Zanthoxylum zanthoxyloides (Rutaceae): Ethnopharmacological Perspectives

Study design

This review was conducted as a narrative and integrative literature review, designed to systematically synthesize published research on Zanthoxylum zanthoxyloides across multiple scientific domains. Given the multidisciplinary nature of existing studies—spanning ethnobotany, pharmacognosy, phytochemistry, pharmacology, toxicology, and plant biology-a narrative review approach was considered most appropriate to allow contextual interpretation, chronological tracking of scientific progress, and thematic integration of findings.

Literature sources and search strategy

The literature included in this review was obtained exclusively from the articles and documents provided by the authors, accessed through established academic databases and repositories such as Multidisciplinary digital publishing institute (MDPI), ScienceDirect, Wiley Online Library, Thieme Connect, Hyper articles online (HAL), African journal online (AJOL), Ingenta Connect, Academia.edu, ResearchGate, Google Books, and regional scientific journals. No additional external sources were introduced beyond the supplied links to maintain consistency and transparency.

The selected sources span several decades, allowing analysis of both early foundational studies and recent advances. Emphasis was placed on peer-reviewed journal articles, authoritative reviews, pharmacognostic monographs, and experimentally driven studies relevant to Z. zanthoxyloides.

Inclusion and exclusion criteria

Exclusion criteria

• Studies addressing other Zanthoxylum species without specific reference to Z. zanthoxyloides.

• Abstract-only publications lacking methodological or result details.

• Opinion pieces or anecdotal reports without scientific validation.

Data extraction and organization

Relevant information was extracted manually from each eligible article and categorized into thematic domains:

1. Ethnobotanical and traditional uses

2. Botanical, anatomical, and pharmacognostic characteristics

3. Phytochemical composition and analytical methods

4. Pharmacological activities (antimicrobial, antiplasmodial, antioxidant, anti-inflammatory, antisickling, gastroprotective, oral health-related effects)

5. Toxicological and safety evaluations

6. Conservation, genetic diversity, and sustainability considerations

Studies were organized into thematic domains (ethnobotany, phytochemistry, pharmacology, and toxicology), with chronological trends discussed within sections where relevant to highlight the evolution of research focus over time.

Critical appraisal and comparative analysis

A qualitative critical analysis was undertaken to assess:

• Experimental design robustness (in vitro, in vivo, ex vivo, or clinical relevance).

• Sample size adequacy and model selection.

• Extraction methods, solvent systems, and phytochemical identification techniques.

• Consistency or divergence of findings across studies.

• Translational relevance and reproducibility of reported outcomes.

Comparisons were made across studies to identify recurring bioactivities, corroborated phytochemical markers, and methodological strengths. Limitations such as lack of standardization, insufficient toxicological evaluation, and absence of clinical trials were systematically noted to identify research gaps.

Ethical considerations

As this study is based solely on previously published data, no ethical approval or informed consent was required. All original authorship and sources are acknowledged through proper citation using the Vancouver referencing style.

Ethnobotanical uses and traditional knowledge

Ethnobotanical evidence consistently identifies Zanthoxylum zanthoxyloides as one of the most widely utilized medicinal plants in West Africa. Across multiple regions—including Nigeria, Ghana, Benin, Burkina Faso, Mali, and Senegal— the plant is traditionally employed for both preventive and therapeutic purposes.^[1,3,6,7] The root and stem bark are the most frequently used parts, commonly prepared as chewing sticks, decoctions, infusions, or powdered formulations.

Early ethnobotanical surveys documented the use of Z. zanthoxyloides in the management of toothache, gingivitis, and halitosis, which later became one of the most scientifically investigated traditional applications.^[6,9] Beyond oral health, the plant is traditionally prescribed for malaria, sickle cell disease, dysentery, diarrhea, gastric ulcers, rheumatism, wounds, fever, and inflammatory conditions.^[4,10] In some communities, preparations are also used as aphrodisiacs and postpartum tonics.^[11]

Comparative ethnobotanical studies reveal strong concordance in usage patterns across geographically distinct populations, suggesting a long-standing and culturally conserved therapeutic relevance.^[1,5] However, despite the breadth of traditional applications, most ethnomedicinal claims remain supported primarily by preclinical evidence, highlighting a persistent gap between traditional use and clinical validation. Ethnomedicinal uses are derived from cross-regional surveys and experimental validations reported in West African populations [Table 1].

Botanical, anatomical, and pharmacognostic characteristics

Botanically, Z. zanthoxyloides is described as a spiny shrub or small tree reaching 5–14 m in height, characterized by pinnate leaves, thorny stems, and aromatic bark.^[1,12] Pharmacognostic studies emphasize the diagnostic features of the root and stem bark, including cork cells, sclerenchyma fibers, secretory canals, and abundant calcium oxalate crystals.^[13,14]

Recent pharmacognostic evaluations comparing root and stem bark powders demonstrated minimal variation in physicochemical parameters such as moisture content, ash values, and extractive values, suggesting that stem bark could potentially substitute root bark to reduce destructive harvesting practices.^[14] This finding has important conservation implications, as root harvesting remains a major threat to wild populations.

Microscopic and macroscopic standardization studies further contribute to the establishment of quality control benchmarks necessary for herbal drug development.^[14] Nevertheless, many earlier studies lacked harmonized standards, limiting reproducibility and cross-study comparison.

Phytochemical composition

Phytochemical investigations reveal that Z. zanthoxyloides possesses a chemically diverse profile dominated by alkaloids, phenolic compounds, flavonoids, terpenoids, tannins, saponins, and essential oils.^[2,8,15] Early studies focused on qualitative phytochemical screening, while more recent research employed chromatographic and spectroscopic techniques for compound isolation and characterization.

Benzophenanthridine alkaloids, including fagaronine and chelerythrine derivatives, are among the most frequently reported bioactive constituents and are strongly associated with antimicrobial and antiplasmodial activity.^[16,17] Furoquinoline alkaloids and divanilloylquinic acid derivatives (burkinabins) have also been identified and linked to antioxidant and anti-inflammatory effects.^[1,18]

Essential oil analyses identified constituents such as α-pinene, β-caryophyllene, and caryophyllene oxide, compounds known for antimicrobial and anti-inflammatory properties.^[16] Comparative studies indicate that solvent polarity significantly influences extract composition, with alcoholic extracts generally exhibiting higher phenolic and flavonoid content than aqueous preparations.^[9,14]

Despite these advances, most phytochemical studies remain descriptive, with limited structure–activity relationship analyses, underscoring the need for targeted bioassay-guided fractionation. Phytochemical composition varies depending on plant part, geographical origin, harvesting period, and extraction solvent [Table 2].

Pharmacological activities

Antiplasmodial and antiparasitic activity

Antiplasmodial investigations demonstrated significant activity of root bark extracts and isolated alkaloids against both chloroquine-sensitive and resistant strains of Plasmodium falciparum.^[17] These findings corroborate the traditional use of the plant in malaria management and position Z. zanthoxyloides as a potential source of novel antimalarial leads. The available evidence is derived from a limited number of studies, primarily involving in vitro parasite assays and a few rodent models, with sample sizes in animal studies typically ranging from 5–10 subjects per group.^[17] Standard antimalarial agents are commonly employed as positive controls, although reporting of negative controls and experimental replication is inconsistent across studies. While antiplasmodial activity has been demonstrated across different extracts and isolated compounds, reproducibility across independent laboratories remains limited, and variability in extraction methods and assay conditions constrains direct comparison of potency. Nonetheless, most studies employed in vitro or rodent models, and pharmacokinetic data remain largely unavailable, highlighting a significant gap in translational relevance.

Antioxidant and anti-inflammatory properties

Several studies report antioxidant activity of Zanthoxylum zanthoxyloides, primarily assessed using in vitro assays such as DPPH, ABTS, and FRAP.^[8,14,21] However, the number of studies within this domain remains limited (approximately 3–5 key reports), and methodological approaches vary considerably. Among these, detailed antioxidant evaluation is provided by^[21], where assays were conducted in replicated experimental runs (typically in triplicate) using appropriate positive controls such as standard antioxidant compounds and solvent controls. Reported DPPH IC₅₀ values fall within a relatively narrow range (approximately 20.8–25.8 µg/ mL), suggesting moderate consistency under controlled experimental conditions.

In contrast, other studies cited in this domain are largely descriptive or pharmacognostic in nature,^[8,14] lacking standardized antioxidant assays, defined sample sizes, or explicit reporting of controls and replication. Consequently, cross-study comparability is limited. Anti-inflammatory effects have been inferred from reductions in inflammatory mediators and tissue damage in experimental models^[15], although these findings are based on a small number of studies with limited methodological detail and minimal independent replication.

Overall, while methanolic extracts consistently demonstrate higher antioxidant capacity than aqueous extracts—highlighting the influence of extraction methodology—the evidence base remains predominantly in vitro, with heterogeneous protocols, insufficient reporting of experimental design parameters, and absence of negative or null findings, indicating a potential risk of publication bias.

Gastroprotective and other systemic effects

Gastroprotective studies revealed that ethanolic root bark extracts significantly reduced experimentally induced gastric lesions, potentially through antihistaminic or cytoprotective mechanisms.^[10] These findings are derived from a limited number of in vivo animal studies, typically employing small sample sizes (commonly fewer than 10 animals per group) and acute ulcer models such as ethanol- or indomethacin-induced gastric injury. While some studies incorporated standard anti-ulcer agents as positive controls, detailed reporting of experimental design parameters— including randomization, blinding, and replication—is generally lacking. Additional reports suggest antisickling activity, lending support to ethnomedical use in sickle cell disease management^[22], although these findings are primarily based on in vitro erythrocyte assays with limited methodological standardization. Reproducibility across independent laboratories remains minimal, and negative or null findings are rarely reported, raising the possibility of publication bias. However, these effects are often reported in isolation, with limited replication across independent laboratories. Pharmacological activities were evaluated using diverse experimental models, predominantly in vitro assays and animal studies [Table 3].

Pharmacognostic parameters are essential for quality control and authentication of herbal raw materials [Table 4].

Toxicological considerations and safety

Toxicological evaluations of Zanthoxylum zanthoxyloides are comparatively sparse. Acute toxicity studies generally suggest a relatively wide margin of safety at commonly used doses, with no mortality observed in experimental animals at doses up to 2000–5000 mg/kg body weight following oral administration, indicating a high median lethal dose (LD₅₀).^[23,24] These findings are consistent with general toxicity profiles reported for several species within the genus Zanthoxylum, which are typically regarded as having low acute toxicity.^[25]

Despite this apparent safety in acute settings, sub-chronic and chronic toxicity data remain limited. Some studies have reported alterations in liver function markers, including elevated alanine aminotransferase (ALT) and alkaline phosphatase (ALP) levels, suggesting possible hepatobiliary or cholestatic effects following prolonged exposure.^[23,26] Histopathological evaluations, where available, indicate mild hepatic cellular changes and occasional renal alterations at higher doses, although these findings are not consistently reported across studies.^[26]

Organ-specific toxicity assessments remain insufficiently characterized. Available data suggest that while short-term administration may be relatively safe, prolonged use could pose risks to the liver and, to a lesser extent, the kidneys, particularly at higher or unstandardized doses. However, inconsistencies in dosing regimens, extract types, and experimental duration limit definitive conclusions.

Comparative evidence from related Zanthoxylum species indicates similar safety patterns, with low acute toxicity but potential for organ-specific effects at higher doses or prolonged exposure, further underscoring the need for cautious interpretation.^[25,27]

Overall, the lack of standardized dosing protocols, limited reporting of LD₅₀ values, absence of well-designed sub-chronic and chronic toxicity studies, and scarcity of human clinical safety data represent significant gaps. These limitations constrain the translational application of Z. zanthoxyloides and highlight the urgent need for comprehensive toxicological evaluation, including dose– response relationships, organ-specific toxicity profiling, and long-term safety assessment.

Synthesis of trends and research gaps

Chronological analysis reveals a progression from ethnobotanical documentation and antimicrobial screening toward phytochemical characterization, pharmacognostic standardization, and applied formulations. Despite this evolution, key gaps persist, including:

• Limited clinical trials,

• Inadequate toxicological profiling,

• Poor standardization across studies,

• Insufficient conservation-oriented research.

Critical appraisal of included studies

The body of evidence reviewed is predominantly preclinical, comprising mainly in vitro assays and a limited number of in vivo animal studies. Across pharmacological domains, the number of studies varies considerably, with antimicrobial and antioxidant activities being the most frequently investigated (approximately 10–15 studies), while antiplasmodial, gastroprotective, and antisickling activities are supported by fewer reports (typically fewer than 5 studies per domain).

Sample sizes in animal studies are generally small, commonly ranging from 5 to 10 animals per experimental group, with limited reporting of power calculations. Methodological quality indicators such as randomization, blinding, and allocation concealment are rarely described, which introduces potential bias in outcome assessment.

The use of appropriate controls varies in studies. While positive controls (e.g., standard antimicrobial or antioxidant agents) are frequently included in in vitro assays, their use in vivo models is less consistently reported. Additionally, extraction methods, solvent systems, and phytochemical standardization differ substantially between studies, limiting reproducibility and cross-study comparability.

Reproducibility across independent laboratories is evident for antimicrobial and antioxidant activities, where similar findings have been reported using different extracts and experimental conditions. However, other pharmacological effects, including antisickling and gastroprotective activities, have not been extensively replicated, reducing confidence in their generalizability.

Notably, negative or null findings are rarely reported in the literature, suggesting the possibility of publication bias. This bias may contribute to an overestimation of the pharmacological potential of Zanthoxylum zanthoxyloides. Overall, while the existing evidence indicates promising biological activity, it is characterized by methodological heterogeneity, limited standardization, and insufficient reporting of key quality parameters, highlighting the need for more rigorous and reproducible study designs.

Limitations

This review is a narrative integrative synthesis and does not follow a systematic review protocol; therefore, it is inherently subject to selection bias and limited reproducibility. The included studies exhibit substantial heterogeneity in experimental design, plant parts investigated, extraction procedures, and outcome measures, which limit direct comparability across pharmacological domains.

Across the reviewed literature, the evidence base is predominantly preclinical, consisting mainly of in vitro assays and small-scale animal studies. Methodological quality indicators such as randomization, blinding, allocation concealment, and power calculations are inconsistently reported, limiting robust assessment of internal validity. In addition, reporting of key quantitative parameters—including MIC, IC₅₀ values, and dose–response relationships—is uneven, and control selection (positive and negative controls) is not standardized across studies.

Reproducibility is further constrained by variability in extraction methods and incomplete phytochemical characterization in several reports. The limited reporting of negative or null findings also suggests potential publication bias. Finally, the absence of well-designed clinical trials remains a major gap, restricting the translation of preclinical findings into clinically validated therapeutic applications.

Future directions and recommendations

1. Standardization and quality control

   • Development of standardized extracts with defined phytochemical markers.

   • Adoption of pharmacognostic benchmarks for raw material authentication.

2. Mechanistic and molecular studies

   • Elucidation of molecular pathways underlying antiplasmodial, antisickling, and anti-inflammatory effects.

   • Structure–activity relationship studies of key alkaloids and phenolics.

3. Toxicological and safety evaluation

   • Comprehensive sub-chronic and chronic toxicity studies.

   • Assessment of herb–drug interactions, particularly for long-term use.

4. Clinical translation

   • Well-designed human clinical trials focusing initially on oral health and malaria adjunct therapy.

   • Development of validated phytopharmaceutical formulations.

5. Conservation and sustainability

   • Promotion of stem bark use over root bark.

   • Integration of cultivation and conservation strategies into phytomedicine development programs.