Characterization of the bark of Plathymenia reticulata Benth. from the Brazilian Cerrado in the context of bioprospecting natural products for multiple uses

2.1 Location and sampling

Bark samples of P. reticulata were collected in Brazil from a Cerrado stricto sensu area located at Albino Farm, Coração de Jesus, in northern Minas Gerais (16°41′07″ S, 44°21′54″ W; altitude: 760 m) under SISBIO license no. 85831 and SISGEN registration no. A0B9C7E. The climate of the region is classified as Aw (tropical savanna) according to Köppen, with a dry summer lasting approximately three months.

Sampling was performed on five trees with 17–23 years of age, an height of 4–6 m and a diameter at breast height (DBH, measured at 1.3 m) of 12–17 cm. Bark was carefully removed from standing trees in strips of approximately 15 × 15 cm at breast height, taking care to avoid injury to the vascular cambium or xylem tissue.

2.2 Cellular structure characterization of bark

Preparation of microscopic slides was conducted following the methodology of Araujo et al. (2020). Transverse sections were obtained from the phloem region of five individuals, impregnated with polyethylene glycol (PEG) 1,500, and sectioned using a Leica SM 2,000 sliding microtome (USA). Fiber dimensions (length, diameter, wall thickness) and sieve tube elements were measured from macerated material following Franklin’s method (1945). Microscopic images were acquired with an Olympus EP50 camera mounted on an Olympus CX23 microscope. Anatomical terminology and classification followed IAWA standards for bark structure (Angyalossy et al. 2016).

2.3 Sample preparation for chemical analyses

Bark samples from five trees were combined to create a composite sample. The phloem was carefully separated from the cork using a chisel. Both the phloem and the whole bark were ground in a hammer mill and fractionated by sieving. The material retained on a 60 mesh sieve was used for distinct analytical procedures. For summative chemical characterization, only the phloem was used. For the preparation of hydroalcoholic extracts, the composite sample containing the entire bark (phloem and cork) was employed.

2.4 Chemical characterization

Extractives were quantified following TAPPI T 204 cm-97 (1997). Triplicate samples of 2 g (dry basis) underwent sequential extraction in a Soxhlet system using dichloromethane (6 h), ethanol (16 h), and water (16 h). The resulting samples (∼1.5 g dry basis, duplicate) were analyzed for suberin content using sodium methoxide depolymerization (Pereira 1988). Insoluble lignin was quantified in triplicate from 0.35 g (dry basis) using 72 % sulfuric acid, following TAPPI T 222 om-02 (2002). Soluble lignin was determined from filtrate using TAPPI UM-250 (1991), with absorbance read at 205 nm using a Shimadzu UV-1601 spectrophotometer (Japan).

Ash content and mineral composition were determined from samples retained on the 60 mesh sieve. Triplicate samples of 2 g (dry basis) were combusted at 550 °C for 4 h (TAPPI T 211 om-02, 2002). Elemental composition was obtained using an inductively coupled plasma optical emission spectrometer (ICP-OES; Germany). Holocellulose content was estimated by subtracting the mean values for extractives, lignin, suberin, and ash from 100 %.

2.5 Total phenolics, flavonoids, and condensed tannins

Dried bark samples (0.5 g) were extracted using ethanol, methanol, and acetone (1:1 v/v with deionized water) in 20 mL total volume via ultrasound (Quimis ISO 9001, Brazil) at 60 °C for 1 h. Each extract was diluted to 50 mL and analyzed in triplicate for total phenolics, flavonoids, and condensed tannins following Miranda et al. (2016).

Total phenolics were quantified by mixing 100 µL of extract with 4 mL of 10 % Folin–Ciocalteu reagent; after 8 min, 4 mL of 7.5 % Na₂CO₃ was added and incubated at 45 °C (Singleton and Rossi 1965). Absorbance was read at 765 nm and expressed as mg gallic acid equivalents (GAE) per g of bark extract.

Flavonoid content was determined using aluminum chloride (Zhishen et al. 1999), and condensed tannins via the vanillin-H₂SO₄ method (Abdalla et al. 2014), both calibrated against catechin standards (50–500 mg/mL). Readings were taken at 510 and 500 nm, respectively, using a Biospectro SP-22 spectrophotometer (Brazil). Results were expressed as mg catechin equivalents (CE) per g of bark extract.

2.6 Identification and quantification of bioactive compounds

Bioactive compounds from ethanol, methanol, and acetone extracts were profiled using high-performance liquid chromatography (HPLC). Standards included trigonelline, gallic acid, theobromine, catechin, chlorogenic acid, caffeic acid, syringic acid, vanillin, p-coumaric acid, ferulic acid, m-coumaric acid, o-coumaric acid, resveratrol, rosmarinic acid, and trans-cinnamic acid. Extracts were filtered through 0.45 µm syringe filters prior to injection.

Analysis was performed on a Shimadzu^® LC system equipped with LC-20AT pumps, SPD-M20A diode array detector, DGU-20A5 degasser, CBM-20A interface, CTO-20AC column oven, and SIL-20A autosampler. Separation used a Shim-pack GVP-ODS-C18 column (4.6 × 250 mm, 5 μm) with a pre-column (4.6 × 10 mm, 5 μm). Eluents followed Lorenço et al. (2021): Phase A (2 % acetic acid in ultrapure water), Phase B (methanol:water:acetic acid at 70:20:2 v/v). Conditions included 280 nm wavelength, 1.0 mL/min flow rate, 35 °C column temperature, and 20 μL injection volume. Compound concentrations were determined using specific linear calibration equations for each standard.

2.7 Evaluation of condensed tannin content

Whole bark (phloem and cork) from the five individual trees was homogenized using a hammer mill. Tannins were extracted in triplicate from 100 g (dry basis) using 3 % aqueous sodium sulfite (15:1 solution:bark ratio) in a water bath at 70 °C for 3 h (da Silva Araujo et al. 2021). Filtrate was concentrated to 150 mL at 70 °C. Duplicate samples were used to determine total solids (TS), Stiasny index (SI), and condensed tannin content (CT).

TS was calculated from 20 g of extract dried at 103 ± 2 °C to constant mass. SI determination involved heating 20 g of extract with 10 mL water, 2 mL 10 N HCl, and 4 mL 37 % formaldehyde under reflux for 35 min after boiling. Precipitate was filtered using a sintered glass funnel (porosity 2), dried at 103 ± 3 °C, and weighed. SI was calculated as: SI (%) = (dry mass of precipitate/total solids in 20 g extract) × 100 CT content was calculated as: CT (%) = (SI × TS)/100.

Non-tannic components (NT) were estimated by subtracting CT from TS (da Silva Araujo et al. 2021).

2.8 Tannin characterization

Tannins obtained as described in Section 2.7 were dried at 40 °C, macerated, and sieved to 200 mesh prior to analysis by Raman spectroscopy, ATR-FTIR, and thermogravimetric analysis (TGA).

2.9 Statistical analysis

Experimental data obtained for the extracts obtained with the different solvents (ethanol, methanol, and acetone) were subjected to analysis of variance (ANOVA) using SPSS software version 21. A significance level of 5 % (p < 0.05) was adopted. Post hoc comparisons were performed using the Tukey HSD test. Results are expressed as mean values ± standard deviations from three independent measurements.

3.1 Anatomical structure

The bark of P. reticulata is distinctively constituted by the phloem inner bark and an outer bark with a well-developed single cork periderm that showed deep longitudinal fractures resulting from tree diameter growth. The periderm develops continuously around the stem with a substantial content of cork, without forming a rhytidome, indicating that its lifespan corresponds to the age of the sampled trees. This is similar to a few other cork-rich species namely to Q. suber (Leite and Pereira 2017).

The phellem (cork) is non-stratified and well developed, exhibiting a radial alignment of thin-walled cells with an average 20.6 μm radial dimension, and growth rings marked by reduced radial cell dimensions to an average 12.2 μm (Figure 1A and B). The development and cellular features of the cork resemble those observed in the first cork layer of Q. suber (Mota et al. 2016). The phelloderm forms a narrow layer comprising 2–3 radially aligned, thick-walled cells (Figure 1B). The phellogen, the meristem responsible for producing both phellem outwardly and phelloderm inwardly, is composed of thin-walled, rectangular cells (Figure 1B). Both phellem and phelloderm cells contain dark contents, presumably phenolic compounds (Figure 1B and C).

Figure 1:

Cross-section of the periderm and phloem of Plathymenia reticulata. (A) Periderm with phellem [Pm] showing a growth ring marked by the smaller radial dimension of the cells [black arrows] and phloem [Ph] with a ring of sclerified cells below the periderm [arrow]. (B) Periderm with phellem [Pm], phelloderm [Pd], phellogen [black arrow] and cells with contents [red arrow]. (C) Non-conductive and conductive phloem [red star], fibres [F], dilatation tissue [Dt], ray [R], cells with contents [red arrow], collapsed tube element [black arrow] and canals [*]. Scale bars: (A and C): 100 μm; (B): 25 μm.

The inner bark is composed of secondary phloem, which includes a conductive and a non-conductive zone, separated by a gradual transition (Figure 1C). The conductive phloem, located near the vascular cambium, represents a narrow portion of the total phloem and comprises rays (transverse conduction and storage), fibers (mechanical support), sieve tubes and companion cells (conductive tissue), and axial parenchyma. Sieve tube elements possess compound sieve plates, averaging 12 per element. Their tangential diameters range from 12 μm to 26 μm, with average lengths between 300 μm and 389 μm. Companion cells are visible in both transverse and longitudinal sections (Figure 2A–C). Fibers are thick-walled, lignified, and often associated with crystals, with an average length of 897 μm and a wall thickness of 20 μm (Figure 2B and F). Rays are uniseriate and multiseriate (2–3 cells wide), homocellular and composed of procumbent cells (Figure 2F and G).

Figure 2:

Transverse and longitudinal sections of the secondary phloem of Plathymenia reticulata: transverse sections (B–E), tangential sections (A and G) and radial sections (F). (A) Sieved area of tube element [red arrow] and companion cells [black arrow]. (B) Tube element with sieved plates [black arrow], companion cells [*] and crystals adjacent to the fibres [red arrow]. (C) Tube element with sieved plates [red arrows] with companion cells [black arrow]. (D) Canal [*] and cells with contents [red arrow]. (E) Scar tissue, sclerified cells (S), cells with contents [red arrows] and tube elements [black arrow]. (F) Rays [R] and fibres [F] bordered by a series of crystalliferous cells. (G) Simple rays [red arrow] and compound rays [black arrow]. Scale bars: (A–D): 25 μm; (E–G): 100 μm.

The non-conductive phloem is initiated by the collapse of sieve elements and development of dilatation tissue via division and expansion of parenchyma cells, primarily the axial parenchyma (Figure 1C). This response accommodates the radial growth of the stem (Angyalossy et al. 2016). Non-conductive phloem cells exhibit dense phenolic deposits (Figures 1C and 2D). Parenchyma cells in the outer phloem are often sclerified, forming an irregular ring of sclereids adjacent to the periderm, similar to structures observed in other Fabaceae species such as Anadenanthera peregrina and Anadenanthera colubrina (Mota et al. 2017).

Secretory channels are present throughout the secondary phloem (Figures 1C and 2D). The fresh bark samples release a reddish-orange sticky substance. Scar tissue, characterized by intense cell division, numerous sclerified cells, and cells rich in secondary metabolites, was observed in recently formed regions of the secondary phloem likely as a response to mechanical injury (Figure 2E). Bark serves a key defensive role by storing secondary metabolites that are induced as phytoalexins in response to biotic stress such as insect or pathogen attack (War et al. 2012).

3.2 Summative chemical characterization

The chemical composition of phloem and cork composite samples is presented in Table 1. Substantial differences were observed between the two bark fractions, particularly in extractives, suberin, ash, and polysaccharide contents. The phloem showed a particularly high content of extractives (40.0 %) for which ethanol- and water-soluble polar compounds contributed most, accounting for 67.5 % and 27.0 % of total phloem extractives. Phloem exhibited approximately 3.5 times more extractives than the cork (12.7 %).

Non-polar extractives obtained via dichloromethane accounted for 3.4 % in P. reticulata cork. This value is lower than values reported for Q. suber cork (5.8 %; Pereira 2013), a commercially exploited temperate species used here as reference, and also lower than those found in other tropical hardwoods from the Cerrado biome, such as Kielmeyera coriacea (5.2 %; Rios et al. 2014), Agonandra brasiliensis (4.8 %; Silva et al. 2021), and Erythrina mulungu (6.9 %; da Silva Mota et al. 2025). However, it exceeds the value reported for the tropical Enterolobium gummiferum cork (1.7 %; da Silva Mota et al. 2025), placing P. reticulata cork in an intermediate position among Cerrado species in terms of non-polar extractive content.

Cork samples of P. reticulata also revealed substantial suberin content (24.7 %) although lower than that of Q. suber cork that averages 42.8 % (Pereira 2013). Suberin levels vary considerably among tropical hardwoods from Cerrado. Lower contents were reported for corks of E. mulungu (4.7 %) and E. gummiferum (3.8 %; da Silva Mota et al. 2025), as well as of A. brasiliensis (5.7 %; Silva et al. 2021). In contrast, K. coriacea cork exhibited a suberin content equal to that of P. reticulata (24.7 %; Rios et al. 2014).

Lignin levels were notably high in both bark fractions: 40.6 % in the phloem and 34.5 % in the cork. These values exceed those of commercial Q. suber cork (22.0 %) and fall below reported lignin contents in K. coriacea (45.0 %) and A. brasiliensis (45.6 %).

A key compositional difference was also noted in polysaccharide content: 10.8 % in phloem and 20.9 % in cork. This difference was also found in other species. In Q. suber, phloem had 49.1 % polysaccharides and cork 16.2 % (Costa et al. 2019) and in Douglas-fir bark 43.3 % and 48.1 % in phloem and 10.0 % and 7.3 % in cork (Cardoso et al. 2018).

3.3 Inorganic elemental composition

Ash content also varied significantly between fractions, with 8.6 % in phloem and only 0.8 % in cork. Similar patterns have been reported in Quercus cerris, which displays 13.0 % ash in phloem and 2.6 % in cork (Şen et al. 2010), and in Douglas-fir bark (1.4–2.1 % in phloem vs. 0.2–0.3 % in cork; Cardoso et al. 2018).

Nitrogen was the most abundant element (58.1 %), followed by calcium (18.1 %) and potassium (8.6 %) (Table 2). These elements play vital roles in the metabolic functions of living organisms and underscore the bark’s nutritional potential.

Calcium abundance may be linked to the crystals observed in axial parenchyma cells (Figure 2C). High levels of nitrogen, calcium, and potassium reinforce the potential of P. reticulata bark as a source of bioelements and as a promising component in plant growth substrates (Sousa et al. 2022).

In a biorefinery context, following extraction of high-value compounds such as tannins, the remaining biomass may be repurposed as fertilizer, contributing to sustainable resource management. The macronutrient composition of P. reticulata bark closely resembles that of Stryphnodendron rotundifolium, where nitrogen also predominates (50.15 %), followed by calcium (24.07 %) and potassium (16.55 %) (Sousa et al. 2021).

Micronutrient concentrations in P. reticulata bark included iron (117.45 mg/kg), magnesium (65 mg/kg), manganese (32.84 mg/kg), phosphorus (21.00 mg/kg), and zinc (18.38 mg/kg). Despite being required in smaller quantities, these elements are essential for regulating key biological processes across diverse organisms.

3.4 Composition of bark extract

The phenolic composition of P. reticulata bark polar extracts is presented in Table 3. The total phenolic content was relatively low, with values of 34.36, 34.49, and 53.62 mg GAE g⁻¹ of extract for ethanol, methanol, and acetone extracts, respectively. Similarly, the contents of flavonoids and condensed tannins were also low. Aqueous acetone proved to be the most effective extraction solvent, yielding higher concentrations of phenolics, flavonoids, and tannins compared to aqueous ethanol and aqueous methanol. The total phenolic content in the acetone extract (53.62 mg GAE g⁻¹) was significantly greater and statistically distinct from the ethanol (34.36 mg GAE g⁻¹) and methanol (34.49 mg GAE g⁻¹) extracts. However, the values for total flavonoids and condensed tannins showed no statistically significant differences among the solvents used.

The extraction efficiency of bioactive compounds, such as total phenols, from P. reticulata bark is influenced by the polarity of the solvent system. The combination of water and organic solvents provides a medium of intermediate polarity, ideal for the extraction of phenolics due to variability in chemical polarity (Jdaini et al. 2023). Aqueous acetone is considered a green solvent suitable for food-grade applications (Agarwal et al. 2021).

In comparison with other tropical hardwood species, the total phenolic content of P. reticulata bark was higher than the 4.06 mg GAE g⁻¹ reported for Moringa oleifera (Vats and Gupta 2017), but lower than the values reported for M. eximia (242.4 mg GAE g⁻¹) and A. peregrina (583 mg GAE g⁻¹) and A. colubrina (682 mg GAE g⁻¹) (Araujo et al. 2020; Mota et al. 2017). In a broader context bark of temperate species, such as black locust (35.93 mg GAE g⁻¹) and pedunculate oak (105.88 mg GAE g⁻¹) also exhibit lower phenolic levels than the aforementioned tropical species (Agarwal et al. 2021). These comparisons highlight the moderate phenolic profile of P. reticulata, which may be advantageous in biorefinery contexts targeting polysaccharide recovery, where lower phenolic interference is desirable, as high levels of phenolic compounds can hinder the conversion of biomass into fermentable sugars, compromising the yield and quality of biofuels and biomaterials (Yao et al. 2022).

The flavonoid content (7.15–7.26 mg CE g⁻¹) exceeded that of M. oleifera bark (1.47 mg CE g⁻¹; Vats and Gupta 2017) but was markedly lower than values found in M. eximia (300.8 mg CE g⁻¹), A. peregrina (148.4 mg CE g⁻¹), and A. colubrina (145.3 mg CE g⁻¹) (Araujo et al. 2020; Mota et al. 2017).

Condensed tannin levels (9.58–9.97 mg CE g⁻¹) in P. reticulata bark were also considerably lower than reported values for M. eximia (877.3 mg CE g⁻¹), A. peregrina (586.9 mg CE g⁻¹), and A. colubrina (97.5 mg CE g⁻¹) (Araujo et al. 2020; Mota et al. 2017).

3.5 Identification and quantification of bioactive compounds

Among the secondary metabolites identified in P. reticulata bark extracts, four major compounds were quantified: trigonelline, theobromine, gallic acid, and caffeic acid. Extraction efficiency varied depending on the solvent employed (Figure 3A). Trigonelline showed its highest concentration in the methanol extract (192.71 mg/100 g), followed by ethanol (101.28 mg/100 g), while acetone yielded the lowest amount (25.61 mg/100 g). Conversely, gallic acid reached its peak concentration with acetone (350.14 mg/100 g), compared to much lower levels in methanol (2.95 mg/100 g) and ethanol (1.82 mg/100 g).

Figure 3:

Bioactive compound concentrations and characterization parameters of Plathymenia reticulata bark extracts. (A) Concentration of bioactive compounds (Trigonelline, Gallic acid, Theobromine, and Caffeic acid) extracted using ethanol, methanol, and acetone. (B) Average values for total solids yield (TSY), Stiasny index (SI), condensed tannin yield (CTY) and non-tannic compounds (NTY).

Theobromine was most abundant in the methanol extract (46.73 mg/100 g), followed by ethanol (17.01 mg/100 g) and acetone (7.43 mg/100 g). Caffeic acid was also extracted most efficiently with methanol (21.79 mg/100 g), followed by ethanol (10.58 mg/100 g) and acetone (3.77 mg/100 g). These findings highlight methanol as the most effective solvent for recovering a broad spectrum of bioactive compounds, although acetone was superior for gallic acid extraction.

Trigonelline, an alkaloid derived from nicotinic acid, offers multiple health benefits, including glycemic control, reduction of LDL cholesterol, and elevation of HDL cholesterol (Subramanian and Prasath 2014). Its antihyperglycemic activity is associated with inhibition of dipeptidyl peptidase-4 (DPP-4) and α-amylase enzymes (Gilbert and Pratley 2020). Additionally, trigonelline is linked to improvements in central nervous system function and gastrointestinal motility (Mathur and Kamal 2012).

Theobromine, a methylxanthine structurally related to caffeine, exhibits neuroprotective, anti-inflammatory, and antioxidant properties. It has been associated with weight management, kidney stone prevention, and enhancement of cognitive and metabolic functions (Zhang et al. 2024). Moreover, its antimicrobial activity against Streptococcus mutans, Lactobacillus acidophilus, and Enterococcus faecalis has been demonstrated (Lakshmi et al. 2019).

Phenolic acids such as gallic acid and caffeic acid are potent antioxidants with anti-aging, anti-tumor, antimicrobial, and anti-inflammatory effects (Pei et al. 2016). Widely used in cosmetics and sunscreens, these compounds protect against UV-induced skin damage (Abdelkader Bensid et al. 2022). Gallic acid, in particular, demonstrates superior antioxidant efficacy and has been identified in bark extracts of several Amazonian species including M. eximia, Astronium lecointei, and Protium tenuifolium (Araujo et al. 2020; de Jesus dos Santos et al. 2024; Mota et al. 2021).

3.6 Condensed tannin content

Condensed tannin yields (CTY) from P. reticulata bark are illustrated in Figure 3B. The extraction yielded 19.0 % tannins, surpassing the yields from Eucalyptus urophylla hybrid clones (1.37 % and 3.41 %) using sodium sulfite (Sartori et al. 2018). This value is comparable to that of other tropical hardwood species, such as A. colubrina bark (19.2 %; Paes et al. 2013), but lower than the 32.6 % reported for M. eximia (da Silva Araujo et al. 2021). In a broader context, A. mearnsii, a species native to subtropical and temperate regions and widely cultivated for commercial tannin production, presents a tannin content of 30.0 % (Mangrich et al. 2014).

The Stiasny index (SI) of 83.1 % recorded here exceeds values reported for A. colubrina (64.5 %; Paes et al. 2013) and E. urophylla hybrids (28 % and 45 %; Sartori et al. 2018), though it remains slightly below that of M. eximia bark (93.3 %; da Silva Araujo et al. 2021). High SI values are desirable as they indicate the fraction of tannins suitable for polymerization in adhesive production. The SI estimates the proportion of polyflavonoids reactive with formaldehyde (Matsumae et al. 2019), making it a crucial metric for industrial applications of tannin-based adhesives.

3.7 Characterisation of Plathymenia reticulata tannins

Raman spectroscopy data for the tannins are shown in Figure 4A. The spectrum reveals multiple characteristic peaks, including bands between 451 and 985 cm⁻¹ corresponding to Na₂SO₃ stretching vibrations, derived from the sodium sulphite extraction solution. Peaks observed at 1,333, 1,423–1,482, and 1,566 cm⁻¹ are associated with tannin structures, specifically reflecting B-ring and C–C stretching, A-ring and C–OH stretching, and C=C stretching in the A and B rings, respectively. These spectral features are consistent with observations reported by Reyer et al. (2016) and Zidanes et al. (2021).

Figure 4:

Spectroscopic and thermal analysis of tannins extracted from Plathymenia reticulata bark. (A) Raman spectrum of the tannin sample. (B) ATR-FTIR spectrum of the tannins. (C) TGA (black line) and DTG (red line) curves of the tannins.

The ATR-FTIR spectra of tannins extracted from P. reticulata bark are presented in Figure 4B. The spectra exhibit prominent bands indicative of hydroxyl groups, aliphatic chains, and aromatic structures. Key absorption bands include 3,241 cm⁻¹ (O–H stretching), 1,604 cm⁻¹ (C=O stretching), 1,519 cm⁻¹ (N–H bending), 1,440 cm⁻¹ (CH₂ bending), 1,334 cm⁻¹ (CH₃ bending), 1,037 cm⁻¹ (C–O stretching), and approximately 900 cm⁻¹ (C–H deformation in aromatic rings). These observations confirm the presence of polyphenolic compounds and aromatic functionalities, consistent with findings by Chupin et al. (2013) and Ricci et al. (2015).

Similar FTIR profiles were reported by Sartori et al. (2018) for tannins extracted from E. urophylla bark, featuring bands at 3,500–3,000 cm⁻¹ (O–H), 1,715 cm⁻¹ (C=O), 1,610, 1,520, and 1,444 cm⁻¹ (C=C), and additional peaks at 1,286, 1,103 cm⁻¹ (C–O), and 1,138 cm⁻¹ (S=O) for extracts treated with 5 % Na₂SO₃. Zidanes et al. (2021) observed a strong O–H signal near 3,235 cm⁻¹ and a distinct peak at 840 cm⁻¹ in A. peregrina bark tannins, absent in adhesive formulations. Tannins derived from M. eximia bark (da Silva Araujo et al. 2021) exhibited bands at 1,606, 1,442, 1,328, 1,031, and 900 cm⁻¹, with a broad O–H absorption spanning 3,500–3,000 cm⁻¹. These interspecific variations reflect the chemical diversity among bark tannins and their implications for functional properties and potential applications.

Thermogravimetric analysis (TGA) of P. reticulata bark tannins (Figure 4C) revealed a structurally stable and thermally resilient profile. Initial mass loss between 25 °C and 125 °C corresponds to moisture evaporation. From 150 °C onward, degradation of residual sugars associated with tannins occurs (Rachwal et al. 2001), followed by gradual breakdown of phenolic rings beginning around 270 °C. The thermogravimetric derivative (DTG) curve highlights distinct degradation phases, illustrating the complex molecular nature of the tannins. Likewise, M. eximia tannins (da Silva Araujo et al. 2021) displayed high thermal stability with approximately 25 % residual mass at 900 °C. Although P. reticulata tannins exhibited a marked mass loss near 600 °C, their robust thermal resistance below this threshold suggests promising applications in heat-intensive industrial processes.