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Metabolic Diversity and Therapeutic Potential of Holarrhena pubescens: An Important Ethnomedicinal Plant

Kulsoom Zahara
Sujogya Kumar Panda
Shasank Sekhar Swain
2 and
Walter Luyten
Department of Biology, Katholieke Universiteit Leuven, 3000 Leuven, Belgium
Division of Microbiology and NCDs, ICMR-Regional Medical Research Centre, Bhubaneswar 751023, Odisha, India
Author to whom correspondence should be addressed.
Authors contributed equally.
Biomolecules 2020, 10(9), 1341;
Submission received: 16 August 2020 / Revised: 13 September 2020 / Accepted: 14 September 2020 / Published: 18 September 2020
(This article belongs to the Collection Pharmacology of Medicinal Plants)


Holarrhena pubescens is an important medicinal plant of the Apocynaceae family that is widely distributed over the Indian subcontinent. The plant is extensively used in Ayurveda and other traditional medicinal systems without obvious adverse effects. Beside notable progress in the biological and phytochemical evaluation of this plant over the past few years, comprehensive reviews of H. pubescens are limited in scope. It has economic importance due to the extensive use of seeds as an antidiabetic. Furthermore, the plant is extensively reported in traditional uses among the natives of Asia and Africa, while scientifical validation for various ailments has not been studied either in vitro or in vivo. This review aims to summarize information on the pharmacology, traditional uses, active constituents, safety and toxicity of H. pubescens. Chemical analysis of H. pubescens extracts revealed the presence of several bioactive compounds, such as conessine, isoconnessine, conessimine, conimine, conessidine, conkurchicine, holarrhimine, conarrhimine, mokluangin A-D and antidysentericine. Overall, this review covers the ethnopharmacology, phytochemical composition, and pharmacological potential of H. pubescens, with a critical discussion of its toxicity, biological activities (in vitro and in vivo), the mechanism of action, as well as suggestions for further basic and clinical research.

Graphical Abstract

1. Introduction

Plants are long been used as a source of medicine by human beings. However, the compound(s) responsible for their therapeutic activities remained unknown for centuries. At the end of 19th century a shift occurred from natural to synthetic drugs, and phytomedicine has since progressively fallen out of use [1]. Though synthetic medicines are in use all over the world, some ancient medicinal systems still persist, e.g., Unani Tibb, Ayurveda, and traditional Chinese medicine. They make ample use of medicinal plants, which provide a source to identify chemical compounds and use them to treat different ailments [2].
Holarrhena pubescens Wall. ex G.Don, Syn. Holarrhena antidysenterica (Roth) Wall. ex A.DC. is a medicinally important plant of Africa as well as tropical and subtropical regions of Asia [3]. It is widely used in Indian medicine for treating diseases viz. diarrhea, amoebic dysentery, liver disorders, irritable bowel syndrome, and bleeding piles. The plant is astringent and bitter in taste. It is used traditionally to treat several diseases (Table 1) and there are clinical and pharmacological studies suggesting its use for various enteric, skin diseases and diabetes [4].

1.1. Geographical Distribution

The geographical distribution of H. pubescens is shown in Figure 1. It is native to South-central China, Cambodia, Myanmar, Thailand, Vietnam, India, Nepal, Bhutan, Pakistan, Bangladesh, Laos, Malawi, Mozambique, Kenya, Northern Tanzania, Zaïre, Zambia and Zimbabwe. It was introduced in South-east China, Hainan, Taiwan, and Mauritius, but its presence in Malaysia is doubtful.

1.2. Morphological Description

H. pubescens is a deciduous tree, with oblong and elliptic leaves. Flowers are white, fragrant corymbose cymes. The corolla is lobed and oblong. Fruits are slender, terete follicles, with white spots. Seeds are glabrous and linear-oblong. Its flowering season is from April–July, and fruiting is from August–October [5].

2. Phytoconstituents

A wide range of phytochemicals has been documented in H. pubescens.
  • Steroidal alkaloids: conarrhimine, conessine, holantosines a, b, c, d, e and f, holarrhessimine, holarrhidine, holarrhine, holonamine, hydroxyconessine, kurchiline, kurchine, kurchiphylline, norconessine, n,n,n′,n′tetramethylholarrhenine, holacin, kurchinine, conamine, holadysine, 12-hydroxyconessimine, holarrhimine, holadysamine, conessimine, isoconessimine, holarosine a, conessidine, kurchiphyllamine, 7α-hydroxyconessine [26,27].
  • Uncharacterized alkaloid: lettocine [28].
  • Triterpenes: betulinaldehyde, ursolic acid, lupeol, 20(29)-lupadien-3β-ol, betulinic acid, lupeol β-hydroxyhexad-ecanoate [29,30].
  • Sterols: sitosta-5,23-dien-3β-ol, stigmasterol [31].

3. Traditional Uses

H. pubescens is widely used in Ayurveda and traditional Chinese medicine. Its seeds are used as anthelminthic, and its bark is reported to have antidiarrheal properties [32]. In Ayurvedic medicine it is used for treating anemia, jaundice, dysentery, stomach pains, diarrhea, epilepsy and cholera [33]. It is widely known for the treatment of Asra (blood or blood-related disorders), Atisara (diarrhea), Kustha (leprosy), Pravahika (amebiasis), Jwaratisara (secondary diarrhea) and Tŕṣṇā (thirst) [34].
As described in Table 2, different parts of this plant are used by tribal communities throughout various regions of the world.

3.1. Bark

  • In Ayurvedic medicine, its bark is used extensively for the treatment of piles, diarrhea, leprosy, biliousness and diseases of the spleen [35,36].
  • In Unani medicine, bark is used to treat excessive menstrual flow, piles and headache [37].
  • In British Materia Medica, its bark is used as antiprotozoal agent, for malaria, against chest infections, for asthma, bronchopneumonia, gastric disorders, dyspepsia, diarrhea and dysentery [38].

3.2. Leaf

  • In Ayurveda, H. pubescens leaves are not reported to have medicinal value.
  • In Unani medicine, leaves are used as aphrodisiac, tonic, astringent and galactagogue, and are thus used for treating chronic bronchitis, urinary discharges, wounds, ulcers, as well as for muscles relaxation; they are also useful to regulate menstruation [72].

3.3. Roots

Roots are reported to be aphrodisiac and abortifacient [73]. They are also used against venereal diseases, gonorrhea, ascariasis, malaria and severe abscesses [74].

3.4. Flowers

In Ayurveda, flowers are used as anthelmintic, antidiarrheal, and reportedly to treat leukoderma and diseases related to blood and spleen [75].

3.5. Seeds

  • In Ayurveda medicine, the seeds are used as anthelmintic, astringent, and to cure dysentery, biliousness, leprosy, fatigue, skin diseases, bleeding piles, and hallucinations [76,77].
  • In Unani medicine, seeds are used as carminative, aphrodisiac, astringent and lithotriptic [78].
  • In Tibetan medicine, they are used as alexipharmic, antidiarrheal, cholagogue, and analgesic [79].
  • In the indigenous Bangladesh system of medicine, they are used as astringent, anthelmintic, febrifuge, stomachic, anti-dysenteric and anti-diarrheal [80].
  • In other parts of the world, they are reportedly used against diuresis, chronic chest infection, asthma, malaria and vaginitis [81], diabetes [82], arthritis, hematuria, epilepsy, bronchitis, diarrhea, eczema and jaundice (Figure 2).
Different H. pubescens parts are also used by local communities in India to treat a wide array of diseases at different dosage (Table 3).

4. Pharmacology

4.1. Anti-diabetic Property

H. pubescens has been used for treating diabetes in various medicinal systems. Its methanol, aqueous and petroleum ether extract of seeds are reported to have antihyperglycemic activity at a dose of 250 mg/kg body weight (BW) in rat models [108]. Keshri [109] also reported its activity against streptozotocin-induced diabetes. Especially the methanol extract of its seeds successfully protects diabetic rats at a dosage of 300 mg/kg BW (Table 4).
The ethanolic extract of its seeds significantly reduced diabetes in rats at a dose of 300 mg/kg. In a similar study on diabetic rats, decreased levels of serum cholesterol, uric acid, aspartate transaminase, triglycerides, creatinine and blood glucose were observed. Another study also demonstrated that a hydro-methanolic seed extract of H. pubescens causes inhibition of α glucosidase (a gut exoenzyme that releases glucose from di- and oligo-saccharides and aryl glucosides in the diet, thereby increasing absorption of glucose from the intestine) [110]. An ethanol extract of its seeds also prevented weight loss in diabetic rats and corrected biochemical parameters. With the administration of 300 mg/kg and 600 mg/kg, a significant reduction of serum cholesterol, blood glucose concentration, mean alanine aminotransferase, triglycerides, uric acid, aspartate transaminase, alanine and creatinine were reported [109].
Besides seeds, its leaves are also effective against diabetes; Hedge and Jaisal [111] reported this for an ethanolic extract of H. pubescens leaves at a dose of 400 mg/kg BW of rats when administrated for 21 consecutive days. The methanol extract of this plant also possesses significant (p < 0.05) hypoglycemic activity in vivo [112]. A study by Bhusal [113] indicated a notable antidiabetic activity, specifically with an alcoholic extract, nearly equal to standard glibenclamide.

Mechanism of Action

Ali et al., [10] demonstrated the effect of H. pubescens on α glucoside activity. α glucosidase is an enzyme which converts polysaccharides into monosaccharides. Intestines are only able to transport sugar to the blood in monosaccharide form. It was observed that an H. pubescens extract significantly inhibits intestinal α glucosidase with IC50 = 0.52 mg/mL, thus successfully limiting carbohydrate absorption. This study suggested that inhibition of α glucosidase is an important approach to limit postprandial hyperglycemia in diabetes.

4.2. Anti-Diarrheal Property

Diarrhea is a condition of increased secretion, volume, fluidity, and frequency of bowel movements, thus causing loss of electrolytes and water. An ethanol extract of H. pubescens seeds when tested on castor oil-induced diarrhea in rats is reported to cause a significant increase in the feces dry weight, and reduced defecation. At a dosage of 200 and 400 mg/kg BW, a significant reduction (p < 0.05) of castor oil-induced diarrhea is observed [114] (Table 4).
In another study on alkaloids isolated from H. pubescens, seeds were tested against clinical isolates of enteropathogenic Escherichia coli in vitro, and castor oil-induced diarrhea in vivo. This successfully reduced diarrhea at a dose of 200–800 mg/kg [118].
Phytochemicals i.e., saponins, steroids, alkaloids, tannins and flavonoids are reported to be responsible for the antidiarrheal activity of plants. H. pubescens seeds extract tests positive for alkaloids and flavonoids; therefore, these may be responsible for this activity. Aqueous and methanol extract of H. pubescens leaves were found effective against the diarrheal pathogens Salmonella typhimurium, Salmonella typhi, Vibrio cholerae and Vibrio alginolyticus [12].
Daswani et al. [115] tested the effect of H. pubescens root bark aqueous extract on Escherichia coli. It was observed that this plant significantly inhibits stable toxin production and reduces intestinal secretions, thus causing a decreased virulence of these enterotoxigenic E. coli strains. Srivastava and Saxena evaluated in vitro activity of H. pubescens seeds aqueous extract against diarrhea caused by bacteria like Staphylococcus aureus, E. coli, Shigella, and Salmonella typhi, and found this extract highly effective on the tested bacterial strains [119].

4.3. Anti-inflammatory and Analgesic Properties

H. pubescens extract can inhibit rat carrageenan-induced paw edema at doses of 100 and 200 mg/kg BW [116]. The methanol extract of its bark showed a decreased level of malondialdehyde and nitric oxide, but an increase in glutathione and superoxide dismutase in colitis induced in male albino Wistar rats [120]
Studies also suggest the anti-inflammatory efficacy of H. pubescens in a dose-dependent manner; a 400 mg/kg dose showed 74% (p < 0.01) inhibition when tested on carrageenan-induced rat paw edema [116]. In another study by Haque et al. [121] the methanol, petroleum ether, chloroform, dichloromethane and aqueous extract of its stem was evaluated. The chloroform extract produced maximum analgesic effect with 71% abdominal writhing inhibition, 88.5% inhibition in the open field test and CNS-depressant activity with 83% inhibition in locomotion at a 200 mg/kg dose.
In an acute inflammation model, the methanolic extract of H. pubescens in doses of 100 and 200 mg/kg produced dose-dependent inhibition of paw edema. The test and the standard drugs produced significant inhibition of paw edema as compared to the control (p < 0.001) at 3 and 4 h duration [122].
The methanolic leaf extract of H. pubescens (100 and 200 mg/kg) suppressed the acetic acid-induced writhing response significantly in a dose-dependent manner. In the same study, the analgesic effect was also tested in the tail flick model. Fifteen min. after drug administration, there is a significant increase in reaction time compared to the pre-drug reaction time. The extract enhances the stress tolerance capacity in animal models. The analgesic effect is proposed to be mediated by the prostaglandin pathways [123] and peritoneal mast cells [124].

4.4. Antioxidant/Free Radical Scavenging Properties

The aqueous and methanol extract of H. pubescens show a very strong radical scavenging activity with 90% DPPH free radical inhibition. The methanol extract also significantly reduces hydroxyl - and superoxide ions. In addition, it also causes a reduction of Fe3+ → Fe2+ conversion. Another study reported that its application decreases the damage to deoxyribose by OH ions. Similarly, H2O2 degradation, nitrite inhibition and lipid peroxidation were inhibited by the ethyl acetate fraction [125].
Zahin et al. [126] investigated the antioxidant capacity of H. pubescens by the ferric thiocyanate (FTC) method, thiobarbituric acid (TBA) method and DPPH radical scavenging method. Their results revealed a fair antioxidant effect using the FTC and TBA method, but a very low DPPH radical scavenging activity (20%). In another study conducted by Bhusal, [113] H. pubescens bark was evaluated for antioxidant effects, whereby both methanol and ethanol extracts had strong DPPH inhibition activity (methanol extract 96% at 0.1 mg/mL), whereas the hexane extract appeared to have the weakest activity.

4.5. Anti-Urolithic Property

The methanol extract of H. pubescens seeds was reported to have an inhibitory effect on the formation of calcium oxalate crystals. When tested in male Wistar rats, it shows a significant decrease in polyurea, Ca++ excretion, Ca++ crystal formation and water intake. These finding suggest that the plant has the potential to reduce kidney stones [117].

4.6. Diuretic Property

At a dose of 30–100 mg/kg in Wistar rats, the aqueous seed extract of H. pubescens was reported to increase urine output notably. A significant increase of excretion of Na+ and K+ ions was also observed. The chloroform extract of H. pubescens was also reported to cause a dose-dependent increase in urine output. In addition to this, an elevated level of urinary Na+ and K+ was also observed, thus showing that increased electrolyte excretion is probably responsible for its diuretic effect [127].

Gut Activities

The gut motility activity of H. pubescens was investigated by Gilani et al., [128]. They investigated the mechanism behind this activity of H. pubescens by testing extracts on high K+-induced contractions. The high K+ (> 30 mM) causes contraction of smooth muscles by opening L-type Ca++ channels, thus allowing entry of Ca++ in the cell and producing a contractile effect. Thus, an inhibitor of K+-induced contraction is an inhibitor of the Ca++ influx. In their study, hydro-ethanolic crude extract of H. pubescens relaxed the high K+-induced contractions just like a standard Ca++ antagonist. Thus, it was concluded that it effectively causes Ca++ channel blocking. Therefore, these extracts may be effective for treating gut disorders such as abdominal cramps and diarrhea.

4.7. Inhibition of Acetylcholinesterase and CNS-Stimulant Activity

In a study on alkaloids isolated from H. pubescens, five alkaloids were tested for CNS-stimulant activity i.e., conimine, isoconessimine, conessine, conarrhimine and conessimine. Conessimine showed the highest activity with an IC50 value of 4 μM. This study suggests that these alkaloids can be used for the treatment of neurological disorders [129]. Another study on Swiss albino mice showed that a methanolic bark extract notably decreased the grip strength and lowered locomotive activity, thus showing a depressant effect on the CNS [130].

4.8. Anti-Microbial Activity

Ethanol extracts of H. pubescens seeds showed a concentration-dependent antibacterial activity against enteropathogenic Escherichia coli (EPEC). The petroleum ether extract of its bark also showed inhibition of E. coli at a 50 μg/mL minimum inhibitory concentration. However, compared to other plants, it showed a moderate activity [131]. Studies [118] showed that adherence of the EPEC strain to INT407 cells leads to cytoplasmic membrane damage (electron microscopic studies), apoptotic bodies by the condensation of chromatin, and mitochondrial swelling and damage (fluorescence microscopy). These effects were diminished in EPEC treated with H. pubescens extracts.
Methanol extracts of H. pubescens exhibited antibiofilm activity against V. cholerae. Results of gene expression studies revealed that both leaf and bark extracts down-regulate aph A or aph B, the major regulator genes modulating both virulence and biofilm formation [132].
The alkaloidal fraction of H. pubescens showed a borderline antifungal activity, with a minimum inhibitory concentration (MIC) of 15.6 µg per disc. The methanol extract of H. pubescens bark showed significant antifungal potential against Candida albicans [133].
Conessine is the principal alkaloid demonstrated to have antibacterial activity. Till today it has not been proven whether the antimicrobial activity is due to a single alkaloid or due to a mixture of alkaloids present in this plant. Bioassay-guided purification is lacking so far, although this approach has been reported over the past 4 decades.

4.8.1. Synergy and Mechanism of Action

Acinetobacter baumannii and Pseudomonas aeruginosa are important nosocomial pathogen, and treatment options are limited. Their resistance mechanisms include the production of beta-lactamases, efflux pumps, and target-site or outer membrane modifications. When tested at 250 μg/mL, the H. pubescens ethanol extract showed low intrinsic antibacterial activity against Acinetobacter baumannii and significantly enhanced the activity of the antibiotic novobiocin (concentration = 1 μg/mL, 1/8th of its MIC). Moreover, the extract at 7.8 μg/mL, confirmed resistance-modifying ability (RMA) and may be a candidate as an alternative treatment for MDR infections due to A. baumannii [134]. Novobiocin was chosen because of its weak antibacterial activity against Gram-negative pathogens due to an effective permeability barrier. Interestingly, when tested at different concentration using a two-fold dilution starting from 250 µg/mL, the ethanol extract enhanced the inhibitory effects of novobiocin as well as synergetic effects against all tested clinical isolates [135]. However, the authors observed no enhancement of the accumulation of ethidium bromide after treatment with the extract, suggesting that it does not act by inhibiting MDR pumps. However, it weakened the outer membrane of the pathogen as exhibited by an increase in the N-phenyl-1-naphthylamine uptake [136].
Siriyong et al. [137] investigated the efficacy of an H. pubescens extract and conessine as resistance-modifying agents (RMAs) on the susceptibility of A. baumannii to novobiocin and rifampicin. The authors observed significant synergistic activity: the fractional inhibitory concentration (FIC) index was ≤ 0.5. To investigate the mechanism of synergism, the authors used fluorescent dyes and different efflux pump inhibitors and concluded that neither the extract nor conessine act as permeabilizers [138]. The authors also noticed an increase in pyronin Y (p < 0.05), while there was no accumulation of ethidium bromide, suggesting the synergism was due to interference with the AdeIJK pump, but no involvement of the AdeABC pump [139].
Antibacterial activity of H. pubescens against P. aeruginosa was confirmed by several scientist using different assays. However, its mechanism is still unclear, although the principal compound with bioactivity was the alkaloid conessine.

4.8.2. Conessine is Major Compound Responsible for Antimicrobial Activity

Siriyong et al. concluded that conessine in H. pubescens is responsible for antibacterial activity and can be useful as a combinatory therapy to restore antibiotic susceptibility in the extensively drug-resistant A. baumannii [137]. Siriyong et al. [138] studied the synergistic activity of conessine in combination with various antibiotics against P. aeruginosa PAO1 strain K767 (wild-type), K1455 (MexAB-OprM overexpressing), and K1523 (MexB deletion). An H33342 accumulation assay was used to evaluate efflux pump inhibition, while NPN uptake was used to assess membrane permeabilisation. Except for novobiocin, all other antibiotics tested such as cefotaxime, levofloxacin, tetracycline, erythromycin, and rifampicin showed synergistic activities. The authors observed that conessine might inhibit other efflux systems present in P. aeruginosa as indicated by synergy inhibition in the MexB deletion strain, while the inhibition of the MexAB-OprM pump was confirmed (H33342 efflux). However, membrane permeabilisation was not observed. This suggest that conessine may be applied as a novel efflux pump inhibitor to restore antibiotic activity by inhibiting efflux pump systems in P. aeruginosa and other Gram-negatives. Later, the authors tested a “P. aeruginosa strains with defined mutations that result in the overexpression of the MexAB-OprM, MexCD-OprJ and MexEF-OprN efflux pumps” as well as a mutated strain with deletion of all these pumps. The authors also studied the effects in an in vivo Galleria mellonella infection model. Conessine along with levofloxacin enhanced bacterial inhibition in vitro, and restored antibiotic efficacy in vivo compared to the corresponding monotherapies. The authors conclude that conessine from H. pubescens, enhanced the efficacy of several antibiotics, and inhibited efflux mediated MDR, without showing any toxicity in G. mellonella larvae [138]

4.9. Anti-malarial Activity

H. pubescens bark chloroform extracts showed significant in vitro and in vivo anti-malarial activity when tested on Plasmodium falciparum isolates, and when administered to Swiss mice infected with Plasmodium falciparum isolates, with average IC50 value of 5.7 μg/mL [139,140].
Nondo et al. [141] reported that ethanol and methanol extracts exhibit significant antiplasmodial activity against Plasmodium (P.) falciparum with an IC50 = 2.43 μg/mL and 2.05 μg/mL, respectively. Moreover, the fractions isolated from H. pubescens roots are highly active against chloroquine-resistant P. falciparum (K1, Dd2) and artemisinin-resistant P. falciparum. Another study [24], showed that the steroidal alkaloid conessine isolated from H. pubescens bark showed anti-plasmodial activity with an IC₅₀ value of 1.9 μg/mL.
Verma [140] tested petroleum ether and chloroform extracts of H. pubescens in P. berghei-infected mice and showed that its bark actively inhibits parasitemia. Simonsen et al. [142] also reported that a crude extract of H. pubescens bark has significant vitro anti-plasmodial activity with an IC50 value of 28 μg/mL against a chloroquine-susceptible strain of P. falciparum. The stem, root and seeds of this plant are reported to contain a large amount of steroidal alkaloid compounds i.e., conessine, kurchine, conessidine, isoconessine, conkurchicine, and holarrhimine. The chemical compound that is thought to be responsible for the antimalarial activity is conessine, isolated from its stem [142]. Verma et al. also reported that a methanol extract of H. pubescens inhibits P. berghei growth with a 43% suppression rate [140].

4.10. Stracture Activity Relationship (SAR) Study

Currently, isolated natural products capture a lot of attention for ‘lead compound’ selection in the primary stage of drug discovery, where the SAR analysis plays a vital role in describing the structural configuration connected with biological activity and potential mechanisms of action [143,144]. For H. pubescens, the most common steroidal-alkaloid class of phytoconstituents; conarrhimine, conessimine, conessine, conimine and isoconnessine was reported to have multiple and dose-dependent biological activities. Structurally, the above-mentioned five compounds are derived from the “conanine” moiety: a chemical structure containing an extra pyrrolidine or tetrahydropyrrole group to the steroid moiety. For example, all five analogues showed acetylcholinesterase (AChE)/neuroprotective activity within an IC50 range of 4 ± 0.1 to > 300 (μM) in vitro [129]. The SAR revealed that the attachment of a methyl group (-CH3) to the pyrrolidine (N-atom at the position C-19 of the alkaloid moiety), as well as adding a tetradecahydro-cyclopenta-phenanthrene ring (N-atom at position C-10 of the steroid moiety), influenced AChE inhibition (Figure 3) [129]. Conessimine was the most potent AChE inhibitor, with an IC50 of 4 μM, where a double -CH3 group is present on the steroid N-atom (at position C-10), but no -CH3 group on the pyrrolidine N-atom (at position C-19). Similarly, the absence of -CH3 group at the C-19 position in both conimine and conarrhimine and the presence of a single -CH3 group/lack of -CH3 group at C-10 vary the IC50 between 23 to 28 μM. On the other hand, the presence of a -CH3 group at C-19 in conessine and double -CH3 groups at C-10 yielded an IC50 of 21 μM, but the presence of a single -CH3 group (elimination of one -CH3 group) at position C-10 in isoconessimine, drastically lowered the AChE inhibition (IC50 > 300 μM).
Another similar group of natural products; mokluangin A-C and antidysentericine, also exhibited AChE inhibition with an IC50 range of 1.44 to 23.22 μM by the presence of a carboxylic group (-C=O) at C-18 and C-20 on the pyrrolidine ring, and a -CH3 group at C-10 on the steroid moiety [145]. Regarding the SAR, the presence of a double carboxylic group at C-18 and C-20 in mokluangin B reduces AChE inhibition (IC50 = 23.22 μM) compared to mokluangin C (IC50 = 1.44 μM), mokluangin A (IC50 = 2.12 μM) and antidysentericine (IC50 = 4.09 μM), which have a -C=O at C-18 and a -CH3 group at C-20 (Figure 4 and [145]). Thus, in both cases, the position of the -CH3 group on the isolated natural novel steroid-alkaloid moiety plays a significant role in AChE inhibition [129,145]. Additionally, Zhao’s research demonstrated that the novel steroid-alkaloid conessine potentially crosses the brain-blood barrier at a higher rate in human and mice brain than the imidazole-containing compounds thiopermide and ciproxican, based on a cell and tissue functional assay [146].

4.11. Molecular Docking Studies with Conessine

Molecular docking is another artificial intelligence-based computational method for finding potential biological activities of a natural product through binding energy/docking score calculations (kcal/mol) based on “target-ligand” docking complexes [147]. Typically, the target is a macromolecule associated with the disease of interest, and the ligand is a therapeutic agent used to inhibit or activate the macromolecule and its pathways/function. A similar docking approach was also used by Cheenpracha et al., taking mokluangin A-C and antidysentericine as ligand, and the crystallographic structure of AChE reported from Electrophorus electricus (PDB ID: 1C2B) [145]. Similarly, using information from previous studies, six different biological activities of conessine such as antibacterial, anti-CNS, antidiabetic, antifungal, anti-inflammatory and antimalarial were analyzed through a blind docking approach [147,148]. To find out more molecular details on the binding mode of this steroid-alkaloid moiety to the presumptive molecular targets (Figure 5). Based on the individual docking score (kcal/mol), conessine would be expected to exhibited more anti-CNS activity with docking score −11.18 kcal/mol (PDB ID: 1C2B), than anti-inflammatory activity with docking score −10.18 kcal/mol (PDB ID: 1F19), antibacterial activity with docking score −9.40 kcal/mol (PDB ID: 1HNJ), antifungal activity with docking score −9.38 kcal/mol (PDB ID: 6TZ6), antidiabetic activity with docking score −8.84 kcal/mol (PDB ID: 5NN4) and antimalaria activity with docking score −8.71 kcal/mol (PDB ID: 1LDG) (Figure 5). Thus, a docking study may a cost-effective computational analysis to help understand different biological activities in the form of binding energy and possible molecular interaction-cum-mode of inhibition. Nowadays, molecular docking is also a useful tool in drug development to identify potential hit and better lead compounds, as well as insights into their mode of action [145,148].

5. Safety and Toxicity Studies

Various crude extracts from H. antidysenterica seeds such as water, ethanol, hydro alcoholic etc. were studied for their acute oral toxicity by Sheikh et al. [108] and Pathak et al. [149]; they were found to be safe up to 2000 mg/kg BW in albino rats. Singh [150], conducted a pre-clinical safety study of H. antidysenterica stem bark in both mice and rats. Albino mice (Swiss) treated with 2000, 1000 & 500 mg/kg, p.o. showed dullness and writhing, and a 30% mortality was recorded within 96 h. However, the authors further studied subacute toxicity in rats with lower doses (50, 100, 200 mg/kg, p.o.) and found no significant changes in hematological or biochemical parameters and histopathological examinations. An acute toxicity study conducted by Keshri et al. [109] and Kumar and Yadav [151] in albino rats revealed that ethanolic extracts of H. antidysenterica seeds showed no toxicity at 3000 mg/kg. Saha and Subrahmanyam [116] and Hegde and Jaisal [111] found ethanolic extracts of H. antidysenterica seed and leaves, respectively, to be safe when administrated at 3000 mg/kg. Other studies showed the nontoxic nature of H. antidysenterica when administered in albino rats at different oral doses, including 200 and 400 mg/kg [110,111,116]. A similar study conducted by Bhusal et al. [113] in male Swiss albino mice also detected no toxicity at 250 and 500 mg/kg (p.o) for methanolic extract of H. pubescens stembark.
In a subchronic toxicity study, an ethanol extract of H. pubescens along with polyvinyl pyrrolidone administrated at dosages of 270 and 530 mg/kg BW/day (which is 10 and 20 times more than the dosage used for humans), caused hepatotoxicity in rats when given for 3 consecutive months [152].

6. Clinical Trials

The search term “Holarrhena” was used to search PubMed while specifying article type “Clinical Trials”. Although no published papers with clinical studies were retrieved in this way, when we searched with the same term in Google scholar, more than a dozen publications were retrieved, most of them from India. The first clinical trial with alkaloids of H. pubescens for the treatment of amoebic hepatitis was carried out by Chopra and De [153]. However, they concluded that four doses of intramuscular injection of the total alkaloid mixture yields no improvement, while the patient complained of severe pain in the right hypochondriac region. Singh [154] carried out a clinical study with 40 patients suffering from intestinal amoebiasis and/or giardiasis using H. pubescens. In 70% of patients a response was observed in the Entamoeba histolytica cysts, and the authors conclude that H. pubescens (Kutaja) still remains a valuable remedy for amoebic infections.
Piles are a common chronic painful anal disease, and a clinical trial (n = 22) used a formulation containing H. pubescens bark (25 mg/capsule) along with six other plants [155]. All patients completed their entire study period of four weeks, and 20 out of 22 patients showed a status of wellbeing (P-0.01). Later, another group [156] also performed a clinical trial for piles, and observed that powder has significant role in stopping the bleeding in the disease Shonitarsha (bleeding piles).
A randomized controlled trial carried out by Kadam et al. [157] using a mixture of 8 herbal plants, including H. pubescens (1.02 g/10 g), for control of dental plaque and gingivitis. Both UDM toothpowder and standard control treatment yielded a statistically significant reduction in scores of gingival index and plaque. Another randomized clinical study with 32 children (3–12 years) with Bhunimbadi-Vati that contains nine herbals including H. pubescens, shows relief of “Mukha Vairasya” (bad taste in mouth) and “Tikta Amlodgara” (sour and better belching efficacy) [158].
The same authors carried out another randomized clinical study in 43 patients with ulcerative colitis; “Kutaja Ghana vati” (H. pubescens, 1 g three times a day) helps in reducing the bowel movement frequency [159]. Johari and Gandhi [160] carried out a randomized single-blind parallel group study comparing a monoherbal formulation containing H. pubescens extract with mesalamine in chronic ulcerative colitis patients, with special emphasis on side effects and relapse. The study supports the efficacy of the monoherbal formulation in resolving chronic ulcerative colitis, with fewer chances of relapse and side effects. However, the authors recommend that the study be used to conduct Phase II and III clinical trials with larger sample sizes. Recently, Kumari et al. [161] studied the efficacy of Kutaja syrup (H. pubescens) on 30 infants suffering from acute diarrhea. The trial drug was given to infants at a dose of 15 mg/kg, every 8 h for two days; the drug had a significant role in reducing signs and symptoms of diarrhea. However, the authors advised a randomized controlled trial with adequate sample size.
Pathadi Kwatha” contains a mixture of five plants, including H. pubescens; it was tested in patients with polycystic ovarian disease (n = 34) in a randomized clinical trial, and proved statistically significantly effective in regularizing menstruation, achieving considerable reduction in body weight, substantial growth of follicles, and thus ovulation [159].
Mundhe et al. [162], performed an open-label, prospective, multi-center clinical study to evaluate the efficacy of Ayuartis capsules (contains H. pubescens stem bark 30 mg, along with several other plants) in patients suffering from osteoarthritis of the knee(s). Three months of treatment with Ayuartis capsule led to a significant reduction in joint pain and joint stiffness. Therefore, this can be an effective treatment option for the management of chronic degenerative joint disorders such as osteoarthritis.
In all trials where multi-herbal treatments were tested, no conclusions can be drawn about the contribution of H. pubescens to the therapeutic effect, if any. Even for H. pubescens monotherapy, the quality and size of the trials preclude definitive conclusions, and more convincing trials are needed, as many of the authors concede.

7. The Way Forward

H. pubescens has been demonstrated by different in vitro studies to have a wide range of medicinal properties, particularly anti-malarial, anti-mutagenic, anti-hypertensive, anti-diarrheal, anti-microbial, CNS-stimulant, diuretic, anti-amoebiasis, anti-urolithic, antioxidant, anti-inflammatory, gut-relaxant and anti-diabetic properties. Several bioactive chemical compounds have been isolated from this plant i.e., conimine, isoconessimine, conessine, conarrhimine and conessimine. In in vitro studies conessimine appeared to have considerable CNS-stimulant activity, as well as antibacterial effects. However, clinical trials on the therapeutic potential of this plant are limited and largely preliminary.
Therefore, it would be useful to explore the individual compounds isolated from H. pubescens in order to validate its ethnomedicinal uses, and to develop clinical applications of this plant. Further efforts are required to identify the active compounds using bioassay-guided purification.

8. Conclusions

H. pubescens is a well-known plant that is mostly used by indigenous communities of Asia. It is used by multiple communities for treating various ailments such as rheumatism, leprosy, skin diseases, diarrhea, dysentery, gastrointestinal infections, stomach-ache, piles, cough and cold, typhoid fever and malaria, etc. Conessine, an active compound of H. pubescens has demonstrated biological properties. Other major bioactive components are holarrhemine, conkurchine, kurchicine, holarrhenine, kurchine, and conkurchinine. From our brief overview, it is evident that several in vitro effects of crude extracts were reported, but in most cases further work is required to isolate and characterize the bioactive compounds. Moreover, except for the antimicrobial and acetylcholinesterase/neuroprotective activity, all other ones need further follow-up, with the mechanism of action and structure-activity relationship studies to assess more fully their potential as drug candidates.

Author Contributions

Conceptualization, S.K.P.; methodology, K.Z.; software, K.Z., S.S.S.; validation, W.L.; formal analysis, S.K.P.; investigation, S.K.P., S.S.S.; resources, K.Z., S.K.P.; data curation, S.K.P., S.S.S. and K.Z.; writing—original draft preparation, K.Z.; writing—review and editing, S.K.P. and W.L.; visualization, S.S.S., K.Z.; supervision, W.L.; project administration, W.L.; funding acquisition, S.K.P. All authors have read and agreed to the published version of the manuscript.


This research received no external funding. This publication was made possible through funding support of the KU Leuven Fund for Fair Open Access


K.Z. was the recipient of a scholarship from the HEC Pakistan. S.K.P. and W.L. largely funded themselves.

Conflicts of Interest

The authors declare no conflict of interest.


  1. Hamilton, A.C. Medicinal plants, Conservation, and livelihoods. Biodiv. Conserv. 2004, 13, 1477–1517. [Google Scholar] [CrossRef]
  2. Balunas, M.J.; Kinghorn, A.D. Drug discovery from medicinal plants. Life Sci. 2005, 78, 431–441. [Google Scholar] [CrossRef] [PubMed]
  3. Royal Botanic Gardens, Kew, Plants of the World online. Available online: (accessed on 16 September 2020).
  4. Jamadagni, P.S.; Pawar, S.D.; Jamadagni, S.B.; Chougule, S.; Gaidhani, S.N.; Murthy, S.N. Review of Holarrhena antidysenterica (L.) Wall. ex A. DC.: Pharmacognostic, Pharmacological, and Toxicological Perspective. Pharmacogn. Rev. 2017, 11, 141–144. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  5. Bharat, K.; Pradhan, K.; Hemant, K.; Badola, K. Ethnomedicinal plant use by Lepcha tribe of Dzongu valley, bordering Khangchendzonga Biosphere Reserve, in North Sikkim, India. J. Ethnobiol. Ethnomed. 2008, 4, 22–40. [Google Scholar]
  6. Panda, S.K. Ethnomedicinal uses and screening of plants for antibacterial activity from Similipal Biosphere Reserve, Odisha, India. J. Ethnopharmacol. 2014, 151, 158–175. [Google Scholar] [CrossRef]
  7. Devi Prasad, A.G.; Shyma, T.B.; Raghavendra, M.P. Plants used by the tribes of for the treatment of digestive system disorders in Wayanad district, Kerala. J. Appl. Pharm. Sci. 2013, 3, 171–175. [Google Scholar]
  8. Manika, N. Identification of Alternative Plant Parts of Some Important Bark Drugs for Sustainable Harvesting. Ph.D. Thesis, University of Lucknow, Lucknow, India, 2014. [Google Scholar]
  9. Ali, K.M.; Chatterjee, K.; De, D.; Bera, T.K.; Ghosh, D. Efficacy of aqueous extract of seed of Holarrhena antidysenterica for the management of diabetes in experimental model rat: A correlative study with antihyperlipidemic activity. Int. J. Appl. Res. Nat. Prod. 2009, 2, 13–21. [Google Scholar]
  10. Ali, K.M.; Chatterjee, K.; De, D.; Jana, K.; Bera, T.K.; Ghosh, D. Inhibitory effect of hydro-methanolic extract of seed of Holarrhena antidysenterica on alpha-glucosidase activity and postprandial blood glucose level in normoglycemic rat. J. Ethnopharmacol. 2011, 135, 194–196. [Google Scholar] [CrossRef]
  11. Aqil, F.; Ahmad, I. Antibacterial properties of traditionally used Indian medicinal plants. Exp. Clin. Pharmacol. 2007, 29, 79–92. [Google Scholar] [CrossRef]
  12. Husain, N.; Trak, T.H.; Chauhan, D. Floristic diversity of Jammu and Kashmir, India, especially in context to skin care. Int. J. Adv. Sci. Technol. 2020, 29, 2358–2386. [Google Scholar]
  13. Diallo, A.; Traore, M.S.; Keita, S.M.; Balde, M.A.; Keita, A.; Camara, M.; Van Miert, S.; Pieters, L.; Balde, A.M. Management of diabetes in Guinean Traditional Medicine: An ethnobotanical investigation in the Coastal Lowlands. J. Ethnopharmacol. 2012, 144, 353–361. [Google Scholar] [CrossRef] [PubMed]
  14. Rout, S.D.; Pandey, A.K. Ethnomedicobiology of Similipal Biosphere Reserve, Orissa. In Advances in Ethnobotany; Das, A.P., Pandey, A.K., Eds.; Bishen Singh Mahendra Pal Singh: Dehera Dun, India, 2007; pp. 247–252. [Google Scholar]
  15. Bikram, K.; Mallik, R.; Panda, T.; Rabindra, N. Traditional herbal practices by the ethnic people of Kalahandi District of Odisha, India. Asian Pac. J. Trop. Biomed. 2012, 2, S988–S994. [Google Scholar]
  16. Dey, A.; Jitendra, N.D. Ethnobotanical survey of Purulia district, West Bengal, India for medicinal plants used against gastrointestinal disorders. J. Ethnopharmacol. 2012, 143, 68–80. [Google Scholar] [CrossRef] [PubMed]
  17. Biswas, A.; Bari, M.A.; Roy, M.; Bhadra, S.K. Inherited folk pharmaceutical knowledge of tribal people in the Chittagong Hill tracts, Bangladesh. Indian J. Tradit. Knowl. 2010, 9, 77–89. [Google Scholar]
  18. Chopra, R.N.; Chopra, I.C.; Handa, K.L.; Kapur, I.D. Chopras Indigenous Drugs of India; Academic Press: New Delhi, India, 1982; p. 342. [Google Scholar]
  19. Rout, S.D.; Panda, T.; Mishra, N. Ethno-medicinal plants used to cure different diseases by tribals of Mayurbhanj district of North Orissa. Stud. Ethno Med. 2009, 3, 27–32. [Google Scholar] [CrossRef]
  20. Jena, M.; Sahoo, S.; Sahu, R.K. Some ethno medicinal plants for the treatment of common health problems in Mayurbhanj District, Orissa, India. N. Y. Sci. J. 2011, 4, 87–92. [Google Scholar]
  21. Gaur, R.D.; Sharma, J.; Painuli, R.M. Plants used in traditional healthcare of livestock by Gujjar community of Sub-Himalayan tracts, Uttarakhand, India. Indian J. Nat. Prod. Resour. 2010, 1, 243–248. [Google Scholar]
  22. Chopra, R.N.; Nayar, S.L.; Chopra, I.C. Glossary of Indian Medicinal Plants; Council of Scientific and Industrial Research: New Delhi, India, 1956. [Google Scholar]
  23. Ghani, A. Medicinal Plants of Bangladesh: Chemical Constituents and Uses, 1st ed.; Asiatic Society of Bangladesh: Dacca, Bangladesh, 1998; pp. 195–196. [Google Scholar]
  24. Dua, V.K.; Gaurav, V.; Bikram, S.; Aswathy, R.; Upma, B.; Dayal, D.; Gupta, N.C.; Sandeep, K.; Ayushi, R. Anti-malarial property of steroidal alkaloid conessine isolated from the bark of Holarrhena antidysenterica. Malar. J. 2013, 12, 194. [Google Scholar] [CrossRef] [Green Version]
  25. Maroyi, A. Holarrhena pubescens Wall. ex G. Don. In Plant Resources of Tropical Africa 11, Medicinal Plants 1; Schmelzer, G.H., Gurib-Fakim, A., Eds.; PROTA Foundation: Wageningen, The Netherlands, 2008; pp. 332–335. [Google Scholar]
  26. Begum, S.; Usmani, S.B.; Siddiqui, B.S.; Siddiqui, S. Alkaloidal constituents of the bark of Holarrhena antidysenterica. Heterocycles 1993, 36, 717–723. [Google Scholar]
  27. Alauddin, M.; Martin-Smith, M. Biological activity in steroids possessing nitrogen atoms. J. Pharm. Pharmacol. 1962, 14, 469–495. [Google Scholar] [CrossRef]
  28. Daniel, M. Medicinal Plants: Chemistry and Properties; Science: Enfield, NH, USA, 2006. [Google Scholar]
  29. Stephenson, R.P. The pharmacological properties of conessine, isoconessine and neoconessine. Br. J. Pharmacol. 1948, 3, 237–245. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  30. Kumar, N.; Singh, B.; Bhandari, P.; Gupta, A.P.; Kaul, V.K. Steroidal alkaloids from Holarrhena antidysenterica (L.) WALL. Chem. Pharma. Bull. 2007, 55, 912–914. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  31. Siddiqui, B.S.; Usmani, S.B.; Ali, S.T.; Begum, S.; Rizwani, G.H. Further constituents from the bark of H. pubescens. Phytochemistry 2001, 58, 1199–1204. [Google Scholar] [CrossRef]
  32. Singh, A.K.; Raghubanshi, A.S. Medical ethnobotany of the tribals of Sonaghati of Sonbhadra district, Uttar Pradesh, India. J. Ethnopharmacol. 2002, 81, 31–41. [Google Scholar] [CrossRef]
  33. Kadir, M.F.; Sayeed, M.S.B.; Mia, M.M.K. Ethnopharmacological survey of medicinal plants used by traditional healers in Bangladesh for gastrointestinal disorders. J. Ethnopharmacol. 2013, 147, 148–156. [Google Scholar] [CrossRef] [PubMed]
  34. Anantha Narayana, D.B. The Ayurvedic Pharmacopoeia of India: Part II. A Good Beginning. (Formulations). Curr. Sci. 2008, 94, 1086–1087. [Google Scholar]
  35. Rudolf, H. Hagers Handbuch der Pharmazeutischen Praxis; Springer: New York, NY, USA, 1976; pp. 92–95. [Google Scholar]
  36. Marius, H.; Yetein, A.N.; Houessou, L.G.; Toussaint, O.; Lougbe´gnon, A.C.; BriceTente, O.K. Ethnobotanical study of medicinal plants used for the treatment of malaria in plateau of Allada, Benin (West Africa). J. Ethnopharmacol. 2013, 146, 154–163. [Google Scholar]
  37. Ghimire, K.A.; Bastakoti, R.R. Ethnomedicinal knowledge and healthcare practices among the Tharus of Nawalparasi district in central Nepal. For. Ecol. Manag. 2009, 257, 2066–2072. [Google Scholar] [CrossRef]
  38. Van Wyk, B.E.; Gericke, N. People’s Plants. A Guide to Useful Plants of Southern Africa; Briza Publications: Pretoria, South Africa, 2000. [Google Scholar]
  39. Steenkamp, V. Traditional herbal remedies used by South African women for gynecological complaints. J. Ethnopharmacol. 2003, 86, 97–108. [Google Scholar] [CrossRef]
  40. Rout, S.D.; Panda, S.K. Ethnomedicinal plant resources of Mayurbhanj district, Orissa. Indian J. Trad. Knowl. 2010, 9, 68–72. [Google Scholar]
  41. Sawadogo, W.R.; Schumacher, M.; Teiten, M.H.; Dicato, M.; Diederich, M. Traditional West African Pharmacopeia, Plants and derived. Biochem. Pharmacol. 2012, 84, 1225–1240. [Google Scholar] [CrossRef] [PubMed]
  42. Koudouvoa, K.; Karoua, D.S.; Kokoua, K.; Essiena, K.; Aklikokoua, K.; Glithob, I.A.; Simporec, J.; Sanogod, R.; De Souzaa, C.; Gbeassora, M. An ethnobotanical study of antimalarial plants in Togo Maritime Region. J. Ethnopharmacol. 2011, 134, 183–190. [Google Scholar] [CrossRef] [PubMed]
  43. Ngarivhume, T.; Charlotte, I.E.A.; Klooster, C.; Joop, T.V.M.; Dejong, C.; Jan, H.V. Westhuizen medicinal plants used by traditional healers for the treatment of malaria in the Chipinge District in Zimbabwe. J. Ethnopharmacol. 2015, 159, 224–237. [Google Scholar] [CrossRef] [Green Version]
  44. Gelfand, M.; Mavi, S.; Drummond, R.B.; Ndemera, B. The Traditional Medical Practitioner in Zimbabwe: His Principles of Practice and Pharmacopoeia; Mambo Press: Gweru, Zimbabwe, 1985. [Google Scholar]
  45. Chhabra, S.C.; Mahunnah, R.L.A.; Mshiu, E.N. Plants used in traditional medicine in Eastern Tanzania. I. Pteridophytes and Angiosperms (Acanthaceae to Canellaceae). J. Ethnopharmacol. 1897, 21, 253–277. [Google Scholar] [CrossRef]
  46. Bruschi, P.; Morganti, M.; Mancini, M.M.A. Signorini traditional healers and laypeople: A qualitative and quantitative approach to local knowledge on medicinal plants in Muda (Mozambique). J. Ethnopharmacol. 2011, 138, 543–563. [Google Scholar] [CrossRef] [PubMed]
  47. Olorunnisola, O.S.; Adetutu, A.; Afolayan, A.J.A. An inventory of plants commonly used in the treatment of some disease conditions in Ogbomoso, South West, Nigeria. J. Ethnopharmacol. 2014, 161, 60–68. [Google Scholar] [CrossRef]
  48. Sheng, P.J. Preliminary study of ethnobotany in Xishuang Banna, People’s Republic of China. J. Pharmacol. 1985, 13, 121–137. [Google Scholar]
  49. Khuankaew, S.; Srithi, K.; Tiansawat, P.; Jampeetong, A.; Inta, A.; Wang, P.P.W. Ethnobotanical study of medicinal plants used by tai yai in Northern Thailand. J. Pharmacol. 2014, 151, 829–838. [Google Scholar] [CrossRef]
  50. Jayaswal, S.B. Amoebicidal activity of steroidal alkaloids of Wrightia tomentosa in vitro. Indian J. Pharm. 1976, 38, 112–113. [Google Scholar]
  51. Juyal, P.; Ghildiyal, M. Medicinal phytodiversity of Bhabar Tract of Garhwal Himalaya. J. Med. Plant. Stud. 2013, 1, 43–57. [Google Scholar]
  52. Girach, R.D.; Singh, S.; Brahmam, M.; Misra, M.K. Traditional treatment of skin diseases in Bhadrak District, Orissa. J. Econ. Taxon. Botany 1999, 27, 754–760. [Google Scholar]
  53. Kumar, M.; Bussmann, R.W.; Mukesh, J.; Kumar, P. Ethnomedicinal uses of plants close to rural habitation in Garhwal Himalaya, India. J. Med. Plant Res. 2011, 11, 2252–2260. [Google Scholar]
  54. Haerdi, F. Die Eingeborenen-Heilpflanzen Des Ulanga-Distriktes Tanganjikas (Ostafrika). Acta Trop. 1964, 8, L-278. [Google Scholar]
  55. Kapur, S.K. Traditionally important medicinal plants of Dudu valley-Jammu. J. Econ. Taxon. Botany 1991, 15, 1–10. [Google Scholar]
  56. Priya, V.K.; Gopalan, R. Ethnomedicinal studies in selected medicinal plants of Dhoni Forest, Western Ghats, Kerala. Asian J. Pharm. Clin. Res. 2014, 7, 3–6. [Google Scholar]
  57. Bhat, P.; Hegde, G.; Ganesh, R. ethnomedicinal practices in different communities of Uttara Kannada district of Karnataka for treatment of wounds. J. Pharmacol. 2012, 143, 501–514. [Google Scholar] [CrossRef]
  58. Mahato, S.; Mehta, A.; Roy, S. Studies on antibacterial effects of bark, seed and callus extracts of Holarrhena antidysenterica Wall. Bioscan 2013, 8, 717–721. [Google Scholar]
  59. Guha Bakshi, D.N.; Sensarma, P.; Pal, D.C. A Lexicon of Medicinal Plants in India; Naya Prokash Publishers: Calcutta, India, 2001; Volume II, pp. 356–358. [Google Scholar]
  60. Panda, S.K.; Bastia, A.K.; Dutta, S.K. Antidiarrheal activity of Terminalia arjuna Roxb from India. J. Biol. Act. Prod. Nat. 2011, 1, 236–247. [Google Scholar]
  61. Hossan, M.S.; Hanif, A.; Agarwala, B.; Sarwar, M.S.; Karim, M.; Taufiq-Urrahman, M.; Jahan, R.; Rahmatullah, M. Traditional use of medicinal plants in Bangladesh to treat urinary tract infections and sexually transmitted diseases. Ethnobot. Res. Appl. 2010, 8, 61–74. [Google Scholar] [CrossRef] [Green Version]
  62. Rahmatullah, M.; Azam, M.N.K.; Malek, I.; Nasrin, D.; Jamal, F.; Rahman, M.A.; Khatun, Z.; Jahan, S.; Seraj, S.; Jahan, R. An ethnomedicinal survey among the Marakh sect of the garo tribe of Mymensingh district, Bangladesh. Int. J. PharmTech Res. 2012, 4, 141–149. [Google Scholar]
  63. Rahmatullah, M.; Ayman, U.; Akter, F.; Sarker, M.; Sifa, R.; Sarker, B.; Chyti, H.N.; Jahan, F.I.; Chowdhury, M.H.; Chowdhury, S.A. Medicinal formulations of a Kanda tribal healer—A tribe on the verge of disappearance in Bangladesh. Afr. J. Trad. Complement Alt. Med. 2013, 10, 213–222. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  64. Tumpa, S.I.; Hossain, M.I.; Ishika, T. Ethnomedicinal uses of herbs by indigenous medicine practitioners of Jhenaidah District, Bangladesh. J. Pharmacogn. Phytochem. 2014, 3, 23–33. [Google Scholar]
  65. Uddin, M.Z.; Hassan, M.A.; Sultana, M. Ethnobotanical survey of medicinal plants in Phulbari Upazila of Dinajpur District, Bangladesh. Bangladesh J. Plant Taxon. 2006, 13, 63–68. [Google Scholar] [CrossRef] [Green Version]
  66. Zerin, Z.; Harun-ar-Rashid, M.; Islam, A.; Khatun, Z.; Rahmatullah, M. Medicinal plants and formulations of a community of the Tonchongya tribe in Bandarban District of Bangladesh. Am. Eur. J. Sustain. Agric. 2012, 6, 292–298. [Google Scholar]
  67. Shaheen, H.D.; Potter, M.F.; Qaseem, A.; Qureshi, R. Heliotropium pakistanicum sp. nov. (Boraginaceae) from Pakistan. Planta Daninha 2019, 37, 1–7. [Google Scholar] [CrossRef] [Green Version]
  68. Qaseem, M.; Qureshi, R.; Amjad, M.S.; Ahmed, W.; Masood, A.; Shaheen, H. Ethnobotanical evaluation of indigenous flora from the communities of Rajh mehal and Goi union councils of district Kotli, Azad Jammu Kashmir, Pakistan. Appl. Ecol. Environ. Res. 2019, 17, 2799–2829. [Google Scholar] [CrossRef]
  69. Qureshi, R. Medicinal Uses of Trees and Shrubs by the Inhabitants of Nara Desert, Pakistan. In Plant and Human Health; Volume 1: Ethnobotany and Physiology; Ozturk, M., Hakeem, K.R., Eds.; Springer International Publishing Ag, Springer Nature: Cham, Switzerland, 2018; pp. 391–407. [Google Scholar]
  70. Khan, A.M.; Qureshi, R.; Saqib, Z. Multivariate analyses of the vegetation of the western Himalayan forests of Muzaffarabad District, Azad Jammu and Kashmir, Pakistan. Ecol. Indic. 2019, 104, 723–736. [Google Scholar] [CrossRef]
  71. Rahmatullah, M.; Jahan, R.; Hossan, M.S.; Seraj, S.; Rahman, M.M.; Chowdhury, A.R.; Begum, R.; Nasrin, D.; Khatun, Z.; Hossain, M.S.; et al. A Comparative analysis of medicinal plants used by three tribes of Chittagong Hill tracts region, Bangladesh to treat leucorrhea. Adv. Nat. Appl. Sci. 2010, 2, 148–152. [Google Scholar]
  72. Dubey, D.; Padhy, R.N. Surveillance of multidrug resistance of two Gram-positive pathogenic bacteria in a teaching hospital and in vitro efficacy of 30 ethnomedicinal plants used by an aborigine of India. Asian Pac. J. Trop. Dis. 2012, 2, 273–281. [Google Scholar] [CrossRef]
  73. Rajakumar, N.; Shivanna, M.B. Ethno-medicinal application of plants in the eastern region of Shimoga District, Karnataka, India. J. Pharmacol. 2009, 126, 64–73. [Google Scholar] [CrossRef]
  74. Khaleel, S.B.; Sudarsanam, G. Traditional use of plants against snakebite in Sugali tribes of Yerramalais of Kurnool District, Andhra Pradesh, India. Asian Pac. J. Trop. Biomed. 2012, 2, 575–579. [Google Scholar]
  75. Jain, S.P.; Puri, H.S. Ethnomedicinal plants of Jaunsar-Bawar Hills, Uttar Pradesh, India. J. Pharmacol. 1984, 12, 213–222. [Google Scholar] [CrossRef]
  76. Painuli, R.M.; Maheswari, J.K. Medicinal plants used by tribals of Panchmahals District, Gujarat. Ancient Sci. Life 1994, 3, 253–258. [Google Scholar]
  77. Harpreet, B.; Sharma, Y.A.; Manhas, R.K.; Kumar, K. Ethnomedicinal plants used by the villagers of District Udhampur, J&K, India. J. Pharmacol. 2014, 151, 1005–1018. [Google Scholar]
  78. Fotie, J.; Bohle, D.S.; Leimanis, M.L.; Georges, E.; Rukunga, G.; Nkengfack, A.E. Lupeol long-chain fatty acid esters with antimalarial activity from Holarrhena floribunda. J. Nat. Prod. 2006, 69, 62–67. [Google Scholar] [CrossRef]
  79. Mahishi, P.; Srinivasa, B.H.; Shivanna, M.B. Medicinal plant wealth of local communities in some villages in Shimoga District of Karnataka, India. J. Pharmacol. 2005, 98, 307–312. [Google Scholar] [CrossRef]
  80. Gangwar, K.K.; Deepali, G.R.; Gangwar, R.S. Ethnomedicinal plant diversity in Kumaun Himalaya of Uttarakhand, India. Nat. Sci. 2010, 8, 66–78. [Google Scholar]
  81. Kabir, A.K.L.; Begum, M.M.; Islam, T. Study of Bioactivities of Holarrhena pubescens growing in Bangladesh. Dhaka Univ. J. Pharm. Sci. 2018, 17, 131–137. [Google Scholar] [CrossRef]
  82. Rajakumar, N.; Shivanna, M.B. Traditional veterinary healthcare practices in Shimoga District of Karnataka, India. Indian J. Trad. Knowl. 2012, 11, 283–287. [Google Scholar]
  83. Sanjib, S.; Shil, N.; Choudhury, M.S.; Das, S. Indigenous knowledge of medicinal plants used by the Reang tribe of Tripura state of India. J. Pharmacol. 2014, 152, 135–141. [Google Scholar]
  84. Tarafdar, R.G.; Nath, S.; Talukdar, A.D.; Manabendra, D.C. Antidiabetic plants used among the ethnic communities of Unakoti District of Tripura, India. J. Pharmacol. 2015, 160, 219–226. [Google Scholar] [CrossRef] [PubMed]
  85. Sen, S.; Chakraborty, R.; De, B.; Devanna, N. An ethnobotanical survey of medicinal plants used by ethnic people in west and south District of Tripura, India. J. For. Res. 2011, 22, 417–426. [Google Scholar] [CrossRef]
  86. Shukla, A.N.; Srivastava, S.; Rawat, A.K.S. A survey of traditional medicinal plants of Uttar Pradesh (India)-used in treatment of infectious diseases. Nat. Sci. 2013, 11, 24–36. [Google Scholar]
  87. Tripathi, S.; Mondal, A.K.; Verma, K. Rare ethno medicinal plants of South West Bengal, India with their different medicinal uses: Needs conservation. Int. J. Life Sci. Pharm. Res. 2013, 2, 114–122. [Google Scholar]
  88. Sikarwar, R.L.S.; Pathak, B.; Jaiswal, A. Some unique ethnomedicinal perceptions of tribal communities of Chitrakoot, Madhya Pradesh. Indian J. Trad. Knowl. 2008, 7, 613–617. [Google Scholar]
  89. Kosalge, S.B.; Fursule, R.A. Investigation of ethnomedicinal claims of some plants used by tribals of Satpuda Hills in India. J. Pharmacol. 2009, 121, 456–461. [Google Scholar] [CrossRef]
  90. Sen, S.K.; Pattnaik, M.R.; Behera, L.M. Traditional use of herbal medicines against rheumatism by the tribals of Bargarh District in Odisha (India). Life Sci. Leaflets 2014, 51, 59–68. [Google Scholar]
  91. Mallick, S.N.; Ram, J.P.; Parida, N. Study of ethnomedicinal values of some shrubs in Rourkela steel city and its surroundings, Sundargarh, Odisha. Int. J. Appl. Biol. Pharm. Technol. 2014, 5, 123–130. [Google Scholar]
  92. Rout, S.; Panda, S.P.; Patra, H.K. Ethnomedicinal studies on Bondo tribe of Malkangiri District, Odisha, India. Int. J. Biodiv. Conserv. 2014, 6, 326–332. [Google Scholar]
  93. Panda, S.K.; Patro, N.; Sahoo, G.; Bastia, A.K.; Dutta, S.K. Anti-diarrheal activity of medicinal plants of Similipal Biosphere Reserve, Odisha, India. Int. J. Med. Aromat. Plants 2012, 1, 123–134. [Google Scholar]
  94. Panda, S.K.; Bastia, A.K.; Sahoo, G. Process characteristics and nutritional evaluation of handia—A cereal based ethnic fermented food from Odisha. Indian J. Trad. Knowl. 2014, 13, 149–156. [Google Scholar]
  95. Panda, S.K.; Rout, S.D.; Mishra, N.; Panda, T. Phytotherapy and traditional knowledge of tribal communities of Mayurbhanj district, Orissa, India. J. Pharmacog. Phytother. 2011, 3, 101–113. [Google Scholar]
  96. Panda, S.K.; Padhi, L.; Leyssen, P.; Liu, M.; Neyts, J.; Luyten, W. Antimicrobial, anthelmintic, and antiviral activity of plants traditionally used for treating infectious disease in the Similipal Biosphere Reserve, Odisha, India. Front. Pharmacol. 2017, 8, 658. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  97. Padal, S.B.; Viyayakumar, Y. Traditional knowledge of Valmiki tribes of G. Madugula Mandalam, Visakhapatnam District, Andhra Pradesh. Int. J. Innov. Res. Dev. 2013, 2, 723–738. [Google Scholar]
  98. Manjula, R.R.; Koteswara Rao, J.; Seetharami Reddi, T.V.V. Ethnomedicinal plants used to cure jaundice in Khammam district of Andhra Pradesh, India. J. Phytol. 2011, 3, 33–35. [Google Scholar]
  99. Sandhya Sri, B.; Padal, S.B.; Ramakrishna, B. New traditional phytotherapy for gynecological disorders among the tribes of Visakhapatnam District, Andhra Pradesh, India. BMR Phytomed. 2014, 2, 1–7. [Google Scholar]
  100. Rajakumar, N.; Shivanna, M.B. Traditional herbal medicinal knowledge in Sagar Taluk of Shimoga District, Karnataka, India. Indian J. Nat. Prod. Res. 2010, 1, 102–108. [Google Scholar]
  101. Shivanna, M.B.; Rajakumar, N. Traditional medico-botanical knowledge of local communities in Hosanagara Taluk of Shimoga District in Karnataka, India. J. Herbs Spices Med. Plant. 2011, 17, 291–317. [Google Scholar] [CrossRef]
  102. Shivanna, M.B.; Rajakumar, N. Ethno-medico-botanical knowledge of rural folk in Bhadravathi taluk of Shimoga district, Karnataka. Indian J. Trad. Knowl. 2010, 9, 158–162. [Google Scholar]
  103. Bhandary, M.J.; Chandrashekar, K.R.; Kaveriappa, K.M. Medical ethnobotany of the Siddis of Uttara Kannada district, Karnataka, India. J. Pharmacol. 1995, 47, 149–158. [Google Scholar] [CrossRef]
  104. Sharma, J.; Gairola, S.; Sharma, Y.P.; Gaur, R.D. Ethnomedicinal plants used to treat skin diseases by Tharu community of District Udham Singh Nagar, Uttarakhand, India. J. Pharmacol. 2014, 158, 140–206. [Google Scholar] [CrossRef] [PubMed]
  105. Gairola, S.; Sharma, J.; Gaur, R.D.; Siddiqi, T.O.; Painuli, R.M. Plants used for treatment of dysentery and diarrhea by the Bhoxa community of district Dehradun, Uttarakhand, India. J. Pharmacol. 2013, 150, 989–1006. [Google Scholar]
  106. Sharma, J.; Gairola, S.; Gaur, R.D.; Painuli, R.M. The treatment of jaundice with medicinal plants in indigenous communities of the Sub-Himalayan region of Uttarakhand, India. J. Pharmacol. 2012, 143, 262–291. [Google Scholar] [CrossRef]
  107. Kunwar, R.M.; Uprety, Y.; Burlakoti, C.; Chowdhary, C.L.; Bussmann, R.W. Indigenous use and ethnopharmacology of medicinal plants in Far-west Nepal. Ethnobot. Res. Appl. 2009, 7, 005–028. [Google Scholar] [CrossRef] [Green Version]
  108. Sheikh, Y.; Manral, M.S.; Kathait, V.; Prasar, B.; Kumar, R.; Sahu, R.K. Computation of in vivo antidiabetic activity of Holarrhena antidysenterica seeds extracts in Streptozotocin-induced diabetic rats. Iranian J. Pharmacol. Ther. 2016, 14, 22–27. [Google Scholar] [CrossRef]
  109. Keshri, U. Antidiabetic efficacy of ethanolic extract of Holarrhena antidysenterica seeds in streptozotocin induced diabetic rats and influence on certain biochemical parameters. J. Drug Deliv. Ther. 2012, 2, 159–162. [Google Scholar] [CrossRef]
  110. Bandawane, D.D.; Bibave, K.H.; Jaydeokar, A.V.; Patil, U.S.; Hivrale, M.G. Antihyperglycemic and antihyperlipidemic effects of methanolic extract of Holarrhena antidysenterica bark in alloxan induced diabetes mellitus in rats. Pharmacologia 2013, 4, 95–106. [Google Scholar] [CrossRef]
  111. Hegde, K.; Jaisal, K.K. Anti-diabetic potential of ethanolic extract of Holarrhena antidysenterica Linn leaves. Int. J. Pharm. Res. 2014, 5, 429–435. [Google Scholar]
  112. Bibave, K.H.; Bandawane, D.D.; Patil, U.S. Evaluation of in vitro antioxidant and in vivo antihyperglycemic effects of Holarrhena antidysenterica bark. J. Pharm. Res. 2012, 5, 5076–5080. [Google Scholar]
  113. Bhusal, A.; Jamarkattel, N.; Shrestha, A.; Lamsal, N.K.; Shakya, S.; Rajbhandari, S. Evaluation of antioxidative and antidiabetic activity of bark of H. pubescens wall. J. Clin. Diagn. Res. 2014, 8, HC05–HC08. [Google Scholar] [CrossRef]
  114. Sharma, D.K.; Gupta, V.K.; Kumar, S.; Joshi, V.; Mandal, R.S.; Prakash, A.G.; Singh, M. Evaluation of antidiarrheal activity of ethanolic extract of Holarrhena antidysenterica seeds in rats. Vet. World 2015, 8, 1392–1395. [Google Scholar] [CrossRef] [PubMed]
  115. Daswani, P.G.; Birdi, T.J.; Antarkar, D.S.; Antia, N.H. Investigation of antidiarrhoeal activity of Holarrhena antidysentrica. Int. J. Pharm. Res. 2012, 64, 164–167. [Google Scholar]
  116. Saha, S.; Subrahmanyam, E.V.S. Evaluation of anti-inflammatory activity of ethanolic extract of seeds of (Holarrhena pubescens Buch.- Ham.) wall. Int. J. Pharm. Pharm. Sci. 2013, 5, 915–919. [Google Scholar]
  117. Khan, A.; Khan, S.R.; Gilani, A.H. Studies on the in vitro and in vivo antiurolithic activity of Holarrhena antidysenterica. Urol. Res. 2012, 40, 671–681. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  118. Kavitha, D.; Shilpa, P.N.; Devaraj, S.N. Antibacterial and antidiarrhoeal effects of alkaloids of Holarrhena antidysenterica WALL. Indian J. Exp. Biol. 2004, 42, 589–594. [Google Scholar] [PubMed]
  119. Srivastava, N.; Saxena, V. Antibacterial activity of Kutaj (Holarrhena antidysenterica L.) in childhood diarrhoea—In vitro study. Pharm. Innov. J. 2015, 4, 97–99. [Google Scholar]
  120. Darji, V.C.; Deshpande, S.; Bariya, A.H. Effects of methanolic extract of Holarrhena antidysenterica bark against experimentally induced inflammatory bowel disease in rats. Int. Res. J. Pharm. 2012, 3, 152–154. [Google Scholar]
  121. Haque, A.; Islam, A.U. Evaluation of analgesic and central nervous system depressant effects of Holarrhena antidysenterica stem on Swiss albino mice model. Bangladesh Pharm. J. 2017, 20, 205–212. [Google Scholar] [CrossRef] [Green Version]
  122. Ganapathy, S.; Ramachandra, Y.L.; Padmalatha, R. Anti-inflammatory and analgesic activities of Holarrhena antidysenterica Wall. leaf extract in experimental animal models. Int. J. Biomed. Pharm. Sci. 2011, 4, 101–103. [Google Scholar]
  123. Bhuiyan, M.; Bhuiya, N.; Hasan, M.N.; Nahar, U.J. In vivo and in silico evaluation of antinociceptive activities of seed extract from the Holarrhena antidysenterica plant. Heliyon 2020, 6, e03962. [Google Scholar] [CrossRef]
  124. Ribeiro, R.A.; Vale, M.L.; Thomazzi, S.M.; Paschoalato, A.B.P.; Poole, S.; Ferreira, S.H.; Cunha, F.Q. Involvement of resident macrophages and mast cells in the writing nociceptive response induced by zymosan and acetic acid in mice. Eur. J. Pharmacol. 2000, 387, 111–118. [Google Scholar] [CrossRef]
  125. Ganapathy, S.; Ramachandra, Y.L.; Padmalatha, R. In vitro antioxidant activity of Holarrhena antidysenterica Wall. methanolic leaf extract. J. Basic Clin. Pharm. 2011, 2, 175–178. [Google Scholar]
  126. Zahin, M.; Farrukh, A.; Iqbal, A. The in vitro antioxidant activity and total phenolic content of four Indian medicinal plants. Int. J. Pharm. Pharm. Sci. 2009, 1, 88–95. [Google Scholar]
  127. Khan, A.; Bashir, S.; Gilani, A.H. An in vivo study on the diuretic activity of Holarrhena antidysenterica. Afr. J. Pharm. Pharmacol. 2012, 6, 454. [Google Scholar]
  128. Gilani, A.H.; Khan, A.; Khan, A.U.; Bashir, S.; Rehman, N.U.; Mandukhail, S.U. Pharmacological basis for the medicinal use of Holarrhena antidysenterica in gut motility disorders. Pharm. Biol. 2010, 48, 1240–1246. [Google Scholar] [CrossRef] [PubMed]
  129. Yang, Z.D.; Duan, D.Z.; Xue, W.W.; Yao, X.J.; Li, S. Steroidal alkaloids from Holarrhena antidysenterica as acetylcholinesterase inhibitors and the investigation for structure activity relationships. Life Sci. 2012, 90, 929–933. [Google Scholar] [CrossRef] [PubMed]
  130. Solanki, R.; Madat, D.; Chauhan, K. Evaluation of central nervous system activity of Holarrhena antidysenterica Linn Apocynaceae bark. J. Pharm. Res. 2011, 4, 1760–1761. [Google Scholar]
  131. Siddiqui, B.S.; Ali, S.T.; Rizwani, G.H.; Begum, S.; Tauseef, S.; Ahmad, A. Antimicrobial activity of the methanolic bark extract of H. pubescens (Buch. Ham), its fractions and the pure compound conessine. Nat. Prod. Res. 2012, 26, 987–992. [Google Scholar] [CrossRef]
  132. Somanadhan, B.; Varughese, G.; Palpu, P. An ethnopharmacological survey for potential angiotensin converting enzyme inhibitors from Indian medicinal plants. J. Pharmacol. 1999, 65, 103–112. [Google Scholar] [CrossRef]
  133. Raman, M.D.; Sultana, N.; Anwar, A. In vitro antimicrobial activity of Holarrifine 24-ol isolated from the stem bark of Holarrhena antidysenterica. Int. J. Agric. Biol. 2004, 6, 698–700. [Google Scholar]
  134. Phatthalung, P.N.; Chusri, S.; Voravuthikunchai, S.P. Thai ethnomedicinal plants as resistant modifying agents for combating Acinetobacter baumannii infections. BMC Complement. Alt. Med. 2012, 12, 56. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  135. Chusri, S.; Na-Phatthalung, P.; Siriyong, T.; Paosen, S.; Voravuthikunchai, S.P. Holarrhena antidysenterica as a resistance modifying agent against Acinetobacter baumannii: Its effects on bacterial outer membrane permeability and efflux pumps. Microbiol. Res. 2014, 169, 417–424. [Google Scholar] [CrossRef] [PubMed]
  136. Panda, S.K.; Bastia, A.K. Antimicrobial efficacy of potential plants used in the indigenous preparation of traditional rice beverage “Handia”. Int. J. Phytomed. 2014, 6, 23–28. [Google Scholar]
  137. Siriyong, T.; Voravuthikunchai, S.P.; Coote, P.J. Steroidal alkaloids and conessine from the medicinal plant Holarrhena antidysenterica restore antibiotic efficacy in a Galleria mellonella model of multidrug-resistant Pseudomonas aeruginosa infection. BMC Complement. Alt. Med. 2018, 18, 285. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  138. Siriyong, T.; Chusri, S.; Srimanote, P.; Tipmanee, V.; Voravuthikunchai, S.P. Holarrhena antidysenterica extract and its steroidal alkaloid, conessine, as resistance-modifying agents against extensively drug-resistant Acinetobacter baumannii. Microb. Drug Resist. 2016, 22, 273–282. [Google Scholar] [CrossRef]
  139. Aqil, F.; Zahin, M.; Ahmad, I. Antimutagenic activity of methanolic extracts of four Ayurvedic medicinal plants. Indian J. Exp. Biol. 2008, 46, 668–672. [Google Scholar]
  140. Verma, G.; Dua, V.K.; Agarwal, D.D.; Atul, P.K. Anti-malarial activity of Holarrhena antidysenterica and Viola canescens, plants traditionally used against malaria in the Garhwal region of north-west Himalaya. Malar. J. 2011, 10, 20. [Google Scholar] [CrossRef] [Green Version]
  141. Nondo, R.; Moshi, M.; Paul, M.P.; Machumi, F.; Kidukuli, A.; Heydenreich, M.; Zofou, D. Anti-plasmodial activity of Norcaesalpin D and extracts of four medicinal plants used traditionally for treatment of malaria. BMC Complement. Alt. Med. 2017, 17, 167. [Google Scholar] [CrossRef] [Green Version]
  142. Simonsen, H.T.; Nordskjold, J.B.; Smitt, U.W.; Nyman, U.; Palpu, P.; Joshi, P.; Varughese, G. In vitro screening of Indian medicinal plants for anti-plasmodial activity. J. Pharmacol. 2001, 74, 195–204. [Google Scholar]
  143. Itoh, H.; Inoue, M. Comprehensive Structure-Activity Relationship studies of macrocyclic natural products enabled by their total syntheses. Chem. Rev. 2019, 119, 10002–10031. [Google Scholar] [CrossRef]
  144. Ҫiçek, S.S. Structure-Dependent Activity of plant-derived sweeteners. Molecules 2020, 25, 1946. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  145. Cheenpracha, S.; Jitonnom, J.; Komek, M.; Ritthiwigrom, T.; Laphookhieo, S. Acetylcholinesterase inhibitory activity and molecular docking study of steroidal alkaloids from Holarrhena pubescens barks. Steroids 2016, 108, 92–98. [Google Scholar] [CrossRef] [PubMed]
  146. Zhao, C.; Sun, M.; Bennani, Y.L.; Cowart, M.D.; Hancock, A.A.; Miller, T.R.; Witte, D.G.; Browman, K.E.; Krueger, K.M.; Marsh, K.C.; et al. The alkaloid conessine and analogues as potent histamine H3 receptor antagonists. J. Med. Chem. 2008, 51, 5423–5430. [Google Scholar] [CrossRef] [PubMed]
  147. Swain, S.S.; Paidesetty, S.K.; Padhy, R.N. Development of antibacterial conjugates using sulfamethoxazole with monocyclic terpenes: A systematic medicinal chemistry based computational approach. Comput. Methods Programs Biomed. 2017, 140, 185–194. [Google Scholar] [CrossRef]
  148. Swain, S.S.; Paidesetty, S.K.; Dehury, B.; Das, M.; Vedithi, S.C.; Padhy, R.N. Computer-aided synthesis of dapsone-phytochemical conjugates against dapsone-resistant Mycobacterium leprae. Sci. Rep. 2020, 10, 6839. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  149. Pathak, V.K.; Maiti, A.; Gupta, S.S.; Shukla, I.; Rao, C.V. Effect of the standardized extract of Holarrhena antidysenterica seeds against Streptozotocin-induced diabetes in rats. Int. J. Pharm. Sci. Rev. Res. 2015, 4, 1–6. [Google Scholar]
  150. Singh, R.K. Pre-clinical toxicity studies of Holarrhena antidysenterica stem bark in mice and rats. World J. Pharm. Pharm. Sci. 2018, 7, 912–922. [Google Scholar]
  151. Kumar, S.; Yadav, A. Comparative study of hypoglycemic effect of Holarrhena antidysenterica seeds and glibenclamide in experimentally induced diabetes mellitus in albino rats. Biomed. Pharm. J. 2015, 8, 477–483. [Google Scholar] [CrossRef]
  152. Permpipat, U.; Chavalittumrong, P.; Attawish, A.; Chuntapet, P. Toxicity study of Holarrhena antidysenterica Wall. Bark. Bull. Dep. Med. Sci. 2012, 40, 145–157. [Google Scholar]
  153. Chopra, R.N.; De, N. The failure of the alkaloids of Holarrhena antidysenterica (kurchi) in the treatment of amoebic hepatitis. Indian Med. Gaz. 1930, 65, 391. [Google Scholar]
  154. Singh, K.P. Clinical studies on amoebiasis and giardiasis evaluating the efficacy of kutaja (Holarrhena antidysenterica) in Entamoeba histolytica cyst passers. Anc. Sci. Life 1986, 5, 228–231. [Google Scholar] [PubMed]
  155. Prakash, P.; Pralhad, P.; Nishikant, J. Efficacy of an indigenous formulation in patients with bleeding piles: A preliminary clinical study. Fitoterapia 2000, 71, 41–45. [Google Scholar]
  156. Atanu, P.; Sharma, P.P.; Mukherjee, P.K. A clinical study of (Holarrhena antidysentrica Wall) on Shonistarsha. AYU 2009, 30, 369–372. [Google Scholar]
  157. Kadam, A.; Prasad, B.S.; Bagadia, D.; Hiremath, V.R. Effect of Ayurvedic herbs on control of plaque and gingivitis: A randomized controlled trial. AYU 2011, 32, 532–535. [Google Scholar] [PubMed]
  158. Patel, M.V.; Patel, K.B.; Gupta, S.N. Effects of Ayurvedic treatment on forty-three patients of ulcerative colitis. AYU 2010, 31, 478–481. [Google Scholar] [CrossRef]
  159. Patel, K.; Dei, L.; Donga, S.; Nalini, A. Effect of Shatapushpa Taila Matra Basti and Pathadi Kwatha on poly cystic ovarian disease. AYU 2012, 33, 243–246. [Google Scholar] [CrossRef] [Green Version]
  160. Johari, S.; Gandhi, T. A randomized single blind parallel group study comparing monoherbal formulation containing Holarrhena antidysenterica extract with mesalamine in chronic ulcerative colitis patients. Anc. Sci. Life 2016, 36, 19–27. [Google Scholar] [CrossRef]
  161. Kumari, V.; Singh, B.M.; Kumar, A.; Singh, G. Assessment of effect of Kutaja (Holarrhena antidysenterica Wall) in different Doshika Atisara in infants: A clinical study. J. Res. Ed. Indian Med. 1982, 1, 1–6. [Google Scholar]
  162. Mundhe, N.B.; Tamoli, S.M.; Pande, S.P.; Kulkarni, S.A.; Patil, V.G.; Mahadik, S.B. An open-label, prospective, multicenter, clinical study to evaluate efficacy of Ayuartis capsules in patients suffering from osteoarthritis of the knee(s). AYU 2019, 40, 16–22. [Google Scholar]
Figure 1. Worldwide distribution of H. pubescens [3].
Figure 1. Worldwide distribution of H. pubescens [3].
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Figure 2. Medicinal use of H. pubescens in India, Pakistan, Bangladesh and Nepal.
Figure 2. Medicinal use of H. pubescens in India, Pakistan, Bangladesh and Nepal.
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Figure 3. Schematic representation of the Structural-Activity-Relationship (SAR) of the steroid-alkaloid class of phytoconstituents; conarrhimine, conessimine, conessine, conimine and isoconnessine, isolated from H. pubescens. IC50 expressed in µM, range of 4 to >300 for acetylcholinesterase (AChE)/neuroprotective activity.
Figure 3. Schematic representation of the Structural-Activity-Relationship (SAR) of the steroid-alkaloid class of phytoconstituents; conarrhimine, conessimine, conessine, conimine and isoconnessine, isolated from H. pubescens. IC50 expressed in µM, range of 4 to >300 for acetylcholinesterase (AChE)/neuroprotective activity.
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Figure 4. Schematic representation of the Structural-Activity-Relationship (SAR) of the steroid-alkaloid class of phytoconstituents; Mokluangin A-C and antidysentericine, isolated from H. pubescens. IC50 expressed in µM, range of 1.44 to 23.22 for acetylcholinesterase activity.
Figure 4. Schematic representation of the Structural-Activity-Relationship (SAR) of the steroid-alkaloid class of phytoconstituents; Mokluangin A-C and antidysentericine, isolated from H. pubescens. IC50 expressed in µM, range of 1.44 to 23.22 for acetylcholinesterase activity.
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Figure 5. Three-dimensional molecular interaction of connesine with six different biological targets using the software, BIOVIA-DSV after a blind molecular docking study using software, AutoDock 4.2. Herein each protein data bank (PDB) ID represents the putative target proteins’ crystallographic structural information. PDB ID: 1HNJ, beta-ketoacyl-acyl carrier protein synthase III (FabH) of E. coli; PDB ID: 1C2B, acetylcholinesterase (AChE) of E. electricus; PDB ID: 5NN4, human lysosomal acid α glucosidase (GAA); PDB ID: 6TZ6, calcineurin catalytic (CnA) of Candida albicans; PDB ID: 5F19, human cyclooxygenase-2 (COX-2) and PDB ID: 1LDG, L-Lactate dehydrogenase (LDH) of Plasmodium falciparum.
Figure 5. Three-dimensional molecular interaction of connesine with six different biological targets using the software, BIOVIA-DSV after a blind molecular docking study using software, AutoDock 4.2. Herein each protein data bank (PDB) ID represents the putative target proteins’ crystallographic structural information. PDB ID: 1HNJ, beta-ketoacyl-acyl carrier protein synthase III (FabH) of E. coli; PDB ID: 1C2B, acetylcholinesterase (AChE) of E. electricus; PDB ID: 5NN4, human lysosomal acid α glucosidase (GAA); PDB ID: 6TZ6, calcineurin catalytic (CnA) of Candida albicans; PDB ID: 5F19, human cyclooxygenase-2 (COX-2) and PDB ID: 1LDG, L-Lactate dehydrogenase (LDH) of Plasmodium falciparum.
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Table 1. Medicinal properties of H. pubescens.
Table 1. Medicinal properties of H. pubescens.
DiseaseMedicinal PropertyReference
Intestinal parasitesAnthelmintic for Guinea worm, roundworm, tapeworm, thread worm, other internal worms[5]
Animal bitesAntidote for snake bite, scorpion sting, insect bite, dog bite[6]
IndigestionAppetizer, stomachic[7]
Blood-related ailmentsAnemia, blood infection, blood purifier, hemorrhage, nose bleeding, hypertension[8]
Body painAnalgesic for backache, body ache, headache, knee pain and rheumatic arthritis[9]
Brain-related disordersImproves depression and other nervous disorders, acts as memory enhancer [10]
Cold and throat-related ailmentsExpectorant for cold, cough, throat infection[9,10]
Dental or oral ailmentsAnalgesic for toothache[11]
Dermatological problemsActivity against abscess, acne, boils, bruises, dermatitis, leukoderma, pimples, ringworm, scabies, skin allergies, warts[12]
DiabetesRegulates blood sugar[6,13]
FeverAntipyretic, febrifuge for intermittent fever, pyrexia[12]
Gastrointestinal disordersActive against (hyper)acidity, intestinal ulcers, stomachache, dyspepsia, flatulence, cholera, diarrhea, dysentery, food poisoning, gastroenteritis, colic complaints, constipation[7]
General healthMuscle strength, obesity, tonic[14,15]
Gynecological disordersEasy delivery, leucorrhea, toning up vaginal tissues after delivery[16,17]
Joint- and muscle-related ailmentsActive against arthritis, rheumatism[18,19]
Liver complaintsUseful for bilious disorders, bile infection, jaundice[20,21]
PilesActive against piles, fissures, fistula, hemorrhoids[22,23]
Respiratory disordersActive against asthma, bronchitis[24,25]
Urogenital disordersControls urination, cystitis, diuretic, dysuria, urinary problem, urinary tract infection, urine tract burning sensation[25]
Table 2. Common traditional uses of H. pubescens throughout different parts of the world.
Table 2. Common traditional uses of H. pubescens throughout different parts of the world.
Geographic LocationCondition TreatedPlant Part UsedMethod(s) of PreparationDosage Forms, and Method(s) of AdministrationReference
East AfricaFeverLeaves, rootsDecoctionBath is taken[12]
MalariaRootsDecoctionTaken in the form of drink twice daily[21]
Southern AfricaConstipation, abdominal painsRootInfusionDrink[38]
ToothacheStem, barkDecoctionGargle[40]
SnakebiteRootBoiled in milkApplied externally
West AfricaStomach painsLeavesMacerationDrink[40,41]
Togo Maritime regionMalariaLeaves, rootsDecoctionOral administration[42]
ZimbabweAbortifacient/venereal diseasesRootInfusionsOral administration[43]
MalariaRootDecoctionOral administration[34,44]
TanzaniaAbdominal painRootsDecoctionTaken in the form of drink on empty stomach[45]
MozambiqueStomachache/vomitingLeaves, rootsMacerationOral administration[46]
EaracheAll partsMacerationDirectly applied in the form of ear drops
GuineaDiabetesWhole plantNot statedNot stated[13]
South West NigeriaInflammatory diseasesLeavesInfusionOral administration[47]
Republic of ChinaDiarrhea, dysenteryBarkDecoctionOral administration[48]
Northern ThailandDiarrhea and weight lossStem, barkBoiledOral administration[49]
IndiaLow feverSeedsPowderOral administration, 2–3 g mixed in one glass of water[50,51]
Knee painBarkDecoctionOral administration, mixed with about 100 g of jaggery[52,53]
LeprosySeedsDecoctionOral administration[54,55]
SnakebiteRootsPasteDirectly applied to bite wound[56]
DysenteryBark, leavesPowderTaken with water[57,58]
Amoebic dysenteryBarkPowderOral administration[59,60]
NepalParalysisBark, rootPowderOne spoonful powder or paste from a mixture of (5 g H. pubescens root, 5 g Terminalia alata bark, 2 g Cissampelos pareira root, 5 g H. pubescens bark, 2 g Psidum guajava bark, 1 g Allium sativum bulb and 2 g Trachyspermum ammi seeds), given once a day[37]
Backache, high feverBarkInfusionOral administration[4]
BangladeshBloody dysenteryBarkBoil1 cupful bark of H. pubescens is boiled with 4 cups of water to make 1 cup. A 1.5 mL solution with trace amount of honey is licked 3–4 times daily till cure[61]
Stomach pain, food poisoningBarkMacerationA red-hot iron rod is dipped in the juice, and the juice is taken while still warm[62]
Bark Mixed with bark of Cinnamomum camphora and chewed.[63]
JaundiceLeavesMacerated juiceJuice obtained from leaves of Cajanus cajan and H. pubescens are mixed with powdered seeds of Plantago ovata and taken (one glassful) in the morning on an empty stomach for one month[64]
HelminthiasisSeedsPowderTaken with cold water every morning[65]
PilesBarkPowderMixed with honey and taken orally
Abdominal pain, diarrheaBarkJuiceA ½ cup is taken 2–3 times orally[66]
AsthmaRootJuiceTaken 4–5 times daily for a week
Abdominal painBark/leafJuice2–3 spoons along with honey on empty stomach
PakistanDiabetesRootPowderSalacia reticulate, Annona squamosa and H. pubescens roots were Ground with lime and taken orally[67]
MalariaRootDecoctionOral administration[68]
DiarrheaBarkDecoctionOral administration[69]
Gut infectionsLeavesJuiceTaken daily[70,71]
Table 3. Regional uses of H. pubescens by traditional healers across India.
Table 3. Regional uses of H. pubescens by traditional healers across India.
State/Province, Tribe(s)Disease/IndicationDosage Forms, and Method(s) of AdministrationReferences
Tripura state, reang tribesDog bitePills prepared from bark[83]
Unakoti districtAntidiabetic-[84]
West and south district of TripuraDysentery, fever, cold and piles-[85]
Uttar Pradesh stateDysenteryBark decoction[86]
Sonaghati of Sonbhadra districtStimulate discharge of urine and to remove constipation10–20 g of root paste is taken orally with water[32]
Jaunsar-bawar hillsDysentery and stomachacheDry stem bark mixed with dried ginger and black pepper are powdered and made into pills with butter oil, 2–3 of these pills (pea size) are administered daily[75]
West Bengal stateBlood dysentery, piles, leprosy, headacheBark[87]
Diabetes, intestinal worms; roots to stop bleeding from noseSeeds
DropsyThe dried bark is rubbed over the body
Madhya Pradesh state, tribal communities of chitrakoot regionArthritis and diarrhea in cattleLeaf decoction twice a day[88,89]
Odisha stateRheumatismRoot bark[90]
Tribals of Bargarh districtRheumatism10 g of root bark is boiled in water (400 mL) and the prepared decoction (100 mL) is taken 1–2 times daily on empty stomach
Sundargarh districtBoils, cut, abscess and woundsRoot paste[91]
Bondo tribe of Malkangiri districtRheumatic painTwo to three leaves are attached with the latex of the same plant and fomented externally over backbone[92]
DysenteryRoot powder
Tribals of SimilipalMalaria and dysenteryStem bark[93]
DysenteryStem bark infusion with honey in a ratio of 3:1 is taken once a day on empty stomach[94]
DysenteryFrom bark of H. pubescens, Terminalia arjuna and Pterocarpus marsupium (in equal ratio) pill is prepared. One pill is taken orally on empty stomach for three days[95]
Tribes of Mayurbhanj districtStomach pain and blood dysentery [94]
HeadacheDecoction of roots with garlic and mustard is made into paste and applied externally as an ointment[95]
Skin infection, jaundiceLeaf paste[96]
Bhadrak districtDeep cutsBark and latex[52]
Kalahandi districtDysenteryStem bark of Careya arborea and H. pubescens with water[33]
Andhra Pradesh state, visakhapatnam districtNerve disorderSpoonful of shade-dried stem bark powder was taken orally with glass of water daily[97]
Khammam districtPost-partum problems15 g of root is ground with 20 mL country liquor of rice. Five spoons of this were taken immediately after delivery followed by 2 g of Ferula asafoetida rhizome powder[98]
Visakhapatnam districtFeverDecoction prepared by adding 100–400 mL water with leaves of H. pubescens and root of Andographis paniculata, given twice a day[99]
Karnataka state, Hosanagara taluk of Shimoga districtCancerOne handful of roots ground in cow’s buttermilk and given orally, twice daily for one month[100]
StomachacheRoots crushed in water and juice is taken orally, twice daily for 1–2 days[101]
Tribes of the Shimoga districtRingworm and poor milk productionBark[102]
Uttara kannadaUlcer in intestineUsed a mixture of plants viz. Syzygium cumini (bark); Holarrhena pubescens (bark), Madhuca indica (leaves and bark), Careya arborea (bark), Elaegnus conferta (bark), Myristica fragrans (fruit), Syzygium aromaticum (flower bud), Piper nigrum (fruit), Trachyspermum ammi (fruit), Zingiber officinale (rhizome), Cuminum cyminum (fruit) in decoction form[103]
Uttarakhand state, Tharu community of district Udham Singh nagarChronic dysenteryPaste made with flower and cow’s milk taken orally, for 4 days[104,105,106]
Theni district (Western ghats)DysenteryDecoction made from the root bark is taken orally twice a day for two days[107]
Gujrat, Rajstan and Kerala stateDropsy and swellingBark extracts from Bombax ceiba, Hymenodictyon excelsium, Azadirachta indica and H. pubescens made by crushing is given with water in morning and evening for 5 days[56]
SnakebiteThe crushed root is given with ghee
Table 4. In vivo studies with H. pubescens.
Table 4. In vivo studies with H. pubescens.
Biological ActivityPartsExtract/CompoundEffective Concentration/DoseStudy ModelReferences
AntihyperglycemicSeedsAqueous and petroleum ether extract250 mg/kg BWRats[109]
SeedsMethanol extract300 mg/kg BW in ratsRats[110]
SeedsEthanolic extract300 mg/kg and 600 mg/kgRats[110]
LeavesEthanolic extract400 mg/kg BWRats[112]
Anti-diarrhealSeedsEthanolic extract200 and 400 mg/kgRats[114]
SeedsAlkaloids200–800 mg/kgRats[115]
Anti-inflammatoryNot statedNot stated400 mg/kgRats[116]
DiureticSeedsAqueous30–100 mg/kgRats[117]

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Zahara, K.; Panda, S.K.; Swain, S.S.; Luyten, W. Metabolic Diversity and Therapeutic Potential of Holarrhena pubescens: An Important Ethnomedicinal Plant. Biomolecules 2020, 10, 1341.

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Zahara K, Panda SK, Swain SS, Luyten W. Metabolic Diversity and Therapeutic Potential of Holarrhena pubescens: An Important Ethnomedicinal Plant. Biomolecules. 2020; 10(9):1341.

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Zahara, Kulsoom, Sujogya Kumar Panda, Shasank Sekhar Swain, and Walter Luyten. 2020. "Metabolic Diversity and Therapeutic Potential of Holarrhena pubescens: An Important Ethnomedicinal Plant" Biomolecules 10, no. 9: 1341.

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