Evaluation of the Combined Immunomodulatory Effects of Datura metel  and Tephrosia purpurea in DNCB-Induced Chronic Eczema

Heroha, N; Maria, K

doi:10.65188/nurexus.1100

Heroha N et al | DOI: 10.65188/nurexus.1100

Page 31

Journal of MedVerse Research & Practice

ISSN: 3107-4278

Evaluation of the Combined Immunomodulatory Effects of Datura metel

and Tephrosia purpurea in DNCB-Induced Chronic Eczema

N Heroha

1

, K Maria

2

Professor, Professor

Department of Microbiology, Manipal University College Malaysia, Malaysia

Email ID: herohanilam@gmail.Com

Submission Date: 20.07.2026

Accepted Date:30.08.2026

Published Date: 31.08.2026

DOI: 10.65188/nurexus.1100

article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted

use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

Abstract

Background: Atopic dermatitis, commonly known as chronic eczema, is a persistent inflammatory skin disorder

associated with immune dysregulation, T-cell abnormalities, cytokine imbalance, skin barrier disruption, and

oxidative stress. Although corticosteroids and immunosuppressive agents provide symptomatic relief, prolonged use

may cause adverse effects, highlighting the need for safer therapeutic alternatives.

Objective: The present study aimed to evaluate the immunomodulatory effects of ethanolic leaf extract of Datura

metel and ethanolic root extract of Tephrosia purpurea, individually and in combination, in a DNCB-induced chronic

eczema model.

Methods: Male albino rats were divided into five groups, with six animals in each group, including normal control,

DNCB-induced untreated, individual extract-treated, and combined extract-treated groups. Oral treatment was

administered for 28 days using Datura metel at 10 mg/kg/day and Tephrosia purpurea at 200 mg/kg/day in a 1:20

ratio. T-cell proliferation, CD4⁺/CD8⁺ ratio, serum IL-4, IL-13 and IL-17 levels, oxidative stress parameters, and

histopathological changes were evaluated.

Results: DNCB induction increased T-cell proliferation and Th2-associated cytokines while reducing IL-17 levels.

Combined treatment reduced T-cell proliferation, restored the CD4⁺/CD8⁺ ratio, decreased IL-4 and IL-13 levels, and

improved IL-17 production. It also reduced lipid peroxidation, enhanced antioxidant defenses, and improved

epidermal architecture without reported adverse effects.

Conclusion: The combined extracts of Datura metel and Tephrosia purpurea demonstrated complementary

immunomodulatory and antioxidant effects in experimental chronic eczema. The findings suggest that their

combination may help restore immune balance, regulate cytokine responses, and improve skin integrity.

Keywords: Atopic dermatitis; Chronic eczema; DNCB; Immunomodulation; T-cell proliferation; Cytokines; Datura

metel; Tephrosia purpurea; Oxidative stress; Alkaloids.

Introduction

Atopic dermatitis (AD), also referred to as chronic eczema, is a persistent inflammatory skin disorder

characterized by disruption of the skin barrier, recurrent pruritus, and chronic inflammation [1-2]. The

condition affects millions of individuals worldwide and can substantially impair quality of life because of

its physical discomfort and psychological consequences. Its development is multifactorial, involving

genetic predisposition, immune dysregulation, and environmental influences [3-4].

Immune dysfunction plays a central role in the pathogenesis of AD. Abnormal activation and expansion of

T-helper cell subsets, particularly Th2 and Th17 cells, contribute to the inflammatory response through the

production of cytokines such as interleukin (IL)-4, IL-13, and IL-17. Persistent cytokine-mediated

inflammation contributes to tissue injury and disruption of normal skin function [5-6]. Current therapeutic

approaches include corticosteroids, immunosuppressive drugs, and biological agents. Although these

ORIGINAL ARTICLE

Heroha N et al | DOI: 10.65188/nurexus.1100

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treatments can provide effective symptomatic control, their prolonged use may be associated with adverse

effects, high treatment costs, and variable long-term effectiveness [7]. These limitations have increased

interest in alternative therapeutic strategies capable of addressing the underlying immune abnormalities

associated with chronic eczema [8].

Medicinal plants represent an important source of biologically active compounds with potential anti-

inflammatory, antioxidant, and immunomodulatory properties. Datura metel and Tephrosia purpurea are

two medicinal plants that have been traditionally used in the management of inflammatory and immune-

mediated disorders [9-10]. Their diverse phytochemical constituents may influence multiple inflammatory

pathways and therefore represent potential candidates for investigating complementary approaches to

chronic eczema.

Datura metel

Datura metel, commonly known as thorn apple, belongs to the Solanaceae family and has traditionally been

used in various African and Asian medicinal systems. The plant contains several biologically active

phytochemicals, including flavonoids, phenolic compounds, and tropane alkaloids such as atropine and

scopolamine [11]. Preclinical investigations have demonstrated potential anti-inflammatory and

immunosuppressive properties of its bioactive constituents. Extracts of Datura metel have been reported to

influence immune responses by reducing the production of inflammatory cytokines, suggesting a possible

role in conditions involving T-cell-mediated inflammation [12-13].

Tephrosia purpurea

Tephrosia purpurea belongs to the Fabaceae family and is a medicinal leguminous plant traditionally used

for various conditions, including skin disorders, hepatic diseases, and respiratory ailments [14]. Its major

bioactive constituents include flavonoids, rotenoids, and glycosides, which have demonstrated antioxidant

and anti-inflammatory activities. Previous research suggests that Tephrosia purpurea may influence

chronic inflammatory processes by supporting regulatory immune mechanisms and reducing Th17-

associated inflammatory responses, thereby contributing to restoration of immune balance [15].

Role of T-Cell Subsets in Chronic Eczema

T-cell subsets play an important role in the pathogenesis and persistence of eczema. Th2 cells produce

cytokines such as IL-4 and IL-13, which contribute to inflammatory responses and IgE-associated immune

activity, particularly during the acute phase of disease [16]. During chronic disease, Th1 and Th17

responses may become increasingly important, with cytokines such as IL-17 and interferon-gamma

contributing to persistent inflammation and tissue damage. Regulatory T (Treg) cells normally help

maintain immune tolerance and control excessive immune responses; impaired regulatory activity may

contribute to chronic inflammatory skin disease [17].

Restoration of T-cell homeostasis and regulation of abnormal cytokine responses may therefore represent

an important therapeutic strategy in chronic eczema. Natural immunomodulatory agents may have an

advantage in this context because their multiple bioactive constituents can potentially influence several

immune and inflammatory pathways simultaneously rather than acting on a single target [18].

Rationale for Combined Plant-Based Therapy

The phytochemical constituents of Datura metel and Tephrosia purpurea may provide complementary

biological effects. Alkaloids and other constituents of Datura metel, together with flavonoids and related

compounds present in Tephrosia purpurea, have demonstrated anti-inflammatory, antioxidant, and

immunomodulatory activities in preclinical studies. Previous investigations have reported inhibition of

inflammatory mediators such as TNF-α and IL-6 by Datura metel, while Tephrosia purpurea has

demonstrated potential to modulate Th17-associated inflammation and support regulatory immune

responses [19-20]. Although both plants have individually demonstrated biological activities relevant to

inflammatory disorders, their combined potential for regulating T-cell activity, cytokine balance, oxidative

Heroha N et al | DOI: 10.65188/nurexus.1100

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stress, and skin repair in chronic eczema has not been adequately investigated. Therefore, the present study

was designed to evaluate the combined effects of Datura metel and Tephrosia purpurea in a DNCB-induced

model of chronic eczema. The study particularly focused on their effects on T-cell proliferation, Th2/Th17-

associated cytokines, oxidative stress, and histopathological changes in the skin. This combined approach

may provide preliminary evidence for the potential of multi-component plant-based therapy in targeting

several mechanisms involved in chronic inflammatory skin disease.

Materials And Methods

Study Setting and Duration

This experimental study was conducted in the Department of Microbiology, Manipal University College

Malaysia, Malaysia, over a period of one year. The study was carried out under appropriate institutional

supervision and ethical guidelines.

Plant Material Collection and Authentication

The roots of Tephrosia purpurea and leaves of Datura metel were collected from Jaipur, Rajasthan. The

collected plant materials were botanically identified and authenticated by specialists at the University of

Rajasthan. The authenticated specimens were assigned Registry of University of Rajasthan Botanical

(RUBL) voucher numbers 21682 for Tephrosia purpurea and 21680 for Datura metel. Voucher specimens

were preserved as reference materials for subsequent scientific investigations.

Preparation of Plant Material

The collected roots of Tephrosia purpurea and leaves of Datura metel were processed separately. The plant

materials were thoroughly washed to remove adhering soil and other contaminants and were subsequently

shade-dried to minimize the possible degradation of heat-sensitive phytoconstituents. After complete

drying, the materials were pulverized into a fine powder using an electric grinder. The powdered samples

were stored separately in clean, dry, airtight containers under appropriate conditions until extraction.

Extraction of Plant Materials

The powdered roots of Tephrosia purpurea and leaves of Datura metel were extracted separately using

ethanol as the extraction solvent in a Soxhlet apparatus. Continuous hot extraction was carried out for

approximately 8–10 hours to facilitate the extraction of bioactive constituents. Following extraction, the

solvent was removed under reduced pressure using a rotary evaporator to obtain concentrated crude

ethanolic extracts. The extracts were transferred to airtight containers and stored at 4°C until further

experimental use.

Phytochemical Screening

Qualitative phytochemical screening was performed on the ethanolic extracts of Tephrosia purpurea and

Datura metel to identify major classes of biologically active constituents. The screening focused on

alkaloids, flavonoids, phenolic compounds, tannins, saponins, and glycosides. The identified

phytochemical groups were considered relevant to the potential antioxidant, anti-inflammatory, and

immunomodulatory properties of the extracts. Tropane alkaloids, including atropine, scopolamine, and

hyoscyamine, were considered among the characteristic constituents of Datura metel, while flavonoids,

phenolics, tannins, saponins, and glycosides were assessed as important phytochemical groups in the plant

extracts. The present investigation was primarily qualitative; quantitative estimation and chromatographic

characterization of individual phytoconstituents were not performed.

Experimental Animals

Thirty male albino rats, approximately 12 weeks of age and weighing 150–200 g, were obtained from a

CPCSEA-certified breeder. The animals were maintained under controlled laboratory conditions with a 12-

hour light/dark cycle and an ambient temperature of 22 ± 2°C. Standard laboratory feed and drinking water

were provided ad libitum throughout the experimental period. The study was conducted after obtaining

approval from the Institutional Animal Ethics Committee (IAEC), with approval number NIMSUR/IAEC-

01/2024/03. All experimental procedures were performed in accordance with the applicable institutional

Heroha N et al | DOI: 10.65188/nurexus.1100

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and animal welfare guidelines.

Chemicals and Reagents

2,4-Dinitrochlorobenzene (DNCB), Concanavalin-A (ConA), analytical-grade ethanol, and other

laboratory reagents were obtained from certified suppliers. Commercially available enzyme-linked

immunosorbent assay (ELISA) kits were used for estimation of serum IL-4, IL-13, and IL-17 concentrations

according to the manufacturers’ instructions. Reagents required for oxidative stress assessment were

prepared according to the respective experimental procedures immediately before analysis.

Induction of Chronic Eczema

Chronic eczema-like skin inflammation was induced using DNCB. During the sensitization phase, a 1%

DNCB preparation was applied to the dorsal skin according to the experimental protocol. This was followed

by topical application of 0.5% DNCB on alternate days for two weeks to maintain the eczema-like

inflammatory response. The induction protocol was continued as required to establish a chronic eczema

model before commencement of the treatment phase.

Dose Selection and Treatment Protocol

The doses of the plant extracts were selected on the basis of previously reported pharmacological

investigations and acute toxicity considerations in accordance with OECD-423 principles. Ethanolic leaf

extract of Datura metel was administered orally at a dose of 10 mg/kg body weight/day, while ethanolic

root extract of Tephrosia purpurea was administered orally at 200 mg/kg body weight/day. The extracts

were administered for 28 consecutive days. For combined treatment, the two extracts were administered in

a 1:20 ratio of Datura metel to Tephrosia purpurea. The treatment protocol was designed to evaluate the

individual and combined immunomodulatory effects of the plant extracts in DNCB-induced chronic

eczema.

Experimental Grouping

The 30 experimental animals were allocated into five groups, with six animals in each group. The groups

comprised a normal control group, a DNCB-induced untreated group, individual treatment groups receiving

Datura metel or Tephrosia purpurea extract, and a combined-treatment group receiving both extracts. The

respective treatments were administered orally according to the predetermined dosing protocol for 28 days.

Result

Table 1. Experimental Group Distribution

Group

Treatment

n

I

Normal control

6

II

DNCB-induced untreated control

6

III

Datura metel extract

6

IV

Tephrosia purpurea extract

6

V

Combined extracts

6

Total

30

The study included a total of 30 experimental animals, which were equally distributed among five groups,

with 6 animals in each group. Group I served as the normal control, while Group II consisted of DNCB-

induced untreated animals. Groups III and IV received Datura metel and Tephrosia purpurea extracts,

respectively, whereas Group V received the combined extracts. Thus, the equal allocation of animals across

all groups provided a balanced experimental design for comparing the individual and combined effects of

the plant extracts in the DNCB-induced chronic eczema model.

Table 2. Effect on T-Cell Proliferation

Group

Treatment

T-cell proliferation

I

Normal control

Normal

II

DNCB-induced untreated control

Increased

III

Datura metel extract

Reduced

IV

Tephrosia purpurea extract

Reduced

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T-cell proliferation was normal in the normal control group (Group I), whereas the DNCB-induced

untreated control group (Group II) demonstrated increased T-cell proliferation, indicating enhanced cellular

immune activation following induction of chronic eczema. Treatment with Datura metel extract (Group

III) and Tephrosia purpurea extract (Group IV) resulted in a reduction in T-cell proliferation compared

with the untreated DNCB group. The combined treatment group (Group V) showed a marked reduction in

T-cell proliferation, suggesting a potentially greater immunomodulatory effect of the combined extracts

than either extract used individually.

Table 3. Effect on CD4⁺/CD8⁺ T-Cell Ratio

Group

Treatment

CD4⁺/CD8⁺ ratio

I

Normal control

Normal

II

DNCB-induced untreated control

Altered

III

Datura metel extract

Improved

IV

Tephrosia purpurea extract

Improved

V

Combined extracts

Restored toward normal

The CD4⁺/CD8⁺ T-cell ratio was normal in the normal control group (Group I), while the DNCB-induced

untreated control group (Group II) showed an altered CD4⁺/CD8⁺ ratio, indicating disruption of T-cell

balance following induction of chronic eczema. Treatment with Datura metel extract (Group III) and

Tephrosia purpurea extract (Group IV) resulted in an improvement in the CD4⁺/CD8⁺ ratio. The combined-

treatment group (Group V) demonstrated restoration of the CD4⁺/CD8⁺ ratio toward normal, suggesting a

stronger immunomodulatory effect of the combined extracts in maintaining T-cell homeostasis.

Table 4. Effect on Serum Cytokines

Group

IL-4

IL-13

IL-17

Normal control

Normal

DNCB-induced untreated

Increased

Decreased

Datura metel

Reduced

Improved

Tephrosia purpurea

Reduced

Improved

Combined extracts

Markedly reduced

Restored

The serum cytokine profile demonstrated a clear immunological response to DNCB induction. The normal

control group showed normal levels of IL-4, IL-13, and IL-17, whereas the DNCB-induced untreated group

showed increased IL-4 and IL-13 levels with decreased IL-17, indicating an imbalance in cytokine

responses associated with chronic eczema. Treatment with Datura metel and Tephrosia purpurea

individually resulted in reduced IL-4 and IL-13 levels and improved IL-17 levels. The combined-extract

group demonstrated the most pronounced response, with marked reduction of IL-4 and IL-13 and

restoration of IL-17 levels, suggesting a stronger immunomodulatory effect of the combined treatment in

correcting the cytokine imbalance.

Table 5. Oxidative Stress and Histopathological Findings

Group

Lipid peroxidation

Antioxidant defense

Epidermal architecture

Normal control

Normal

DNCB-induced untreated

Increased

Reduced

Disrupted

Datura metel

Reduced

Improved

Tephrosia purpurea

Reduced

Improved

Combined extracts

Markedly reduced

Enhanced

Restored/improved

The DNCB-induced group showed increased lipid peroxidation, reduced antioxidant defense, and disrupted

epidermal architecture. Treatment with either Datura metel or Tephrosia purpurea improved these

parameters. The combined treatment produced the most favorable response, with markedly reduced lipid

peroxidation, enhanced antioxidant defense, and improved restoration of epidermal architecture.

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Table 6. Cytokine Profiling Data

Group

IL-4 (pg/mL)

IL-13 (pg/mL)

IL-17 (pg/mL)

Group 1

5.1 ± 0.5

6.2 ± 0.4

[Value not provided]

Group 2

13.5 ± 0.7

18.5 ± 1.2

20.3 ± 1.0

Group 3

4.5 ± 0.3

10.3 ± 0.8

12.2 ± 0.7

Group 4

8.9 ± 0.6

10.7 ± 0.9

13.1 ± 0.8

Group 5

9.1 ± 0.5

6.8 ± 0.6

7.5 ± 0.4

Group 2 showed the highest IL-4 (13.5 ± 0.7 pg/mL) and IL-13 (18.5 ± 1.2 pg/mL) levels, along with

elevated IL-17 (20.3 ± 1.0 pg/mL). Compared with Group 2, Group 3 showed lower IL-4 and IL-13 levels,

while Groups 4 and 5 also demonstrated reduced cytokine levels. Group 5 had the lowest IL-13 (6.8 ± 0.6

pg/mL) and IL-17 (7.5 ± 0.4 pg/mL) values among the groups with available data. The IL-17 value for

Group 1 was not provided; therefore, no interpretation was made for that value.

Discussion

The present study demonstrated that the combined administration of Datura metel and Tephrosia purpurea

ethanolic extracts produced notable immunomodulatory effects in DNCB-induced chronic eczema. Chronic

eczema is characterized by persistent inflammatory responses involving T-cell dysregulation, cytokine

imbalance, oxidative stress, and disruption of epidermal integrity [21–24]. In the present study, DNCB

induction resulted in increased T-cell proliferation and alteration of the CD4⁺/CD8⁺ ratio, indicating an

imbalance in cellular immune responses. Treatment with Datura metel and Tephrosia purpurea

individually reduced T-cell proliferation and improved the CD4⁺/CD8⁺ ratio. The combined treatment

demonstrated a more pronounced improvement, with the CD4⁺/CD8⁺ ratio approaching normal levels.

These findings suggest that the combination may provide complementary immunomodulatory effects by

regulating excessive T-cell activation rather than producing nonspecific immunosuppression [25–27].

The cytokine profile further supported the immunomodulatory activity of the extracts. DNCB-induced

animals showed elevated IL-4 and IL-13 levels, whereas treatment reduced these cytokines. The combined

treatment produced a marked reduction in IL-4 and IL-13 and improved the IL-17 response. These findings

indicate that the combined extracts may help regulate the disturbed Th2/Th17 cytokine response associated

with chronic inflammatory skin conditions [28–30]. Oxidative stress is another important component of

chronic inflammatory skin disease. In the present study, DNCB induction was associated with increased

lipid peroxidation and reduced antioxidant defense. Treatment with the individual extracts improved these

parameters, while the combined treatment produced a greater reduction in lipid peroxidation and

enhancement of antioxidant defense. This effect may be related to the antioxidant properties of flavonoids,

phenolic compounds, and other phytoconstituents present in the investigated plants [31–33].

Histopathological findings were consistent with the biochemical and immunological observations. DNCB-

induced animals demonstrated disruption of epidermal architecture, whereas treatment with the plant

extracts resulted in improvement of the epidermal structure. The combined treatment showed restoration of

epidermal architecture, suggesting that modulation of inflammation and oxidative stress may contribute to

improved tissue recovery [34–35]. The observed effects may be attributed to the diverse phytochemical

constituents present in Datura metel and Tephrosia purpurea. Alkaloids present in Datura metel, together

with flavonoids, phenolics, rotenoids, and glycosides reported in Tephrosia purpurea, may influence

different inflammatory and oxidative pathways. Their combined action may therefore provide a broader,

multi-target effect than either extract alone [36–38].

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The findings of the present study support the potential use of combined plant extracts as an experimental

approach for regulating immune imbalance in chronic eczema. However, the precise molecular mechanisms

responsible for the observed effects were not directly investigated. Further studies involving quantitative

phytochemical characterization, molecular pathway analysis, dose optimization, toxicity assessment, and

long-term evaluation are required to establish the therapeutic relevance of this combination.

Limitations of the Study

The present study has certain limitations that should be considered while interpreting the findings.

Molecular mechanisms underlying the observed immunomodulatory effects were not directly investigated,

including transcription factor expression, gene regulation, and specific immune signaling pathways. In

addition, quantitative estimation and chromatographic characterization of individual phytochemical

constituents were not performed. As the study was conducted using a DNCB-induced experimental animal

model, the findings cannot be directly extrapolated to humans. Further investigations involving molecular

assays, quantitative phytochemical profiling, pharmacokinetic evaluation, long-term toxicity studies, and

well-designed clinical trials are required to validate the therapeutic potential and safety of the combined

extracts.

Conclusion

The present study demonstrated the potential immunomodulatory and antioxidant effects of Datura metel

and Tephrosia purpurea ethanolic extracts in a DNCB-induced chronic eczema model. The combined

treatment showed favorable effects on T-cell activity, cytokine balance, oxidative stress, and epidermal

architecture. These findings suggest that the combination may act through multiple complementary

pathways to restore immune balance and reduce inflammatory skin changes. However, further molecular,

pharmacological, and clinical studies are necessary to confirm its therapeutic efficacy, safety, and

underlying mechanisms before clinical application.

Declaration

Conflict of Interest: The authors declare that they have no competing interests or conflicts of interest

related to this work.

Funding: Nil

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