, mass losses recorded after termite exposure of wood blocks impregnated by C. schweinfurthii PR were very low (WLterm < 3.0%), termite mortality was low (TSR < 18.0%) and the visual rating of treated samples ranged from 1 to 2. This indicated that the PR fraction from C. schweinfurthii greatly increased the resistance of the wood specimens tested to termites, for impregnation solution concentrations of 5% and above.

After leaching, the effect of PR diminishes slightly, but remained measurable. A mass loss due to termite degradation of 2.5%, a termite survival rate of 2.0%, and a visual rating of 2 were observed on the wood samples impregnated by PR at a concentration level of 20%.

These results obtained through this work are consistent with those previously described for other resin species belonging to the Burseraceae family and the volatile fraction of Canarium schweinfurthii [15, 27, 49, 50].

The extraction yields of Canarium schweinfurthii differ from those reported in the literature due to various reasons, such as the state of the resins (present resins were still fresh, almost liquid, differing from those available in local market), the type of distillation apparatus used, and the quantities distilled [27].

Some past studied [51,52,53] agree with this result, emphazing that the yield, physical characteristics, and chemical composition of essential oils can be influenced by factors, such as plant species, harvesting period, plant age, moisture content of the plant material, the part of the plant used for distillation process, and the extraction technique employed to obtain the different resin or essential oil fractions.

Variations observed in the chemical composition of the three oleoresin fractions from Canarium schweinfurthii, both in quantity and quality, could be attributed to several factors, including ecological parameters, plant species, and genetic make-up, as noted in the literature [54,55,56].

The chemical composition of the oleoresins and essential oils (EOs) from certain plants can vary within the same species. These chemical variants are commonly referred to as chemotypes. Such variations can occur between populations or even among individuals an may result from exogeneous factors, such as sunlight, soil type and composition, temperature, altitude, etc. as well as endogenous factors like genetic composition [25].

This implies that substantial variability is possible in oleoresins and EOs. Further research is needed to determine whether differences in EO composition stem from genetic variation within the species, the developmental stages of trees, infections, or environmental factors [57].

Additional parameters may partly explain this variability, such as the type of equipment used and the detection threshold of the compounds analyzed. For instance, prior studies identified all compounds present at levels as low as 0.001%, whereas in this present study, the limited detection was 0.1%, focusing on the chemical valorization of oleoresin compounds [27].

Another study conducted on Myristica fragrans oil revealed that myrcene, β-pinene, sabinene, α-pinene, α-thujene and limonene induced termite mortality at a dose of 1 mg/g [58]. Extractives from Madhuca heartwood have shown anti-termite activity against Coptotermes gestroi, attributed to the presence of γ-terpinene, terpinene-4-ol, eicosan and p-cymene [7]. γ-terpinene and p-cymene have been identified as key contributors for the antitermite activity properties of E. camaldulensis against C. formosanus [59]. According to Carson and Riley (1995) [60] and to Inouye et al. (2001) [61], chemical compounds such as α-phellandrene, sabinene, α-pinene, and terpin-4-ol possess antibacterial and antifungal activities. In addition, a mixture of α- and β-amyrin was reported to have antifungal properties [62]. Past study from Cosentino et al. (1999) [63] indicated that essential oils are rich in phenols are generally more effective toward fungal exposure. However, phenols are not the solely molecules responsible for all the biological activities observed. The global chemical composition of the concerned essential oil must be considered, which may explain the efficacy of the essential oil from Canarium schweinfurthii on termites. The antitermite and antifungal activities of C. schweinfurthii EOs can be attributed to their terpene- and terpenoid-rich chemical profiles. Synergy among these compounds likely plays a significant role in their observed efficacy.

According to Eaton and Hale (1993) [64], the disruption of fungal membranes typically leads to the microroganism death. In addition, Voda et al. (2003) [65] highlighted the antioxidant properties of terpenoid compounds, which inhibit the growth of wood-destroying fungi (Trametes versicolor and Coniophora puteana) by trapping free radicals. However, another study carried out by Zhang et al. (2016) [66] noted that the antifungal efficacy, mainly against white rot, of monoterpenes and triterpens, such as α-pinene, γ-terpinene contained in the oleoresins from C. schweinfurthii, is lower compared to phenolic compounds, such as carvacrol, thymol and eugenol. This level of improvement in decay resistance of wood samples is significantly reduced for leached wood blocks. Similar findings have been reported regarding the antifungal activity of various EOs against wood-degrading fungi and their potential applications in wood protection systems [67].

Idris EA (2007) [68] and Lahlou (2004) [69] have shown that essential oils are more active against microorganisms than their major constituents tested individually, and this is true whatever the species tested. The synergetic interaction of all the compounds in the oleoresins from C. schweinfurthii plays a major role in the efficacy observed against termites. In view of their termicidal, rather than fungicidal, properties, the selected oleoresins are being studied in greater depth with a view to leveraging their termicidal potential in the field of wood protection systems on an industrial scale.

The antitermite and antifungal activities of RR and PR from C. schweinfurthii can also be attributed to their chemical composition, which is rich in terpenes and terpenoids [15], such as α-pinene and γ-terpinene, whose anti-termite activities have been well-documented. Numerous examples in the literature highlight that terpene elicit behavioral response in subterrean termites [70]. For instance, Zhu et al. (2010) [71] have shown that terpenoid compounds act as repellents, antifeedants, wood protectants and protozoal inhibitors, against termites.

According to Yoshimura et al. (1992) [72], this process can reduce the vigor of the termite colony and ultimately diminish the colony’s ability to effectively attack wood. Furthermore, Piskorski et al. (2009) [73] established that certain terpene are associated with secretions from the frontal glands of soldier termites and are emitted as repellents to other nestmates during dispersal flight. Finally, some authors noted that α-pinene has a strong antioxidant activity [74].

According to Tondi et al. (2013) [75], the impregnation solution follows the natural sap circulation pathways existing in the wood before they are obstructed, which explains the effective penetration of the different tested fractions in beech and scots pine wood samples.

Mass gains appear to depend on the nature of the resin in the impregnation solution and on the nature of the treated wood: scots pine sapwood tends to impregnate more easily than beech wood.

The differences observed between pine and beech WPGs are likely linked to the different anatomical structures of these two wood species. Scots pine, being a resinous tree, facilitates the absorption of resin fractions due to the presence of tracheids, which serve as pathways for resin in its exudates forms. In contrast, beech, with its larger vessels, is almost exclusively penetrated longitudinally. Tondi et al. (2013) [75] showed through microscopic analysis that tannin resin solutions penetrate scots pine longitudinally via tracheids with open-edged pits and radially through parenchymal rays, whereas beech is almost exclusively penetrated longitudinally via wide, easily accessible vessels.

The greater susceptibility to leaching of EO-treated wood blocks compared to RR-treated and PR-treated blocks might be explained by the solubilization of certain monoterpene alcohols in water during the removal of essential oil compounds. In addition, the drying stage following the leaching process may function as a steam distillation stage, leading to the total loss of volatile compounds from the essential oils.

The chemical composition of the different fractions influences leaching. Fractions rich in terpene alcohols and terpenes (volatile, water-soluble compounds) are more leachable than those containing fewer volatile compounds.

The improvement in the durability properties of treated wood blocks is attributed to the chemical properties of the different fractions, which are rich in monoterpenes, terpene alcohols and triterpenes [15]. The synergetic interaction between these molecular families results in enhanced effects.

Although these concentrations, in oleroresins within impregnated formulations, is slightly too high to be practical for antifungal treatments applied to wood materials, it remains interesting, because it could confer additional benefits, such as improved dimensional stability.

This work is part of the development of continuing education in the forestry and wood industry in Central Africa, as well as the promotion of natural products derived from renewable plant raw materials. These materials have the potential to replace chemical products derived from fossil resources, which are commonly used in wood preservation and are criticized for their harmful effects on humans and the environment.

In the present work, the chemical composition of C. schweinfurthii oleoresins has been analyzed using Perking Elmer Clarus 680 SQ8 gas chromatography–mass spectrometry, in three forms (fractions): raw resin (RR), purified resin (PR) and essential oils (EO). The results reveal very interesting quantitative and qualitative chemical variability within each fraction. The RR and PR studied are primarily composed by triterpenes, which represent the majority of compounds, followed by monoterpenes and monoterpenoids. On the other hand, EO exclusively consist in monoterpenes and monoterpenoids.

The essential oil of C. schweinfurthii has demonstrated notable antifungal activity and strong anti-termite activity. In addition, it has shown a marked improvement in the resistance of impregnated wood samples to both termites and fungi. These activities are all dose-dependent. The essential oil of C. schweinfurthii mainly consists of β-phellandrene, o-cymene, y-terpinene, a-terpinene, 2-menthene and terpinen-4-ol. The other two fractions of the resin, namely, the raw resin (RR) and the purified resin (PR), exhibited moderate antifungal activity but high antitermite activity. These fractions also significantly enhanced the durability of wood blocks against termites and three fungal strains. PR is composed of both monoterpenes and triterpenes (α-amyrin and β-amyrin), whereas RR contains only triterpenes (α-amyrin and β-amyrin), which are heavy molecules known for their anti-inflammatory properties. Despite the reduced effectiveness of the treated wood specimens after leaching, an increase in the biological resistance of the wood blocks was still observed.

The authors thank the National Agency of Grants and Internship of Gabon for the PhD grant allocated to Walter Fiacre Bedounguindzi. LERMAB is supported by a Grant overseen by the French National Research Agency (ANR) as part of the "Investissements d’Avenir" program (ANR-11-LABX-0002-01. Lab of Excellence ARBRE). The authors also gratefully acknowledge the GDR 3544 "Science du Bois" for its financial support for the STSM attributed to Walter Fiacre Bedounguindzi to carry out termite resistance tests at the Research Unit BioWooEB, CIRAD.

Authors and Affiliations

1. Institute of Pharmacopoeia and Traditional Medicine, IPHAMETRA, BP: 1156, Libreville, Gabon

   Walter Fiacre Bedounguindzi

2. University of Lorraine, Inrae, LERMAB, 54000, Nancy, France

   Walter Fiacre Bedounguindzi, Stephane Dumarçay & Philippe Gerardin

3. CIRAD, UPR BioWooEB, 34398, Montpellier, France

   Walter Fiacre Bedounguindzi, Kevin Candelier & Marie-France Thevenon

4. BioWooEB, Univ. Montpellier, CIRAD, Montpellier, France

   Kevin Candelier & Marie-France Thevenon

5. LAPLUS, Pluridisciplinary Science Laboratory, Ecole Normale Superieure of Libreville, BP 17009, Libreville, Gabon

   Prosper Edou Engonga

Corresponding author

Correspondence to Walter Fiacre Bedounguindzi.

Competing interests

All authors certify that they have no affiliations with or involvement in any organization or entity with any financial interest or non-financial interest in the subject matter or materials discussed in this manuscript.

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