Antileishmanial Activities of Medicinal Herbs and Phytochemicals In Vitro and In Vivo: An Update for the Years 2015 to 2021

Abstract

Leishmaniasis is one of the most neglected tropical diseases that present areal public health problems worldwide. Chemotherapy has several limitations such as toxic side effects, high costs, frequent relapses, the development of resistance, and the requirement for long-term treatment. Effective vaccines or drugs to prevent or cure the disease are not available yet. Therefore, it is important to dissect antileishmanial molecules that present selective efficacy and tolerable safety. Several studies revealed the antileishmanial activity of medicinal plants. Several organic extracts/essential oils and isolated natural compounds have been tested for their antileishmanial activities. Therefore, the aim of this review is to update and summarize the investigations that have been undertaken on the antileishmanial activity of medicinal plants and natural compounds derived, rom plants from January 2015 to December 2021. In this review, 94 plant species distributed in 39 families have been identified with antileishmanial activities. The leaves were the most commonly used plant part (49.5%) followed by stem bark, root, and whole plant (21.9%, 6.6%, and 5.4%, respectively). Other plant parts contributed less (<5%). The activity was reported against amastigotes and/or promastigotes of different species (L. infantum, L. tropica, L. major, L. amazonensis, L. aethiopica, L. donovani, L. braziliensis, L. panamensis, L. guyanensis, and L. mexicana). Most studies (84.2%) were carried out in vitro, and the others (15.8%) were performed in vivo. The IC₅₀ values of 103 plant extracts determined in vitro were in a range of 0.88 µg/mL (polar fraction of dichloromethane extract of Boswellia serrata) to 98 µg/mL (petroleum ether extract of Murraya koenigii). Among the 15 plant extracts studied in vivo, the hydroalcoholic leaf extract of Solanum havanense reduced parasites by 93.6% in cutaneous leishmaniasis. Voacamine extracted from Tabernaemontana divaricata reduced hepatic parasitism by ≈30 times and splenic parasitism by ≈15 times in visceral leishmaniasis. Regarding cytotoxicity, 32.4% of the tested plant extracts against various Leishmania species have a selectivity index higher than 10. For isolated compounds, 49 natural compounds have been reported with anti-Leishmania activities against amastigotes and/or promastigotes of different species (L. infantum, L. major, L. amazonensis, L. donovani and L. braziliensis). The IC₅₀ values were in a range of 0.2 µg/mL (colchicoside against promastigotes of L. major) to 42.4 µg/mL (dehydrodieuginol against promastigotes of L. amazonensis). In conclusion, there are numerous medicinal plants and natural compounds with strong effects (IC₅₀ < 100 µg/mL) against different Leishmania species under in vitro and in vivo conditions with good selectivity indices (SI > 10). These plants and compounds may be promising sources for the development of new drugs against leishmaniasis and should be investigated in randomized clinical trials.

1. Introduction

Leishmaniasis is a group of diseases caused by protozoa parasites from more than 20 Leishmania species. In 2018, 92 countries and 83 territories were considered endemic for Leishmania species or had previously reported cases of cutaneous and visceral leishmania, respectively. Today, more than 1 billion people live in areas endemic to leishmaniasis and are at risk of infection. An estimated 30,000 new cases of visceral leishmania and more than 1 million new cases of cutaneous leishmania occur annually [1]. The parasite is categorized into two main groups: Old World leishmaniasis, which is endemic in Africa, Asia, the Mediterranean, and the Middle East. Leishmania tropica, L. major, L. aethiopica, and L. donovani are the four common species causing Old World leishmaniasis. New World leishmaniasis is caused by L. mexicana, L. amazonensis, L. braziliensis, L. panamensis, L. peruviana, L. guyanensis, L. pifanoi, L. venezuelensis, L. shawi, and L. lainsoni [2]. There are three clinical forms of leishmaniasis in humans: namely, cutaneous, mucocutaneous and visceral leishmaniasis. Cutaneous leishmaniasis is a less severe form of the disease which usually manifests in self-healing ulcers. Mucocutaneous leishmaniasis results in disfiguring lesions of mucous membranes in the nose, mouth, and throat. Visceral leishmaniasis is the most severe form of the disease which can result in 95% mortality of infected patients if not treated [3].

In 2020, more than 90% of new cases of visceral leishmaniasis reported to the WHO occurred in Bangladesh, Brazil, China, Ethiopia, Eritrea, India, Kenya, Somalia, South Sudan, Sudan, and Yemen [1]. Over 90% of mucocutaneous leishmaniasis occurred in Bolivia, Brazil, Ethiopia, and Peru, and more than 85% of cutaneous leishmaniasis cases appeared in Afghanistan, Algeria, Brazil, Colombia, Iran, Libya, Pakistan, Peru, Syria, and Tunisia [1]. Depending on the stage of its life cycle, the parasite exhibits two morphological forms in its life cycle: The amastigotes in macrophages of the mammalian host and the promastigotes in the gut of the sand fly vectors. The life cycle of the Leishmania parasite starts if a parasitized female sand fly takes a blood meal from a vertebrate host to produce its eggs. As the sand fly feeds, infective promastigotes enter the vertebrate host via the insect’s proboscis. The promastigotes are then phagocytosed by macrophages which they transform into amastigotes and reproduce by binary fission. They increase in number until the cell eventually bursts and then infects other phagocytic cells to continue the cycle [4]. Over the years, a number of drugs have been employed for the treatment of leishmaniasis. A brief account of the mechanism of action and mode of administration of these drugs has been presented in

Table 1. Drugs used for the treatment of leishmaniasis.

Name of the Drug	Mode of Action	Mode of Administration	Adverse Effects
Pentavalent antimonials	Inhibition of glycolysis and β-oxidation of fatty acids of parasite	Intralesional for CL, Parenteral	Abdominal pain, erythema, nausea, toxicity (hepatic, pancreas, renal, muscular, and skeletal cardiothrombocytopenia or leukopenia)
Amphotericin B	Binding to parasite’s membrane sterols and changing its permeability selective to K+ and Mg2+	Liposomal formulations, Deoxycholate formulations	Fever, nausea, hypokalemia, anorexia, leukopenia, kidney failure, and heart problems
Pentamidine	Interferes with DNA synthesis and modifies the morphology of kinetoplast	Parenteral, Intramuscular administration	Pain, nausea, vomiting, dizziness, myalgia, hypertension, headache, hypoglycemia, and transient hyperglycemia
Miltefosine	Associated with phospholipid biosynthesis and alkyl-lipid metabolism in leishmania	Oral for VL	Nausea, vomiting, diarrhea, and raised creatinine
Paromomycin	Inhibition of protein biosynthesis in sensitive organism	Topical for CL Parenteral for VL	Erythema, pain, edema, and ototoxicity (damage to the internal ear)

[5].

Latest developments in the prevention and treatment regarding a permanent solution for leishmaniasis in terms of successful human vaccination is still a major challenge. However, there are different vaccinations currently being tested in mouse models. One of them uses “killed but metabolically active” parasites to induce host immune system reaction. Using salivary peptides of the sandfly holds the potential to be used as a vaccine component. However, the complex immune response makes it a challenge [6]. Macrophage-targeted drug delivery systems are another novel approach to directly affect Leishmania parasites that live in the macrophages. As getting into macrophages is a challenge, liposomes, microspheres, nanoparticles, and carbon nanotubes are some of the various drug carriers that are studied to target macrophages. In addition, the use of specific receptors expressed by macrophages to actively deliver a drug is also used [7].

The current treatment by chemical drugs has several limitations such as toxic side effects, high costs, frequent relapses, the development of resistance, and the requirement for long-term treatment [8,9]. Thus, investments in novel drug development against this parasitic disease may be a risky affair. Medicinal plants are centuries-old sources in the various traditional herbal medicine systems of the world. For instance, their importance lies in the fact that the WHO concludes that about 80% of the world’s population relies on them for primary health care [10]. Moreover, 25 to 50% of the pharmacopeias worldwide contain plant products and drugs derived from natural products [11]. Therefore, current research approaches for the treatment of leishmaniasis should largely consider medicinal plants as an important area of search.

The aim of this review is to update and summarize the investigations that have been undertaken on the antileishmanial activity of medicinal plants and natural compounds derived from plants from January 2015 to December 2021.

2. Results

As shown in

Table 2. Botanical characteristics of the medicinal herbs in the present study.

No	Family Name	Scientific Name	Part Used
1.	Anacardiaceae	Pistacia lentiscus	Leaves
		Schinus terebinthifolia	Fruits
		Schinus molle	Leaves
		Spondias mombin	Leaves
2.	Annonaceae	Annona senegalensis	Stem bark
		Bocageopsis multiflora	Leaves
		Guatteria latifolia	Branch
		Cleistopholis patens	Stem bark
3.	Apiaceae	Ferula communis	Whole plant
4.	Apocynaceae	Tabernaemontana divaricata	Voacamine
		Mondia whitei	Roots
		Pentalinon andrieuxii	Pentalinon sterol
5.	Araliaceae	Oreopanax floribundus	Leaves
6.	Arecaceae	Phoenix dactylifera	Kernel and date fruit
7.	Asteraceae	Acanthospermum hispidum	Whole plant
		Tessaria integrifolia	Leaves
		Abuta grandifolia	Leaves
		Cynara scolymus	Leaves
		Artemisia absinthium	Leaves
		Artemisia campestris	Leaves
		Artemisia herba-alba	Aerial parts, Leaves
		Bidens pilosa	Whole plant
		Tessaria integrifolia	Whole plant
8.	Balanophoracea	Thonningia sanguinea	Whole plant
9.	Bignoniaceae	Handroanthus serratifolius	Lapachol
		Jacaranda glabra	Bark
10.	Burseraceae	Boswellia serrata	Resin
11.	Cannabaceae	Celtis australis	Leaves
12.	Capparaceae	Capparis spinosa	Fruits
13.	Cistaceae	Citrus sinensis	Leaves
14.	Combretaceae	Terminalia ivorensis	Leaves
15.	Cupressaceae	Juniperus excelsa	Leaves, fruits
16.	Ericaceae	Arbutus unedo	Leaves
		Erica arborea	Flower
17.	Euphorbiaceae	Bridelia ferruginea	Leaves
		Ejije bidu	Leaves
		Croton caudatus	Leaves
18.	Fabaceae	Afzelia africana	Stem bark
		Baphia nitida	Stem bark
		Cassia alata	Leaves
		Cassia gloca	Leaves
		Cassia sieberiana	Roots, leaves
		Prosopis laevigata	Leaves
		Parkia clappertoniana	Stem bark, leaves
		Tamarindus indica	Leaves
		Prosopis juliflora	Leaves
19.	Gentianaceae	Anthocleista nobilis	Leaves, stem bark, root
		Centaurium erythraea	Flowering, stems
20.	Lamiaceae	Marrubium vulgare	Leaves
		Mentha pulegium	Leaves
		Otostegia integrifolia	Whole plant
		Rosmarinus officinalis	Leaves
		Salvia clandestina	Aerial parts
		Vitex fosteri	Stem bark, leaves
21.	Lauraceae	Aniba riparia	Fruits
		Persea ferruginea	Leaves
		Cinnamomum cassia	Bark
22.	Loranthaceae	Loranthus europaeus	Aerial part
23.	Malvaceae	Ceiba pentandra	Stem bark
		Cola acuminata	Stem bark
		Cola cordifolia	Stem bark, leaves
		Glyphaea brevis	Leaves
24.	Marantaceae	Thalia geniculata	Roots
		Iresine diffusa	Flower
25.	Meliaceae	Khaya grandifoliola	Stem bark
		Cedrela spp	Bark
		Azadirachta indica	Leaves
26.	Moraceae	Treculia africana	Stem bark
		Ficus capensis	Stem bark, leaves
27.	Myrtaceae	Eugenia uniflora	Leaves, seed
28.	Ochnaceae	Lophira lanceolata	Stem bark, roots
29.	Olacaceae	Ximenia americana	Stem and twigs
30.	Papaveraceae	Argemone mexicana	Aerial parts
31.	Piperaceae	Piper pseudoarboreum	Leaves
32.	Rhamnaceae	Ziziphus spina-christi	Whole plant
33.	Rosaceae	Pyrus communis	Leaves
		Pyrus pashia	Leaves
		Prunus armeniaca	Leaves
		Eryobotrya japonica	Leaves
34.	Rubiaceae	Mitragyna inermis	Stem bark, leaves
		Psychotria buhitenii	Leaves
35.	Rutaceae	Zanthoxylum zanthoxyloides	Roots, stem bark
		Murraya koenigii	Stem bark
		Clausena anisata	Roots
36.	Scrophulariaceae	Scoparia dulcis	Aerial part
		Licania salicifolia	Leaves
37.	Solanaceae	Solanum havanense	Leaves
		Solanum lycocarpum	Leaves
		Solanum myriacanthum	Leaves
		Solanum nudum	Leaves
		Physalis angulata	Flowers
		Solanum seaforthianum	Leaves
38.	Urticaceae	Urtica dioica	Leaves
39.	Verbenaceae	Lantana camara	Leaves

, 92 plant species distributed in 39 families have been identified with anti-Leishmania activities. The family Fabaceae accounted for the highest percentage (9.7%) followed by Asteraceae (7.6%). Lamiaceae and Solanaceae account for 6.5% each.

The leaves were the most commonly used plant part as compared to other parts (49.5%) followed by stem bark, roots, and whole plant (21.9%, 6.6%, and 5.4%, respectively). Aerial parts and fruits accounted for 4.5% each. Other plant parts (flowers, seeds, resins, branches, and kernels) contributed less (<4%) (Figure 1).

With respect to the test methods, 84.2% of studies were carried in vitro, while 15.8% of them were performed using in vivo assays (

Table 3. Anti-Leishmania activity of medicinal plants in vitro.

No.	Scientific Name	Organism	Stage	Part Used	Most Active Extract/ Essential Oil	IC50 (µg/mL)	Bioactive Compounds	Data Analysis (Activity)	Reference
1.	Abuta grandifolia	L. amazonensis	Promastigotes	Leaves	Ethanol	38.1	Alkaloids, triterpenes, saponins	Moderate	[12]
		L. braziliensis				31.1		Moderate
2.	Acanthospermum hispidum	L. donovani	Promastigotes	Whole plant	50% aqueous ethanol	32.10	Essential oil, alkaloids	Moderate	[13]
3.	Afzelia africana	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	77.10	Alkaloids, tannins, flavonoids, saponins	Weak	[13]
4.	Aniba riparia	L. amazonensis	Amastigotes	Fruits	50% aqueous ethanol	1.30	Riparin E	High	[14]
			Promastigotes			4.70		High
5.	Annona senegalensis	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	10.80	Alkaloids, tannins, flavonoids, saponins, terpenoids, glycosides	Moderate	[13]
				Stem bark		27.80		Moderate
6.	Anthocleista nobilis	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	41.50	Glycosides, saponins, steroids	Moderate	[13]
				Root		79.0	Anthocleistol	Weak
7.	Arbutus unedo	L. infantum	Promastigotes	Leaves	n-Hexane	64.05	Phenolics, flavonoids	Weak	[15]
		L. tropica				79.57		Weak
8.	Argemone mexicana	L. donovani	Promastigotes	Aerial part	Petroleum ether	50.0	-	Moderate	[16]
9.	Artemisia absinthium	L. major	Promastigotes	Leaves	Hydrodistillation	1.49	Essential oil	High	[17]
10.	Artemisia campestris	L. major	Promastigotes	Leaves	Hydrodistillation	2.20	Essential oil	High	[17]
11.	Artemisia herba-alba	L. major	Promastigotes	Leaves	Hydrodistillation	1.20	Essential oil	High	[17]
12.	Artemisia herba-alba	L. infantum.	Amastigote	Aerial part	Methanol extract	68.25	-	Weak	[18]
		L. major				37.87		Moderate
		L. infantum	Promastigotes			77.97		Weak
		L. major				55.21		Weak
13.	Azadirachta indica	L. infantum	Amastigotes	Leaves	Oil	15.3	Phenolics, flavonoids	Moderate	[19]
		L. tropica				17.6		Moderate
14.	Baphia nitida	L. donovani	Promastigotes	Stem-bark	50% aqueous ethanol	34.40	Tannins, flavonoids, saponins, glycosides	Moderate	[13]
15.	Bidens pilosa	L. donovani	Promastigotes	Whole plant	50% aqueous ethanol	28.90	Essential oil, flavonoids, alkaloids, saponins, triterpenes	Moderate	[13]
16.	Bocageopsis multiflora	L. amazonensis	Promastigotes	Leaves	Ethanol	37.9	Essential oil, alkaloids	Moderate	[12]
		L. braziliensis				19.1		Moderate
17.	Boswellia serrata	L. donovani	Amastigotes	Resin	Polar fractions of dichloromethane	0.88	Boswellic acids	High	[20]
18.	Bridelia ferruginea	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	16.50	Flavonoids, tannins, triterpenoids	Moderate	[13]
19.	Capparis spinosa	L. tropica	Promastigotes	Fruits	Methanol	44.6	Tannins, alkaloids, saponins, terpenoids, glycosides	Moderate	[21]
					Aqueous	28.5		Moderate
20.	Cassia alata	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	10.10	Flavonoids, glycosides	Moderate	[22]
21.	Cassia gloca	L. tropica	Promastigotes	Leaves	Methanol	9.62	Flavonoids	High	[22]
22.	Cassia sieberiana	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	62.90	Flavonoids, alkaloids	Weak	[23]
23.	Cedrela spp.	L. amazonensis	Promastigotes	Bark	Ethanol	36.8	Sesquiterpenes, triterpenes	Moderate	[22]
		L. braziliensis				18.2		Moderate
24.	Ceiba pentandra	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	31.10	Isoflavones, sesquiterpenoids	Moderate	[13]
25.	Centaurium erythraea	L. tropica	Promastigotes	Flowering stems	n-Hexane	37.20	Phenolics, flavonoids	Moderate	[23]
		L. major				64.52		Weak
26.	Celtis australis	L. tropica	Promastigotes	Leaves	Methanol	69.13	Flavonoids	Weak	[22]
27.	Cistus crispus	L. major	Promastigotes	Leaves	Methanol	84.29	Phenolics, flavonoids	Weak	[15]
		L. infantum			n-Hexane	82.39		Weak
		L. tropica				96.82		Weak
		L. major				47.29		Moderate
28.	Citrus sinensis	L. tropica	Promastigotes	Leaves	Methanol	12.27	Flavonoids	Moderate	[22]
29.	Cola acuminata	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	47.80	Purine alkaloids, catechins, (tannins)	Moderate	[13]
30.	Cola cordifolia	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	25.10	Tannins, phenolics	Moderate	[13]
				Leaves		18.20		Moderate
31.	Clausena anisata	L. donovani	Promastigotes	Roots	50% aqueous ethanol	12.10	Essential oil, indole alkaloids, coumarins	Moderate	[13]
32.	Cleistopholis patens	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	60.20	Flavonoids, saponins, alkaloids	Weak	[13]
33.	Croton caudatus	L. donovani	Promastigotes	Leaves	Ethyl acetate –hexane (9:1)	10.0	Terpenoids	High	[23]
			Amastigote			2.5		High
34.	Cynara scolymus	L. tropica	Promastigotes	Stem leaf	Ethanol	80.0	-	Weak	[24]
35.	Ejije bidu	L. amazonensis	Promastigotes	Leaves	Ethanol	17.8	-	Moderate	[12]
		L. braziliensis				13.3		Moderate
36.	Erica arborea	L. major	Promastigotes	Flower	Methanol	43.98	-	Moderate	[18]
		L. infantum.				61.27		Weak
		L. major	Amastigotes			36		Moderate
		L. infantum.				53.93		Weak
37.	Eryobotrya japonica	L. tropica	Promastigotes	Leaves	Methanol	10.59	Flavonoids	Moderate	[22]
38.	Eugenia uniflora	L. amazonensis	Amastigotes	Leaves	n-Hexane	9.20	Sesquiterpenes, flavonoids	High	[25]
		L. donovani	Promastigotes	Seeds	50% aqueous ethanol	26.60	Essential oil, flavonoids, tannins	Moderate	[13]
39.	Ferula communis	L. aethiopica	Promastigotes	Whole parts	80% methanol	11.38	Phenolics, flavonoids	Moderate	[26]
		L. donovani				23.41		Moderate
		L. aethiopica	Amastigotes			14.32		Moderate
		L. donovani				31.12		Moderate
40.	Ficus capensis	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	37.0	Alkaloids, phenolics, flavonoids	Moderate	[13]
				Leaves		88.90		Weak
41.	Glyphaea brevis	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	43.40	Tannins, alkaloids, flavonoids	Moderate	[13]
42.	Guatteria Latifolia	L. amazonensis	Promastigote	Branch	n-hexane fraction of ethanol	51.7	Alkaloids	Weak	[27]
43.	Iresine diffusa	L. amazonensis	Promastigotes	Flower	Ethanol	30.5	Sesquiterpenes, triterpenes	Moderate	[12]
		L. braziliensis				11.1		Moderate
44.	Jacaranda Glabra	L. amazonensis	Promastigotes	Bark	Ethanol	29.8	-	Moderate	[12]
		L. braziliensis				17.4		Moderate
45.	Khaya grandifolia	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	43.20	Alkaloids, saponins, tannins	Moderate	[13]
46.	Lantana camara	L. amazonensis	Amastigotes	Leaves	Dichloromethane	21.8	Terpenoids	Moderate	[28]
47.	Licania Salicifolia	L. panamensis	Amastigotes	Leaves	Ethyl acetate	9.8	Triterpenes, flavonoids	High	[29]
48.	Lophira lanceolata	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	68.60	Flavonoids, saponins, alkaloids	Weak	[13]
				Roots		66.0	Alkaloids	Weak
49.	Marrubium vulgare	L. infantum	Amastigotes	Leaves	Methanol	18.64	-	Moderate	[18]
		L. major				32.15		Moderate
		L. infantum	Promastigotes			35.63		Moderate
		L. major				45.84		Moderate
50.	Mentha pulegium	L. infantum	Promastigotes	Leaves	Essential oil	2.0	Menthone, pulegone	High	[30]
		L. tropica				2.2		High
		L. major				1.30		High
51.	Mitragyna Inermis	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	21.90	Indole alkaloids, triterpenoids	Moderate	[13]
				Stem bark		28.0		Moderate
52.	Mondia whitei	L. donovani	Promastigotes	Roots	50% aqueous ethanol	31.0	Glycosides	Moderate	[13]
53.	Murraya koenigii	L. donovani	Promastigotes	Stem	Petroleum ether	98.0	-	Weak	[16]
54.	Oreopanax floribundus	L. panamensis	Amastigotes	Leaves	Dichloromethane	24.6	Triterpenes	Moderate	[29]
					Ethyl acetate	23.7	Triterpenes, flavonoids	Moderate
55.	Otostegia integrifolia	L. aethiopica	Promastigotes Amastigotes	Whole parts	80% methanol	13.03	Phenolics, flavonoids	Moderate	[31]
		L. donovani				17.24		Moderate
		L. aethiopica				16.84		Moderate
		L. donovani				14.55		Moderate
56.	Parkia clappertoniana	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	17.0	Saponins, flavonoids, Tannins	Moderate	[13]
				Stem bark		17.60	Saponins, steroids, triterpenes	Moderate
57.	Persea ferruginea	L. panamensis	Amastigotes	Leaves	Ethyl acetate	25.5	Triterpenes, leucoanthocyanidins, coumarins	Moderate	[29]
58.	Phoenix dactylifera	L. major	Promastigotes	kernel	Methanol	23.0	Gallic acid	Moderate	[32]
59.	Physalis angulata	L. amazonensis	Promastigotes	Flower	Ethanol	17.6	Terpenes, phenolic acids, flavonoids	Moderate	[12]
		L. braziliensis				43.5		Moderate
60.	Piper pseudoarboreum	L. amazonensis	Promastigotes	Leaves	Ethanol	31.4	Alkamides	Moderate	[33]
		L. braziliensis				21.3		Moderate
		L. guyanesis				41.3		Moderate
		L. infantum				32.3		Moderate
61.	Pistacia lentiscus	L. infantum	Promastigotes	Leaves	Essential oil	11.28	Myrcene, α-pinene	Moderate	[23]
		L. tropica				23.50		Moderate
		L. major				17.52		Moderate
		L. infantum		Fruits	Essential oil	8.0	Limonene α-pinene	High
		L. tropica				26.20		Moderate
		L. major				21.42		Moderate
62.	Rosmarinus officinalis	L. infantum	Promastigotes	Leaves	Essential oil	1.20	α-Pinene, 1,8-cineole, borneol	High	[23]
		L. tropica				3.50		High
		L. major				2.60		High
63.	Prosopis juliflora	L. donovani	Promastigotes	Leaves	Methanol	3.12	Saponins, tannins, flavonoids, alkaloids	High	[34]
64.	Prosopis laevigata	L. amazonensis	Amastigotes	Leaves	Aqueous	35.2	Alkaloids, anthraquinones	Moderate	[28]
65.	Prunus armeniaca	L. tropica	Promastigotes	Leaves	Ethanol	16.18	Alkaloids, phenolics, tannins, flavonoids, terpenoids, coumarins	Moderate	[35]
66.	Psychotria buhitenii	L. panamensis	Amastigotes	Leaves	Dichloromethane	21.5	Triterpenes, flavonoids	Moderate	[29]
					Ethyl acetate	14.1	Triterpenes, saponins, Coumarins, anthocyanins	Moderate
					Ethanol	29.4	Saponins, phenolics, tannins, coumarins, anthocyanins	Moderate
67.	Pyrus communis	L. tropica	Promastigotes	Leaves	Ethanol	56.68	Alkaloids, phenolics, tannins, flavonoids, terpenoids, quinones, saponins	Weak	[35]
68.	Pyrus pashia	L. tropica	Promastigotes	Leaves	Ethanol	60.95	Alkaloids, phenolics, tannins, flavonoids, terpenoids, quinones, saponins	Weak	[35]
69.	Salvia clandestina	L. infantum	Promastigotes	Aerial part	n-Hexane	14.11	-	Moderate	[36]
		L. infantum			Dichloromethane	31.57		Moderate
		L. tropica				33.77		Moderate
		L. major				24.56		Moderate
70.	Schinus molle	L. amazonensis	Amastigotes	Leaves	Dichloromethane	25.9	Terpenoids	Moderate	[28]
					Dichloromethane: Methanol (1:1)	21.8	Terpenoids, phenolics	Moderate
71.	Schinus terebinthifolia	L. amazonensis	Promastigotes	Fruits	n-Hexane	13.90	Triterpenes	Moderate	[29]
72.	Scoparia dulcis	L. amazonensis	Promastigotes	Aerial part	Ethanol	23.9	Diterpenes, triterpenes, flavonoids	Moderate	[12]
		L. braziliensis				25.1		Moderate
73.	Spondias mombin	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	81.50	-	Weak	[13]
74.	Tamarindus indica	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	58.12	Phenolics, flavonoids	Weak	[13]
75.	Terminalia ivorensis	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	24.90	Terminolic acid, quercetin, β-glycyrrhetinic acid	Moderate	[13]
76.	Tessaria integrifolia	L. amazonensis	Promastigotes	Leaves	Ethanol	54.20	Sesquiterpenes, flavonoids	Weak	[12]
		L. braziliensis				31.60		Moderate
77.	Thalia geniculata	L. amazonensis	Promastigotes	Roots	Ethanol	29.8	Phytosterols	Moderate	[12]
		L. braziliensis				17.4		Moderate
78.	Thonningia sanguinea	L. donovani	Promastigotes	Whole plant	50% aqueous ethanol	18.60	Alkaloids, tannins, flavonoids	Moderate	[13]
79.	Treculia africana	L. donovani	Promastigotes	Stem bark	50% aqueous ethanol	44.80	Catechin, cyanidin glycosides	Moderate	[13]
80.	Vitex fosteri	L. donovani	Promastigotes	Leaves	50% aqueous ethanol	72.40	Essential oil, flavonoids	Weak	[13]
				Stem bark		49.80		Moderate
81.	Ximenia americana	L. donovani	Promastigotes	Stem and twigs	50% aqueous ethanol	36.10	Tannins, flavonoids, alkaloids	Moderate	[13]
82.	Zanthoxylum zanthoxyloides	L. donovani	Promastigotes	Roots	50% aqueous ethanol	13.50	Alkaloids, tannins, flavonoids, essential oil	Moderate	[13]
				Stem bark		45.20		Moderate
83.	Ziziphus spina-christi	L. major	Amastigotes	Leaves	Methanol	54.6	Tannins, flavonoids, Glycosides, alkaloids, terpenoids	Moderate	[37]

and

Table 4. Anti-Leishmania activity of medicinal plants in vivo.

No.	Plant Species	Leishmania Species	Route, Dose, and Scheme of Treatment	Efficacy	Bioactive Compounds	Reference
1.	Cinnamomum cassia	Visceral leishmaniasis (L. donovani)	Oral: 100 mg/kg/d for 10 days	Reduction of hepatic parasitism by 80.9% and splenic parasitism by 82.9%	Cinnamaldehyde and its derivatives	[38]
2.	Croton caudatus	Visceral leishmaniasis (L. donovani)	Oral: 5 mg/kg/d five consecutive days	Reduction of hepatic parasitism by 65% and splenic parasitism by 69.1%	Terpenoids	[23]
3.	Handroanthus serratifolius	Cutaneous leishmaniasis (L. amazonensis)	Oral: 25 mg/kg/d for 10 days	24.5-fold reduction of parasite number	Lapachol	[39]
		Visceral leishmaniasis (L. infantum)		Reduction parasite number in spleen (4.6-fold) and liver (5.3-fold)
4.	Loranthus europaeus	Cutaneous leishmaniasis (unspecific)	Topical: ointment (40%) once daily at bedtime for 6 h under occlusion for maximal 6 weeks	79.0% cure rate without side effects	Flavonoids, alkaloids, glycosides, triterpenes, phenolic acids	[40]
5.	Pentalinon andrieuxii	Visceral leishmaniasis (L. donovani)	2.5 mg/kg i.v.	Reduction of 64, 83, and 57% of parasites in the liver, spleen, and bone marrow.	Pentalinonsterol	[41]
6.	Piper pseudoarboreum	Cutaneous leishmaniasis (L. amazonensis)	Intralesional: 25 mg/kg/d for 4 days	Reduction of skin lesions by 40% and visceralization by 55%.	(E)-Piplartine	[33]
7.	Prosopis juliflora	Visceral leishmaniasis (L. donovani)	Oral: 100 mg/kg/d for 21 days	85.1% reduction of parasite number in spleen	Saponins, tannins, flavonoids, alkaloids	[34]
8.	Solanum havanense	Cutaneous leishmaniasis (L. amazonensis)	Intralesional: 30 mg/kg every 4 days, 5 doses	93.6% reduction of parasite number	Steroidal alkaloids, saponins, phenolics, triterpenes, coumarins	[42]
9.	Solanum lycocarpum	Cutaneous leishmaniasis (L. mexicana)	Topical: 10 μg/d for 6 weeks	71.4% reduction of parasite number	Alkaloids (solamargine, solasonine)	[43]
10.	Solanum myriacanthum	Cutaneous leishmaniasis (L. amazonensis).	Intralesional: 30 mg/kg every 4 days, 5 doses	56.8% reduction of parasite number	Steroidal alkaloids, saponins, phenolics, triterpenes, coumarins	[42]
11.	Solanum nudum	Cutaneous leishmaniasis (L. amazonensis)	Intralesional: 30 mg/kg every 4 days, 5 doses	80% reduction of parasite number	Steroidal alkaloids, saponins, phenolics, triterpenes, coumarins	[42]
12.	Solanum seaforthianum	Cutaneous leishmaniasis (L. amazonensis)	Intralesional: 30 mg/kg every 4 days, 5 doses	49.9% reduction of parasites in treated animals	Steroidal alkaloids, saponins, phenolics, triterpenes, coumarins	[42]
13.	Tabernaemontana divaricata	Visceral leishmaniasis (L. donovani)	Intraperitoneal: 5 mg/kg twice a week for 3 weeks	Decreased the hepatic parasitism by ≈30 times and splenic parasitism by ≈15 times	Voacamine	[35]
14.	Urtica dioica	Cutaneous leishmaniasis (L. major)	Intramuscular and intralesional: 250 mg/kg for 10 weeks	Intralesional treatment reduced lesions more than amphotericin B (control)	-	[44]
15.	Ziziphus spina-christi	Cutaneous leishmaniasis (L. major)	Topical: 100 and 200 mg/kg/d for 4 weeks	Reduction of lesion size by 6.4- and 8.6-fold	Tannins, flavonoids, glycosides, alkaloids, terpenoids	[37]

). For in vitro assay, 80 medicinal plants were screened in vitro for antileishmanial activities against different Leishmania species (L. infantum, L. tropica, L. major, L. amazonensis, L. aethiopica, L. donovani, L. braziliensis, L. panamensis, L. guyanensis, and L. mexicana) and life cycle forms (amastigotes and/or promastigotes). The IC₅₀ value of 103 plant extracts/essential oils determined in vitro was in a range of 0.88 µg/mL (polar fraction of dichloromethane extract of Boswellia serrata) to 98 µg/mL (petroleum ether extract of Murraya koenigii) (Table 3). B. serrata (resins), R. officinalis (leaves), A. riparia (fruits), M. pulegium (leaves) as extracts had strong anti-Leishmania activity (0.88, 1.2, 1.3, and 1.3 µg/mL, respectively).

For in vivo assay, among the 15 medicinal plants studied in vivo, the highest activity against cutaneous leishmaniasis was exhibited by the hydroalcoholic leaf extract of Solanum havanense, which reduced parasites by 93.6%, and the highest activity against visceral leishmaniasis was shown by the voacamine compound extracted from Tabernaemontana divaricata, which reduced the hepatic and splenic parasitism by ≈30 times and ≈15 times, respectively (Table 4). For cytotoxic activity, 32.4% of tested plant extracts have good cytotoxic activity with a selectivity index of SI > 10. (

Table 5. Cytotoxic activity and selectivity index of medicinal plants in the present study (p = promastigote; a = amastigote).

No.	Plant Species	Leishmania Species	Part Used	Bioactive Extract/ Compounds	Cytotoxicity (CC50 µg/mL)	Selectivity Index (CC50/IC50)	Reference
1.	Abuta grandifolia	L. amazonensis p	Leaves	Ethanol	15.2	0.4	[12]
		L. braziliensis p			15.6	0.5
2.	Acanthospermum hispidum	L. donovani p	Whole plant	50% aqueous ethanol	55.5	1.73	[13]
3.	Afzelia africana	L. donovani p	Stem bark	50% aqueous ethanol	232.8	3.02	[13]
4.	Aniba riparia	L. amazonensis a	Fruits	50% aqueous ethanol	50.6	38.9	[14]
5.	Annona senegalensis	L. donovani p	Leaves	50% aqueous ethanol	273.5	25.32	[13]
			Stem bark		127.9	4.60
6.	Anthocleista nobilis	L. donovani p	Leaves	50% aqueous ethanol	245.7	5.92	[13]
			Root		716.5	9.07
7.	Argemone mexicana	L. donovani p	Aerial part	Petroleum ether	52.1	0.95	[16]
8.	Artemisia absinthium	L. major p	Leaves	Essential oils	11.22	7.5	[17]
9.	Artemisia campestris	L. major p	Leaves	Essential oils	21.12	9.6	[17]
10.	Artemisia herba-alba	L. major p	Leaves	Essential oils	11.24	9.4	[17]
11.	Artemisia herba-alba	L. major p	Aerial part	Methanol	131.5	2.38	[18]
		L. infantum p			131.5	1.86
12.	Azadirachta indica	L. infantum a	Leaves	Oil	703.8	46	[19]
		L. tropica a			721.6	41
13.	Baphia nitida	L. donovani p	Stem bark	50% aqueous ethanol	990.7	28.8	[13]
14.	Bidens pilosa	L. donovani p	Whole plant	50% aqueous ethanol	192.8	6.67	[13]
15.	Bridelia ferruginea	L. donovani p	Leaves	50% aqueous ethanol	392.9	23.81	[13]
16.	Bocageopsis multifolia	L. amazonensis p	Leaves	Ethanol	26.5	0.7	[12]
		L. braziliensis p			26.7	1.4
17.	Boswellia serrata	L. donovani a	Resin	Polar fractions of dichloromethane	33	38	[20]
18.	Capparis spinosa	L. tropica p	Fruits	Methanol	44.6	9.1	[21]
				Aqueous	28.5	8.4
19.	Cassia gloca	L. tropica p	Leaves	Methanol	1030	-	[22]
20.	Cassia alata	L. donovani p	Leaves	50% aqueous ethanol	371.5	36.78	[13]
21.	Cassia sieberiana	L. donovani p	Leaves	50% aqueous ethanol	62.90	0.77	[13]
22.	Cedrela spp.	L. amazonensis p	Bark	Ethanol	66.3	1.8	[12]
		L. braziliensis p			67.4	3.7
23.	Ceiba pentandra	L. donovani P	Stem bark	50% aqueous ethanol	160.7	3.32	[13]
24.	Celtis australis	L. tropica p	Leaves	Methanol	1209	-	[22]
25.	Cinnamomum cassia	L. donovani a	Barks	Dichloromethane fraction	No cytotoxicity at 500 µg/mL	-	[38]
26.	Citrus sinensis	L. tropica p	Leaves	Methanol	1755	-	[22]
27.	Clausena anisata	L. donovani p	Roots	50% aqueous ethanol	29.2	24.23	[13]
28.	Cleistopholis patens	L. donovani p	Stem bark	50% aqueous ethanol	214.9	3.57	[13]
29.	Cola cordifolia	L. donovani p	Stem bark	50% aqueous ethanol	465.6	18.55	[13]
			Leaves		465.6	25.58
30.	Cola acuminata	L. donovani p	Stem bark	50% aqueous ethanol	156.8	3.28	[13]
31.	Cynara scolymus	L. tropica p	Stem leaves	Ethanol	40.0	4.96	[24]
32.	Ejije bidu	L. amazonensis p	Leaves	Ethanol	133.5	7.5	[12]
		L. braziliensis p			133.0	10
33.	Erica arborea	L. major p	Flower	Methanol	89.6	2.04	[18]
		L. infantum p			89.6	1.46
34.	Eryobotrya japonica	L. tropica p	Leaves	Methanol	1903	-	[22]
35.	Eugenia uniflora	L. amazonensis a	Leaves	n-Hexane	50.5	3.6	[25]
36.	Eugenia uniflora	L. donovani p	Seed	50% aqueous ethanol	94.4	3.55	[13]
37.	Ferula communis	L. aethiopica a,	Aerial part	80% methanol	175.22	-	[26]
		L. donovani a,
38.	Ficus capensis	L. donovani p	Stem bark	50% aqueous ethanol	56.6	1.53	[13]
			Leaves		257.8	2.90
39.	Glyphaea brevis	L. donovani p	Leaves	50% aqueous ethanol	962.2	22.17	[13]
40.	Handroanthus serratifolius	L. amazonensis p	Lapachol	Lapachol	3405.8	42.6	[39]
		L. infantum p				33.0
41.	Iresine diffusa	L. amazonensis p	Flower	Ethanol	39.7	1.3	[12]
		L. braziliensis p			11.1	1.7
42.	Jacaranda glabra	L. amazonensis p	Bark	Ethanol	18.9	6.4	[12]
		L. braziliensis p			191.4	11
43.	Khaya grandifolia	L. donovani p	Stem bark	50% aqueous ethanol	50.1	1.16	[13]
44.	Lantana camara	L. amazonensis a	Leaves	Aqueous	125.9	>9	[28]
45.	Licania salicifolia	L. panamensis a	Leaves	Ethyl acetate	>200	>20.4	[29]
46.	Lophira lanceolata	L. donovani p	Stem bark	50% aqueous ethanol	45.962	0.67	[13]
			Roots		38.9	0.59
47.	Marrubium vulgare	L. major p	Leaves	Methanol	107.4	2.34	[18]
		L. infantum p			107.2	3.01
48.	Mitragyna inermis	L. donovani p	Leaves	50% aqueous ethanol	193.2	8.82	[13]
			Stem bark		424.5	15.16
49.	Mondia whitei	L. donovani p	Roots	50% aqueous ethanol	434.5	13.97	[13]
50.	Murraya koenigii	L. donovani p	Stem	Petroleum ether	73.9	1.32	[16]
51.	Oreopanax floribundus	L. panamensis a	Leaves	Dichloromethane	47.4	2.0	[29]
				Ethyl acetate	54.1	2.2
52.	Otostegia integrifolia	L. aethiopica a,	Aerial part	80% methanol	144.55	-	[26]
		L. donovani a,p
53.	Parkia clappertoniana	L. donovani p	Leaves	50% aqueous ethanol	112.7	6.63	[13]
			Stem bark		42.4	2.41
54.	Persea ferruginea	L. panamensis a	Leaves	Ethyl acetate	>200	>7.8	[29]
55.	Physalis angulata	L. amazonensis p	Flower	Ethanol	19.4	1.1	[12]
		L. braziliensis p			17.4	0.4
56.	Piper pseudoarboreum	L. amazonensis p	Leaves	Ethanol	55.0	1.8	[33]
		L. braziliensis p				2.6
		L. guyanesis p				1.3
		L. infantum p				1.7
57.	Prosopis juliflora	L. donovani p	Leaves	Methanol	0.85	0.26	[34]
58.	Prosopis laevigata	L. amazonensis a	Leaves	Dichloromethane	57.0	7	[28]
59.	Prunus armeniaca	L. tropica p	Leaves	Ethanol	1912.31	-	[44]
60.	Psychotria buhitenii	L. panamensis a	Leaves	Dichloromethane	76.8	3.57	[29]
				Ethyl acetate	109.5	7.75
				Ethanol	>200	>6.81
61.	Pyrus communis	L. tropica p	Leaves	Ethanol	1411.30	-	[35]
62.	Pyrus pashia	L. tropica p	Leaves	Ethanol	1230.66	-	[35]
63.	Schinus molle	L. amazonensis a	Leaves	Dichloromethane	69.7	5	[28]
				Dichloromethane: Methanol (1:1)	186.8	6
64.	Schinus terebinthifolia	L. amazonensis p	Fruits	n-Hexane	52.0	3.7	[25]
65.	Scoparia dulcis	L. amazonensis p	Aerial part	Ethanol	71.7	3.0	[12]
		L. braziliensis p			72.8	2.9
66.	Solanum lycocarpum	L. mexicana a	Fruits	Solamargine	1515.5	43.3	[43]
				Solasonine	1397.9	38.3
67.	Spondias mombin	L. donovani p	Leaves	50% aqueous ethanol	55.42	0.68	[13]
68.	Tamarindus indica	L. donovani p	Leaves	50% aqueous ethanol	77.9	1.34	[13]
69.	Terminalia ivorensis	L. donovani p	Leaves	50% aqueous ethanol	939.2	37.72	[13]
70.	Tessaria integrifolia	L. amazonensis p	Leaves	Ethanol	119.2	2.2	[12]
		L. braziliensis p			120.0	3.8
71.	Thalia geniculata	L. amazonensis p	Roots	Ethanol	50.7	1.7	[12]
		L. braziliensis p			50.4	2.9
72.	Thonningia sanguinea	L. donovani p	Whole plant	50% aqueous ethanol	286.1	15.38	[13]
73.	Treculia africana	L. donovani p	Stem bark	50% aqueous ethanol	172.0	3.84	[13]
74.	Urtica dioica	L. major p	Leaves	Aqueous	4500	4.4	[44]
75.	Vitex fosteri	L. donovani p	Leaves	50% aqueous ethanol	114.4	1.58	[13]
			Stem bark		420.3	8.44
76.	Ximenia americana	L. donovani p	Stem and twigs	50% aqueous ethanol	42.3	1.17	[13]
77.	Zanthoxylum zanthoxyloides	L. donovani p	Roots	50% aqueous ethanol	247.1	18.30	[13]
			Stem bark		583.5	12.91
78.	Ziziphus spina-christi	L. major a	Leaves	Methanol	563.3	10.31	[37]

).

For isolated compounds, 49 natural compounds have been identified with anti-Leishmania activities against amastigotes and/or promastigotes of different species (L. infantum, L. major, L. amazonensis, L. donovani and L. braziliensis). The IC₅₀ values were in the range of 0.2 µg/mL (colchicoside against promastigotes of L. major) to 42.4 µg/mL (dehydrodieuginol against promastigotes of L. amazonensis) (

Table 6. Anti-Leishmania activity of isolated natural compounds.

No.	Compound Name	Leishmania Species	Stage	Assay	Values (IC50)	Data Analysis (Activity)	Authors
1	2,3-Dihydrobenzofuran	L. amazonensis	Promastigotes	In vitro	1.04 µg/mL	High	[45]
			Amastigotes		1.4 µg/mL	High
2	Dehydrodieuginol	L. amazonensis	Promastigotes	In vitro	42.4 µg/mL	Moderate	[31]
3	Erytro-manassatin A	L. amazonensis	Promastigotes	In vitro	35.4 µg/mL	Moderate	[46]
			Amastigotes		20.4 µg/mL	Moderate
4	Threo-manassatin A	L. amazonensis	Promastigotes	In vitro	17.6 µg/mL	Moderate	[46]
			Amastigotes		16.0 µg/mL	Moderate
5	Epipinoresinol-4-O-β-D-glucopyranoside	L. major	Promastigotes	In vitro	36.5 µg/mL	Moderate	[47]
6	Calanolide E1	L. major	Promastigotes	In vitro	36.5 µg/mL	Moderate	[48]
7	Calanolide E2	L. major	Promastigotes	In vitro	29.1 µg/mL	Moderate	[48]
8	Caffeic acid	L. infantum	Promastigotes	In vitro	12.5 µg/mL	Moderate	[49]
			Amastigotes		21.9 µg/mL	Moderate	[50]
10	Capsaicin	L. infantum	Promastigotes	In vitro	5.01 µg/mL	High	[51]
			Amastigotes		24.2 µg/mL	Moderate
11	Cassine	L. amazonensis	Promastigotes	In vitro	25.2 µg/mL	Moderate	[52]
12	Spectaline	L. amazonensis	Promastigotes	In vitro	15.8 µg/mL	Moderate	[52]
13	Berberine	L. donovani	Promastigotes	In vitro	4.8 µg/mL	High	[53]
14	Colchicoside	L. major	Promastigotes	In vitro	0.2 µg/mL	High	[54]
			Amastigotes		4.0 µg/mL	High
15	Bisabolol	L. donovani	Visceral leishmaniasis	In vivo	39.4 µM	Moderate	[55]
16	2-Demethyl colchicine	L. major	Promastigotes	In vitro	0.5 µg/mL	High	[54]
			Amastigotes		10.2 µg/mL	Moderate
17	3-Demethyl colchicine	L. major	Promastigotes	In vitro	0.4 µg/mL	High	[54]
			Amastigotes		11.1 µg/mL	Moderate
18	Cornigerine	L. major	Promastigotes	In vitro	0.8 µg/mL	High	[54]
			Amastigotes		11.9 µg/mL	Moderate
19	Piperine	L. infantum	Promastigotes	In vitro	3.03 µg/mL	High	[51]
20	Colchicine	L. major	Promastigotes	In vitro	0.4 µg/mL	High	[54]
			Amastigotes		8.7 µg/mL	High
21	N-deacetyl-N-formyl colchicine	L. major	Promastigotes	In vitro	0.5 µg/mL	High	[54]
			Amastigotes		10.2 µg/mL	Moderate
22	Colchifoline	L. major	Promastigotes	In vitro	0.7 µg/mL	High	[54]
			Amastigotes		14.0 µg/mL	Moderate
23	Demecolcine	L. major	Promastigotes	In vitro	0.7 µg/mL	High	[54]
			Amastigotes		14.8 µg/mL	Moderate
24	Staurosporine	L. amazonensis	Promastigotes	In vitro	0.08 µM	High	[56]
			Amastigotes		10.0 µM	High
		L. donovani	Promastigotes		2.1 µM	High
25	7-Oxostaurosporine	L. amazonensis	Promastigotes	In vitro	3.6 µM	High	[56]
			Amastigotes		0.1 µM	High
		L. donovani	Promastigotes		0.6 µM	High
26	4′-Demethylamine-4′- oxostaurosporine	L. amazonensis	Promastigotes	In vitro	17.1 µM	Moderate	[56]
			Amastigotes		2.0 µM	High
27	Streptocarbazole B	L. amazonensis	Promastigotes	In vitro	10.4 µg/mL	Moderate	[56]
			Amastigotes		2.5 µg/mL	High
28	3-O-acetylspectaline	L. donovani	Promastigotes	In vitro	25.9 µg/mL	Moderate	[53]
29	3-O-acetylcassine	L. donovani	Promastigotes	In vitro	30.3 µg/mL	Moderate	[53]
30	Soranjidiol	L. amazonensis	Promastigotes	In vitro	16.3 J/cm2	Moderate	[57]
31	Epigallocatechin 3-O- gallate	L. infantum	Visceral leishmaniasis	In vivo	ED50 = 12.4 mg/kg/day	Moderate	[58]
32	5-Chlorosoranjidiol	L. amazonensis	Promastigotes	In vitro	13.8 J/cm2	Moderate	[58]
33	Bisoranjidiol	L. amazonensis	Promastigotes	In vitro	15.2 J/cm2	Moderate	[58]
34	Gallic acid	L. major	Promastigotes	In vitro	23.0 µg/mL	Moderate	[32]
35	Calanolides E1	L. infantum.	Amastigotes	In vitro	37.1 µM	Moderate	[48]
36	Calanolides E2				29.1 µM	Moderate
37	Apigenin	L. amazonensis	Promastigotes	In vitro	23.7 µM	Moderate	[59]
			Amastigotes		4.3 µM	High
38	2′-hydroxyflavanone	L. amazonensis	Promastigotes	In vitro	20.5 µM	Moderate	[60]
			Amastigotes		3.09 µM	High
39	5,7,3′,4′-tetrahydroxy- 6,8-diprenylisoflavone	L. amazonensis	Promastigotes	In vitro	2.7 µM	High	[61]
			Amastigotes		1.1 µM	High
40	Brachydin B	L. braziliensis	Promastigotes	In vitro	7.05 µM	High	[62]
41	Brachydin C	L. amazonensis	Promastigotes	In vitro	10.0 µM	High	[62]
			Amastigotes		6.25 µM	High
		L. braziliensis	Promastigotes		8.8 µM	High
42	Ursolic acid	L. amazonensis	Promastigotes	In vitro	6.2 µg/mL	High	[63]
		L. donovani	Amastigotes		1.8 µM	High
43	Aplysulphurin	L. donovani	Amastigotes	In vitro	3.1 µM	High	[64]
44	Tetrahydroaplysulphurin-1	L. donovani	Amastigotes	In vitro	3.5 µM	High	[64]
45	Membranolide	L. donovani	Amastigotes	In vitro	9.7 µM	High	[64]
46	Apigenin	Cutaneous leishmaniasis	Cutaneous leishmaniasis	In vivo	ED50 = 0.73 mg/kg	High	[65]
47	Darwinolide	L. donovani	Amastigotes	In vitro	11.2 µM	Moderate	[63]
48	Pukalide aldehyde	L. donovani	Amastigotes	In vitro	1.9 µM	High	[66]
49	Epigallocatechin 3-O- gallate	L. infantum	Amastigotes	In vitro	2.6 µM	High	[58]

).

Numerous natural compounds were isolated from different parts of the plants that were used in traditional medicine to treat leishmaniasis [67]. These compounds act against Leishmania by various mechanisms including the disintegration of cytoplasmic membranes, electron flow disturbances, active transport of crucial substances, coagulation of the cell contents, and destabilization of proton motive forces [68]. For example:

• Some medicinal plants are enriched with essential oils composed of different hydrophobic molecules which can diffuse easily across cell membranes and consequently gain access to intracellular targets [67,69]. They may also act on ATPases and other proteins located in cytoplasmic membranes that are surrounded by lipid molecules. They can also cause a distortion of lipid–protein interactions in hydrophobic parts of the proteins, or they can interact with the enzymes involved in the synthesis of structural sections.
• The diversity of terpenoids increases their biological activity spectrum, including several Leishmania species [70]. Terpenes can easily penetrate the lipid bilayer of the cell membrane and produce changes in the integrity of cell structure and the mitochondrial membrane of Leishmania parasites [67]. For example, Artemisinin induced apoptosis, depolarization of the mitochondrial membrane potential, and DNA fragmentation [71,72]. Ursolic acid induce programmed cell death independent of caspase 3/7 but dependent on mitochondria. The compound reduced the lesion size and parasite load of cutaneous leishmaniasis in vivo [70]. (−)-α-Bisabolol induced phosphatidylserine externalization and caused cytoplasmic membrane damage, both of which are apoptosis indicators. The compound also decreased ATP levels and disrupted the mitochondrial membrane potential [73].
• Plants enriched with antioxidant compounds such as flavonoids may act by initiating morphological changes and causing a loss of cellular integrity, leading to cell cycle arrest in the G1 phase [59]. They also may act by damaging the mitochondria of the parasites [67]. For example, apigenin increased intracellular reactive oxygen species (ROS) and the number of double-membrane vesicles as well as myelin-like membrane inclusions, which are characteristics of the autophagic pathway. Furthermore, the fusion between autophagosome-like structures and parasitophorous vacuoles was observed [65]. Epigallocatechin 3-O-gallate (EGCG) has increased ROS levels, which decreased the mitochondrial membrane potential and the ATP levels [58].
• The diversity of structures within the coumarin group enables them to exhibit many biological activities, including anti-Leishmania activity. It represents a promising natural compound that can act on two fronts: as a treatment for leishmaniasis (able to induce mitochondrial membrane damage and changes in ultrastructure [74] and as a tool to control Leishmania vectors (might block the transmission of leishmaniasis since they decrease parasite loads [27].
• Many alkaloids have been described as having biological activities against trypanosomatids, such as Leishmania spp. For example, heterocyclic steroids (solamargine and solasonine) induced different immunochemical pathways in macrophages and dendritic cells. Additionally, they were capable of enhancing the expression levels of transcription factors, such as NFκB/AP-1 [43]. In addition, isoquinoline alkaloid (berberine) has leishmanicidal activity through a reduction in the viability of promastigotes and the generation of ROS in these cells. It also increased the levels of mitochondrial superoxide and induced the depolarization of mitochondrial transmembrane potential [53].

3. Methods

3.1. Study Design and Setting

In order to perform this review, the following aspects were addressed: identification and selection of the theme of the research question, establishment of criteria for selection of the sampling, the definition of information to be extracted from selected studies, assessment of the studies included in the integrative review, and final explanation of the results.

3.2. Search Strategies

The databases used for this article were PubMed, Google Scholar, Web of Science, Research Gate, SCOPUS, and Scientific Electronic Library Online (SciELO) using the keywords: neglected tropical disease, Leishmania species, anti-Leishmania activity, natural product, medicinal plants, and promastigote form. We used the search terms separately and in combination with the Boolean operators “OR” or “AND”.

3.3. Inclusion and Exclusion Criteria

The initial total articles (1374) were adjusted for the restriction in the year of publication (from 1 January 2015 to 31 December 2021) (806), duplicates (273), articles that were not available in full (67) and articles in other languages (4). After a review of their titles and abstracts, some articles were discarded, since the anti-leishmanial activity (IC₅₀) values were higher than 100 µg/mL (134), and they tested extract/natural compounds obtained through other natural sources (algae, fungi, etc.) (11). The full texts of the remaining articles were reviewed in detail. However, further articles were discarded after the full text had been reviewed (18) since they did not address much of the required information. Finally, 61 articles were evaluated as valuable to reach the goals of this review. The methodological validity of all 61 studies was proven prior to inclusion in the review by undertaking a critical appraisal using a standardized instrument [75].

3.4. Data Extraction and Analysis

The data extraction protocol included the scientific and family names, parts of the plant used, most active extract/ essential oil employed in the experiment, name of natural compound, Leishmania species and form, IC₅₀ values, potential groups/compounds responsible for activity, clinical form of leishmaniasis, route, the dose of administration and scheme of treatment, the efficacy of the treatments in the experiment, cytotoxic activity, selectivity index, the authors, and year of publication. In the results analysis, an active extract/compound was considered if the IC50 value was less than or equal to 10 µg/mL against the promastigote or amastigote forms. Moderate activity was defined if the IC50 was greater than 10 and less than 50 µg/mL and weakly active if the IC50 value was greater than 50 µg/mL and less than 100 µg/mL.

4. Conclusions and Perspectives

Leishmaniasis threatens about 350 million people around the world and continues to represent a menace on a global scale. Without a doubt, it requires utmost attention due to the lack of vaccines for the prevention and reported resistance against available chemical drugs for treatment. The intolerably high incidence of millions of new cases of leishmaniasis per year worldwide and deficiencies in current treatment point to an urgent need for new medications.

As a means to facilitate the accessibility of information, this review updates and summarizes recent results on medicinal plants and natural compounds against different Leishmania species. The plants presented here have demonstrated a diverse range of activities against different forms of leishmaniasis with some showing high activities that could be reasonable starting points for the further development of effective and affordable novel drugs.

However, it was also evident that the majority of experiments were performed with the promastigote form. We believe that these studies are undoubtedly important because promastigotes are infectious to man and other animals. However, it is urgent that future studies should be conducted to find compounds with anti-amastigote activity too, since the morbimortality associated with Leishmania is caused by this form.

It Is pleasing that more and more investigations report on the anti-Leishmania activity in vivo and more studies are needed in this respect, increasing the number of potential candidate compounds for further drug development. In vitro studies are valuable for the screening of extracts and isolated compounds as well as for investigations of the cellular and molecular modes of action. Since many natural compounds are rapidly metabolized in the human body by liver enzymes and gastrointestinal microflora, animal experiments are indispensable to identify candidates with sufficient half-life times in vivo and anti-Leishmania activities in concentration ranges that are reachable in the human blood. However, in the literature inspected by us, only four plants and two natural compounds have been investigated both in vitro and in vivo, i.e., Prosopis juliflora [34], Ziziphus spina-christi [37], Piper pseudoarboreum [33], and Croton caudatus [76] as well as epigallocatechin 3-O-gallate [58] and apigenin [65], respectively. More investigations are required to allow a direct comparison of in vitro and in vivo data.

Further down this line of argumentation, standardized extracts and/or isolated phytochemicals need to be tested in randomized clinical trials. Without convincing clinical evidence on safety and efficacy, preparations from traditional medicine will hardly reach considerable recognition in the medical world.