Fernandes-Pedrosa F. , Silvana M. Zucolotto Júlia M. Fernandes , Lorena M. Cunha , Eduardo Pereira Azevedo , Estela M.G. Lourenc¸ o ,Matheus ethnopharmacology, phytochemistry, pharmacology andtoxicology Kalanchoe laciniata and Bryophyllum pinnatum : an up

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w ww . e l s e v i e r . c o m / l o c a t e / b j p

Review

Kalanchoe laciniata and Bryophyllum pinnatum: an updated review about ethnopharmacology, phytochemistry, pharmacology and toxicology

Júlia M. Fernandes

^a

, Lorena M. Cunha

^a

, Eduardo Pereira Azevedo

^c

, Estela M.G. Lourenc¸ o

^a

, Matheus F. Fernandes-Pedrosa

^b

, Silvana M. Zucolotto

^a,∗

aLaboratóriodeFarmacognosia,DepartamentodeFarmácia,UniversidadeFederaldoRioGrandedoNorte,Natal,RN,Brazil

bLaboratóriodeTecnologiaeBiotecnologiaFarmacêutica,UniversidadeFederaldoRioGrandedoNorte,Natal,RN,Brazil

cLaboratóriodeFarmacotécnica,DepartamentodeFarmácia,UniversidadeFederaldoRioGrandedoNorte,Natal,RN,Brazil

a r t i c l e i n f o

Articlehistory:

Received19September2018 Accepted30January2019 Availableonline7June2019

Keywords:

Comparison Flavonoid Traditionaluse Phytochemical Pharmacology Toxicology

a b s t r a c t

ThespeciesKalanchoelaciniata(L.)DC.andBryophyllumpinnatum(Lam)Pers.arenativefromBraziland Madagascar,respectively.BothbelongingtotheCrassulaceaefamilyandbeingwidelyusedbypopu- lationasanaturalanti-inflammatoryagent.Thesespecieshavesimilarleafmorphologyandforthis reason,theyareknownbythesamepopularnameas“saião”or“coirama”.Severalstudieshavebeen publishedinvolvingdifferentpartsandpreparationsofthesespecies.Therefore,thisreviewaimstopro- videanupdateoverviewaboutthetraditionaluses,chemicalconstitution,pharmacologyandtoxicology ofK.laciniataandB.pinnatumspecies.Anextensiveliteraturereviewwasconductedindifferentsci- entificdatabases.Variouschemicalconstituentshavebeenidentifiedinextractsfromdifferentpartsof K.laciniataandB.pinnatum,beingflavonoidsthemajorcompounds.Theyhavebeentraditionallyused totreatinflammation,microbialinfection,pain,respiratorydiseases,gastritis,ulcers,diabetesandcan- certumors.Non-clinicalinvitroassaysevaluatedmainlytheantimicrobialandantioxidantactivities, whileinvivoassaysevaluatedtheleishmanicide,anti-inflammatoryandimmunomodulatoryactivities.

Regardingtoxicity,fewstudieshavebeenconductedforthetwospecies.Theinformationreportedin thisworkmightcontributetotherecognitionoftheimportanceofK.laciniataandB.pinnatumspecies, aswellastodirectfurtherstudies.

4.0/).

Introduction

The Crassulaceae family comprises approximately 33 genera and 1500 species distributed worldwide, except for Australia and the Paciﬁc Islands (Allorge-Boiteau,

1996).

This family presents xeromorphic characteristics that allows its species to adapt to bright light and water scarcity (Herrera,

2008).

It has an important role on the research of biochemical, ecophysiological and phyloge- netic aspects related to the Crassulacean Acid Metabolism (CAM), which is an evolutive adaptation of the pathway of photosynthetic carbon assimilation (Osmond,

1978).

Among species, Kalanchoe

∗ Correspondingauthor.

E-mail:silvanazucolotto@ufrnnet.br(S.M.Zucolotto).

laciniata (L.) DC. is native from Brazil and Bryophyllum pinnatum (Lam) Pers. from Madagascar (Allorge-Boiteau,

1996;Gehrigetal., 2001).

Even though these plants are naturalized in Brazil, they are not endemic (Zappi,

2015).

Kalanchoe laciniata and B. pinnatum are both popularly known as “saião” or “coirama” and have been used to treat inﬂammatory disorders (Amaral

etal.,2005).

Due to the various ethnopharmaco- logical reports attributed to these species, several research groups have conducted studies to prove their pharmacological or biological properties. In addition, some researchers have carried out phyto- chemical studies resulting in the identiﬁcation of different classes of secondary metabolites, as well as the isolation of various con- stituents, specially from their leaves and aerial parts. Therefore, this review aims to provide an updated overview about the tradi- tional uses, chemical constitution as well as pharmacological and

https://doi.org/10.1016/j.bjp.2019.01.012

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Fig.1.Leaves(A)andinﬂorescences(B)ofKalanchoelaciniata(L.)DC.

toxicological aspects of K. laciniata and B. pinnatum species. This study might be used as a guide to further investigations involving these species.

Materialandmethods

An extensive literature review was conducted in different scien- tific databases such as PubMed, Web of Science, Scopus, Scielo, The Cochrane Library. The study covered several aspects of the vegetal species like botany, phytochemistry, traditional uses, pharmacol- ogy and toxicology. In addition, the scientific names, synonyms and popular names of major species identified by the botanical databases “Flora do Brasil”, Tropicos, International Plant Names Index and The Plant List were included. The common names found in the book “Coletânea científica de plantas de uso medicinal”

(Amaral

etal.,2005)

were also used. The data were updated in April 2018.

Botanical information

Kalanchoe laciniata and Bryophyllum pinnatum exhibit similari- ties concerning their leaf morphologies, which include decussate, succulent, glabrous, oval to elliptical leaves with crenate border (Hyakutake

and Grotta,1972; Anjooand Saluja, 2010;Moreira etal.,2012).

Due to such similarities, these species are known by the same popular name (Moreira

etal.,2012).

The knowledge of leaf anatomy is important for registration purpose and for quality control of herbal medicines (Moreira

etal.,2012;Anvisa,2014).

The K. laciniata (L.) DC. specie (Fig.

1)

is popularly known as “saião”, “corama-branca”, “folha-da-fortuna”, “para-tudo”,

“fortuna-de-ﬂores-amarelas”, “folha-da-costa”, “folha-grossa” in Alagoas, “coerana” in Pernambuco, and “erva-da-costa” in Bahia.

This species is found in almost all states of the Northeast region (Rio Grande do Norte, Ceará, Pernambuco, Paraíba, Bahia and Sergipe), Southeast (Espírito Santo, Rio de Janeiro, Minas Gerais and São Paulo), South (Paraná and Santa Catarina), Midwest (Federal Dis- trict and Mato Grosso do Sul) and North (Acre) regions, mainly in the coastal zone (Allorge-Boiteau,

1996;Amaraletal.,2005;Zappi, 2015).

The accepted name for this specie is K. laciniata (L.) DC.

and others botanics synonyms are Kalanchoe brasiliensis and Kalan- choe crenata (The

PlantList,2010;Zappi,2015;Tropicos,2019).

Although K. laciniata is the accepted name, most of the works found use its synonym of K. brasiliensis.

Kalanchoe laciniata constitutes a subligneous and perennial veg- etable with 30–100 cm in height. Its leaves are succulent, oval or obval, opposites, shortly petiolate and crenate (Corrêa,

1984;

Barroso,1991;Amaraletal.,2005).

The ﬂowers are yellow-orange in collor, small, abundant, arranged in composite summits of stamps or paniculate, hermaphrodites, gamopetalas with corolla longer than the cup, with the presence of scaly carpels that become polispermos follicles. Their fruit is a follicle with 6 cm long that con- tains brown oblong seeds (Lorenzi

andMatos,2000;Amaraletal., 2005).

The Bryophyllum pinnatum (Lam) Pers. species (Fig.

2)

is popularly known as “saião” and “coirama” throughout Brazil;

“folha-de-pirarucu”, in Pará State; “fortuna” and “roda-da-fortuna”

in Minas Gerais; “zakham-hayat” in Asia and Africa; life-plant in Mexico; love-plant, canterburry, bells and cathedral bells in the United States of America and Europe (Amaral

etal.,2005;Joseph etal.,2011).

This species is found in Brazil, China, India and Africa and in all tropical countries. In Brazil, it is found in Northeast (Bahia, Ceará and Paraíba), North (Acre), Southeast (Espírito Santo, Rio de Janeiro, Minas Gerais and São Paulo), Midwest (Distrito Fed- eral, Mato Grosso do Sul and Mato Grosso) and South (Paraná, Rio Grande do Sul and Santa Catarina) regions, mainly in the coastal and Caatinga zones (Amaral

etal.,2005;Zappi,2015).

B. pinnatum (Lam) Pers. is the accepted name and B. calycinum and Kalanchoe pinnata are its botanic synonyms (The

PlantList,2010;Zappi,2015;

Tropicos,2019).

Bryophyllum pinnatum is a perennial, succulent and corpulent vegetable with glabrous and tuberous stem. This species can reach up to 150 cm in height. The oldest stalks have a light color while the youngest ones are reddish with defilements. Its leaves are vari- able and decussates, being the lowest ones generally simple or sometimes imparinates. The leaves are 30 cm long, the upper are 3-5-7 foliated, long petiolate, thick, fleshy and dark with crenate borders. The inflorescences are hermaphrodites, tubular, pendu- lous, monopetalas, pale green or yellow-red, with cup swollen and corolla longer than the cup. The fruit are in the form of hoods which become scaly polispermos follicles that are housed within the hoods (Amaral

etal.,2005;Jessica,2008;Josephetal.,2011;

Moreiraetal.,2012).

The main characteristic that differentiates K. laciniata and B. pin-

natum species is the leaf aspect as K. laciniata has a corrugated or

subcrenated border (Fig.

3),

whereas B. pinnatum leaf is signiﬁcantly

crenate (Lorenzi

andMatos,2000).

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Fig.2. Leaves(A)andinﬂorescences(B)ofBryophyllumpinnatum(Lam)Pers.

Fig.3. LeavesofKalanchoelaciniata(A)andBryophyllumpinnatum(B).

Chemical constituents

Various chemical constituents for K. laciniata and B. pinnatum have been reported in the literature, where they have been isolated mainly from the leaves of both species. The aqueous and methanolic are the most commonly used extracts of K. laciniata and B. pinnatum, respectively. Among the constituents that have been identiﬁed so far, ﬂavonoids represent the class of secondary metabolites most commonly found, being the major component in both species.

However, for K. laciniata, some patuletin aglycone derivatives have been identiﬁed, whereas for B. pinnatum, quercetin, kaempferol and luteolin aglucons were found. In addition, chlorophylls, carotenoids and polysaccharides have been identiﬁed for K. laciniata.

Despite the widespread use of K. laciniata and B. pinnatum by folk medicine practitioners, their chemical constituents have not been fully elucidated. Moreover, only few studies involved the isolation and elucidation of their chemical constituents, where the major- ity of the studies indicates the presence of glycosylated flavonoids, derived from the acetylation of the rhamnose ring of patuletin in different positions. The acetylation occurs at the end of the biosyn- thetic route of flavonoids, usually through esterifications of the hydroxyl groups (Aguiar

etal.,2007).

Flavonoids derived from pat- uletin have already been described for another species of the genus, K. spathulata (Gaind

etal.,1981).

Acetylated rhamnose ﬂavonoids

have been described for B. pinnatum but derived from canferol (Tatsimo

etal.,2012).

However, a quick search in the literature reveals that acetylated rhamnoses of patuletin have only been described for K. laciniata and K. gracilis by

Costaetal.(1994)

and

Liuetal.(1989),

respectively. Thus, these compounds are consid- ered as potential candidates for speciﬁc markers of these species by presenting themselves as differentiators in relation to most other species, especially B. pinnatum. It is worth pointing out that until the date of this publication, no study have been published about the description of the bioactive compounds for this specie as only two studies were conducted by bioassay-guided isolation by

Costa etal.(1994)

and

Trevisanetal.(2006).Box1

summarizes the main compounds already described for K. laciniata.

Widely used by the general population, B. pinnatum species

has several studies that dealt with the isolation and elucidation

of its major compounds. Several classes of chemical compounds

have been described for B. pinnatum, which includes fatty acids,

acyclic and aromatic organic acids, amino acids, sugars, vitamins,

minerals, bufadienolides, ketones, fenantrenics derivatives, sterols,

ﬂavonoids, long chain hydrocarbons, triterpenoids, phenolic acids,

saponins and gums (Amaral

et al., 2005).

However, ﬂavonoids,

steroids and terpens are the compounds most frequently isolated

from B. pinnatum. In relation to ﬂavonoids, glycosylated derivatives

from the quercetin, kaempferol and luteolin aglycones have been

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Box1:ChemicalcompoundsreportedforKalanchoelaciniataspecies.

Plantpart Classiﬁcation Extract Compound Reference

Freshleaves,green callus

Chlorophylls, carotenoids

Peroxide-freediethyl etherfractionof80%

(v/v)acetone–water extract

– Stobartetal.(1967)

Freshstems,leaves Flavonoids Juice KalambrosideA(1);

KalambrosideB(2);

KalambrosideC(3);

Patuletin3-O-␣-L- rhamnopyranosyl-7-O- (3-O-acetyl-␣-L- rhamnopyranoside)(4);

Patuletin

3-O-(4-O-acetyl-␣-^L- rhamnopyranosyl)-7-O- (3-O-acetyl-␣-L- rhamnopyranoside)(5);

Patuletin3-O-␣-L- rhamnopyranoside(6);

Patuletin3-O-␣-L- rhamnopyranosyl-7-O-

␣-^L-rhamnopyranoside (7)

Costaetal.(1994)

Leaves Flavonoidsand

carbohydrates

Aqueousandbutanolic fractionofjuice

– Almeidaetal.(1997)

Leaves Flavonoids Ethylacetateextract 3,7-Di-O-␣-L-

rhamnopyranosyl-8- methoxyquercetin (8)

Trevisanetal.(2006)

3,7-Di-O-␣- rhamnopyranosyl- kaempferol (9) 3-O-␣-^L- rhamnopyranosyl- 3,3,4,5,7-pentahidroxi- 8-methoxyﬂavone (10)

Leaves Organicsalt Juice Complexedbetweena

hydroaminoacidand malicacid(1:2)

Costaetal.(2006)

Driedleaves Polysaccharides – – Bhattietal.(2013)

Freshleaves Flavonoids Ethanolicextract50% Patuletin3-O-␣-L-

rhamnopyranosyl-7-O-

␣-L-rhamnopyranoside (7)

Costaetal.(2015)

described for this species. Quercetin 3-O-

␣

-

l

-arabinopyranosyl- (1

→

2)-O-

␣

-

l

-rhamnopyranoside was the ﬁrst derivative isolated for B. pinnatum, as reported by

Muzitanoetal.(2006a,b).

Although quercetin is a common aglycone derivate, its 3-O diglycosidic bond is quite peculiar as it is a rhamnose–arabinose dimer that is not very common and in fact, has never been reported for leaves of other species of the genus Kalanchoe (Nascimento

etal.,2015).

This

very peculiar molecule has been described for few species of other botanical families, but not as the major compound.

Nascimento etal. (2018)

state that this unusual ﬂavonoid can be used as a marker for B. pinnatum. In addition, this major component has shown potent anti-inﬂammatory (Ferreira

etal.,2014)

and leish- manicidal (Muzitano

etal.,2009)

activities. The compounds already described for B. pinnatum are summarized in

Box2

.

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Box2:ChemicalcompoundsreportedtoBryophyllumpinnatumspecies.

Plant part

Classiﬁcation Typeofextract Compound Reference

– Polysaccharides,

minerals,ﬂavonoids

Sap Glycosylatedderivativesofquercetin MaksyutinaandZub

(1969) Leaves Alkanes,triterpenes,

steroids

Petroleumether C26–C34;␣-amyrin;␤-amyrin;Sitosterol GaindandGupta(1972) Fresh

leaves

Organicacids,steroids, hydrocarbons,phenolic compounds,ﬂavonoids

Methanolicextract Bryophynol(11);-taraxasterol(12);

Bryophyllol(13);18␣-oleanane(14);

Bryophollone(15);bryophollenone;

24-ethyl-25-hydroxycholesterol(16)

Siddiquietal.(1989)

Shoots Steroids Dichloromethaneextract Stigmast-24-enol(17);(24S)-stigmast-25-enol (18);25-metylergost-24(28)-enol(19);

(24R)-ergost-5-enol(20);(24S)-ergost-5-enol (21);ergost-5,24(28)-dienol(22);

(24S)-ergosta-5,25-dienol(23);

(24R)-stigmast-5-enol(24);

(22E,24S)-stigmast-5,22-dienol(25);

stigmast-5,24-dienol(26);

(24Z)-stigmast-5,24(28)-dienol(27);

(24R)-stigmast-5,25-dienol(28);(24S)- stigmast-5,25-dienol(29);

25-metylergost-5,24(28)-dieno(30);

stigmast-7,24-dienol(31);

(24Z)-stigmast-7,24(28)-dienol(32);

(24R)-stigmast-7,25-dienol(33);

(24S)-stigmast-7,25-dienol(34)

Akihisaetal.(1991)

Whole fresh plant

Bufadienolide orthoacetate

Chloroformfractionofthe methanolicextractofthe aqueousresidue

Bersaldegenin-1,3,5-orthoacetate(35) Xiuzhenetal.(1992)

Leaves – Acidfraction 89.3%ofpalmiticacid(C16),10.7%ofstearic

acid(C18)andtracesofarachidic(C20)and behenicacids(C22)

Almeidaetal.(2000)

Leaves Bufadienolides Methanolicextract BryophyllinA(36) Supratmanetal.(2000,

2001)

Leaves Bufadienolides Methanolicextract BryophyllinC(37) Supratmanetal.(2001)

Leaves Bufadienolides Methanolicextract Bersaldegenin-3-acetate(38) Supratmanetal.(2001)

Dried leaves

Alkaloids,ﬂavonoids, phenols,tannins, vitamins,minerals

Aqueous,ethanolic,diethyl etherand20%aceticacid extracts

– OkwuandJsiah(2006)

Fresh leaves

Flavonoid Aqueousextract Quercitrin(quercetin

3-O-␣-^L-rhamnopyranoside)(39)

Aokietal.(2014),Cruz etal.(2008),Muzitano etal.(2006a,b) Dried

leaves

Minorvinylicaliphatic alcoholdiglycoside

Ethanolicextract 1-Octen-3-O-␣-L-arabinopyranosyl-(1→6)-␤

glucopyranoside

Almeidaetal.(2006)

Leaves Protein TamponTEGN Rubisco(ribulose1,5-bisphosphate

carboxylase-/Oxigenase)

Abatetal.(2008) Powdered

plant

Alkaloids,tannins, steroids,ﬂavonoids, sugars,organicacids

Petroleumether,chloroform, methanolic,aqueousextracts

– Devbhutietal.(2008)

Leaves Flavonoids Aqueousextract Quercetin3-O-␣-L-arabinopyranosyl(1→2)

␣-L-rhamnopyranoside(40);kaempferol 3-O-␣-L-arabinopyranosyl(1→2)

␣-L-rhamnopyranoside(kapinnatoside)(41)

Cruzetal.(2008), Muzitano(2006)

Leaves Phenolicacids, ﬂavonoids

Ethylacetateandmethanol extract

Gallicacid;caffeicacid;coumaricacid;

kaempferol-3-O-rutinoside(42)

Abdellaouietal.(2010) Dried

leaves

Phenanthrenealkaloid Ethanolicextract 1-Ethanamino-7-hex-1-yne-5-onephenanthrene OkwuandNnamdi (2011a)

Powdered plant

Flavonoids Ethanolic

extract

5ⁱMethyl4ⁱ,5,7-trihydroxyﬂavone Okwu and Nnamdi (2011b) 4ⁱ,3,5,7-Tetrahydroxy-5-methyl-5ⁱ-propenamine;

anthocyanidines Leaves Bufadienolides,

ﬂavonoids,cinnamic acids

Juice – Wächteretal.(2011)

Roots Steroids,alkaloids, saponins,glycosides ﬂavonoids,tannins, carbohydrates, proteins,aminoacids

Aqueous,chloroform,ethanol, etherextract

– Majazetal.(2011)

Leaves Steroids,esters Ethanolicextract Stigmast-4,20(21),23-trien-3-one(43);

stigmata-5-en-3␤-ol(44);

␣-amyrin-␤-D-glucopyranoside(45);

n-undecanylnoctadec-9-en-1-oate;n-dodecanyl n-octadec-9-en-1-oate

Afzaletal.(2012)

Leaves Alkaloids,glycosides, gums,saponins, tannins,reducing sugars

Chloroformextract – Biswasetal.(2012)

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Box2:Continued

Leaves Phenols,ﬂavonoids Petroleumether,chloroform, ethanol95%extract

– Bhattietal.(2012)

Leaves Cardiotonicglycosides, alkaloids

Chloroformextract – Mahataetal.(2012)

Leaves Flavonoids Methanolicextract Kaempferitrin(46);kaempferol

3-O-␣-L-(2-acetyl)

rhamnopyranoside-7-O-␣-^L-rhamnopyranoside (47);kaempferol3-O-␣-^L-(3-acetyl)

rhamnopyranoside-7-O-␣-L-rhamnopyranoside (48);kaempferol3-O-␣-L-(4-acetyl)

rhamnopyranoside-7-O-␣-L-rhamnopyranoside (49);kaempferol3-O-␣-D-glucopyranoside-7-O-

␣-L-rhamnopyranoside(50);afzelin(51);

␣-rhamnoisorobin(52)

Tatsimoetal.(2012)

Air- dried powdered leaves

Flavonolglycosides Petroleumetherextract 7-O-methylkaempferol-3-O-␣-^L-rhamnopyranosyl-(1→6)-O-␤-D-galactopyranoside (53);kaempferol-14-O-␣-L-rhamnopyranosyl- (1→3)-O-␣-L-rhamnopyranosyl-(1→6)-O-␤-D- galactopyranoside

(54)

Bodakheetal.(2013)

Dried leaves

Flavonol Methanolicextract 3,4-Dimethoxyquercetin(55) Darmawanetal.(2013)

Leaves Flavonolglycosides, ﬂavonoids,phenolic glycosides

Methanolicextract 4-O-␤-D-glucopyranosyl-cis-p-coumaricacid;

syringicacid␤-D-glucopyranosylester

Füreretal.(2013)

Shoots Steroid Petroleumetherextract Stigmasterol KambojandSaluja

(2017) Leaves Flavonoids,steroids,

terpenoids,phenolics, alkaloids,glycosides, tannins.

Ethanolicextract – JoshiandChauhan

(2013)

Dried leaves

Flavonoid,phenolic acids

Ethylacetatefractionof methanolicextract

Quercetin Aokietal.(2014)

Chiblietal.(2014) Dried

leaves

Flavonoids,saponins Aqueousextract – Alhajietal.(2014)

Leaves Flavonoids Methanolicextract Rutin;luteolin;luteolin7-O-␤-glucoside(56) Chiblietal.(2014) Leaves Steroidalglycoside,

bufadienolides

Methanol,ethylacetate, petroleumetherextracts

– Tariqetal.(2015)

Leaves Bufadienolides Dichloromethane,ethanol extracts

Bersaldegenin-1-acetate(57);

Bersaldegenin-3-acetate(58);

Bersaldegenin-1,3,5-orthoacetate(59);Bufalin (60)

Ouﬁretal.(2015)

Leaves Phenoliccompounds Acetoneandmethanolextracts KPB-100;KPB-200 Cryeretal.(2017)

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Box2:Continued

Traditional uses

The traditional medicinal uses of K. laciniata and B. pinnatum are shown in

Boxes3and4

, respectively. As can be seen from these boxes, the leaf is the part of the plant mostly used by the popula- tion, whereas the juice is the most frequent mode of preparation for both species. Some of the properties are common for both species and those have been used to treat: (i) skin problems, (ii) problems in the respiratory system, (iii) pain, (iv) inﬂammation and (v) dis- orders in the gastrointestinal system. The inﬂammatory and gastric problems (ulcers and gastritis) are the most commonly treated dis- orders. In addition, K. laciniata and B. pinnatum have been used in the form of plaster or poultice to treat dermatological disorders and burn wounds. The antivenom activities of the leaves of both

species against snake and scorpion bites have also been reported elsewhere.

The extract of K. laciniata leaves has been used for the treat- ment of chilblains, burns, erysipelas, wounds, cough, bronchitis, ﬂu, gastritis, ulcers, otitis, kidney stone, diabetes, anxiety, microbial diseases, snakebite, pain and inﬂammation. In addition, it has been used to treat cancerous tumors, osteoarticular rheumatism, jaun- dice, yellow fever, other liver disorders, headache, prostate tumors and hemorrhoids. K. laciniata have been used mainly in the form of decoction, syrup, juice, poultice and maceration of its leaves (Silva

etal.,2002).

The popular uses of K. laciniata species are reported in

Box3.

The extract of B. pinnatum leaves have been used for the

treatment of severe disorders such as gastritis, ulcers, cough,

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Box3:MedicinalpopularusesofKalanchoelaciniatareportedintheliterature.

Plantpart Popularuse Preparation Reference

Leaves,stem Otitis,skindiseases – Cosetal.(2002)

Leaves Healing – Fonseca-KruelandPeixoto(2004)

Leaves Bronchitis,ﬂuandsore Juice,syrupandpoultice Medeirosetal.(2004)

Leaves Ovariananduterine

inﬂammations

Purejuiceormixedwithotherplantsforsyrup preparation

Moraisetal.(2005)

Leaves – Teaandsyrup Pereiraetal.(2005)

Leaves Ulcersandgastritis – Lisboaetal.(2006)

Leaves Ulcersandgastritis Juice Silvaetal.(2006)

Leaves Coughing,gastritis,diabetes,

painsingeneral

– Albuquerqueetal.(2007a)

Leaves Generalpain,ulcer,gastritis,

injury,generalinﬂammation, asthma,lungproblems,kidney stone

– Albuquerqueetal.(2007b)

Leaves Chilblains,burns,wounds,

erysipelas,ulcers,scurvy,ﬂu andbronchitis

– BoscoloandValle(2008)

Leaves Anxiolyticeffects – Rodriguesetal.(2008)

Leaves Respiratorysystemdisorders – Leitãoetal.(2009)

Leaves Healing,ﬂuandcough Plaster,syrupandjuice Albertasseetal.(2010)

Leaves,root Depurative,bloodthinner, uterineinflammation,cough, influenza,expectorant,healing, painsingeneral,inflammation ingeneral

Decoction,tincture,leavesoaking,refreshment, licking,poultice,warminoilorinfusion

Cartaxoetal.(2010)

Aerialpart Fever,fracture,earpain Maceration Hubertetal.(2013)

Leaves Totreatsnakebitevictims Paste Mouraetal.(2015)

Leaves,aerialparts,roots, wholeplant

Microbialdiseases Decoction,sap,maceration,heatingintheash Ngezahayoetal.(2015)

Leaves Facilitationofdelivery,

bleedingduringpregnancy, postpartumabdominalpain

Decoction,maceration Yemeleetal.(2015)

Leaves Snakebite Paste Mouraetal.(2015),Félix-Silvaetal.(2017)

bronchitis, various bacterial, viral and fungal infections, leishma- niasis, pain, inﬂammation, some tumors, respiratory infections, diabetes, hypertension, ﬂu and fever (Perry

andMetzger, 1980;

Silvaetal.,1995;Moreiraetal.,2002;Medeirosetal.,2004;Amaral etal.,2005;KambojandSaluja,2009).

This species is part of the traditional Indian medicine. The part of the plant mostly used is its leaves, prepared as decoction, infusion, juice, syrup, poultice and paste. The popular uses of the species B. pinnatum are summarized in

Box4.

Pharmacological activities

The studies that investigated the pharmacological properties of K. laciniata and B. pinnatum usually assessed the activities of the juice or extracts from the leaves and/or aerial parts of these plants.

Regarding the type of extract, most studies with K. laciniata used hydroethanolic, whereas for B. pinnatum the ethanolic (in vitro) and aqueous (in vivo) were the most commonly used extracts.

However, there is a large difference in the number of studies between the two species, being B. pinnatum the most studied one, therefore, following the trend that has already been observed in the chemical and ethnopharmacological studies, which had been previously discussed in this article.

Fig.4

presents an overview of the main pharmacological activities that have been studied (in vitro and in vivo) for the two species in addition to the number of studies that have been published so far for each plant.

It is clear from

Fig.4

that a larger number of studies have been reported for B. pinnatum, which suggests that there is more room for studies that investigate additional pharmacological activities for K.

laciniata, especially in vivo studies. Moreover, this ﬁgure indicates that the gastroprotective activity (alti-ulcer, for instance) is not well explored yet for these plants, especially for K. laciniata.

0 Other activities Action on immune system Action on nervous system Action on urinary system Gastroprotective action Anti-snake action Antidysiphydeemic action Antidiabetic action Antioxidant action Antimicrobial action

KP KB

10 20 30

Fig.4. GraphicalrepresentationofthenumberofpharmacologicalstudiesforKalan- choelaciniataandBryophyllumpinnatum.

It is worth to point out that no comparative study between these species has been carried out so far, especially those related to phar- macological aspects. There are only two studies,

Fernandesetal.

(2016)

and

Araújoetal.(2018),

which compare the anti-snake and gastroprotective activities, respectively. Thus, more studies that compare some pharmacological activities between K. laciniata and B. pinnatum would be interesting to see, especially those related to their traditional uses.

Kalanchoe laciniata

Several non-clinical assays related to the evaluation of phar-

macological activities of K. laciniata are described in the literature.

(9)

Box4:MedicinalpopularusesofBryophyllumpinnatumdescribedintheliterature.

Plantpart Popularuse Preparation Reference

Leaves Bruises,swellings Rawleafapplieddirectlytotheaffectedarea Arnasonetal.(1980)

Notreported Tonic – Mello(1980)

Leaves Otitis,sinusitis – SandbergandCronlund(1982)

Leaves Boils,sores Warmleafjuiceappliedovertheinjuredarea

twoorthreetimesaweek.Leavesareboundon woundstohastensuppuration

SebastianandBhandari(1984)

Notreported Dermatologicaldisorders – EsquivelandZolla(1986)

Leaves Leproussoresandmotordisorders.

Fever

Theleavesaredry-friedandusedinacomplex preparation.Thejuiceoftheleavesisrubbed ontheforeheadtoreducefever

ElliottandBrimacombe(1987)

Notreported Cancer – MathewandUnithan(1992)

Leaves Scabies,leucoderma Juiceanddecoction Bhandaryetal.(1995)

Leaves Hypertension Decoction LonguefosseandNossin(1996)

Leaves Constipation,fever Juice OngandNordiana(1999)

Notreported Inﬂammation,stomachpain Tea Mu ˜nozetal.(2000)

Leaves Generalpain,cough,bronchitis,ﬂu, pneumonia,dermatitis

– Begossetal.(2002)

Leaves Bronchitis,ulcers,chilblain

treatment,atsyrupform

Juicewithmilkandsmearleaves Moreiraetal.(2002) Wholeplant,leaf Rheumatoidarthritis,tummybug,

injuriesfromfalls,numbnessof limbs,bruise,burn,ulcer

Medicinebath,rubbingormassage LongandLi(2004)

Leaves Cough,bronchitis,chilblain,ulcer Syrup,juicewithmilk,smearleaves Medeirosetal.(2004)

Leaves Bladderproblems,“cooling”,

kidneyandotherurinaryproblems, bladderstones,hypertension,high cholesterol

– Lans(2006)

Leaves Abscess Theleavesarecrushedandthepasteislayered

ontheboils

Saikiaetal.(2006) Leaves Analgesic,psychotropic Decoction,squeezedsapafterhealing,boiled Banzouzietal.(2008)

Leaves Relieveabdominalandbackpain,

stoppostpartumabdominalpain, stopexcessivemenstrual hemorrhage,reducefever

Decoction,poultice Coe(2008)

Leaves Burn,wound Pounded

Linimentandpoultices

Intaetal.(2008)

Leaves Wounds,bruises,boils,jaundice,

snakebite,dysentery,urinary troubleandquickhealingof wounds

– SikdarandDutta(2008)

Leaves Gastritis,ulcer,kidney,drycough withchestpain,cough,erysipelas, anyswelling,cataract,ulcer

Mixture,syrup,cataplasm,mixtureor compress,distillate,“garrafada”

Coelho-Ferreira(2009)

Leaves Fever,headache,nausea,wound Directapplicationoftheleavesonthesite GiovanniniandHeinrich(2009)

Leaves Coughs,mucus,fever,suddenloss

ofconsciousness(epilepsy-like), constipation,piles

Juicesqueezedfromtheleavesistakentwo teaspoonfulstwicedaily.Pasteofleavesis appliedtorectum

Hossanetal.(2009)

Leaves Kidneystone Oneteaspoonfuljuicewastakentwiceadayup

to1month

KosalgeandFursule(2009)

Leaves Asthma Heatedonlowﬁre,sapsqueezed,mixedwith

melasse(juiceofSaccharumofﬁcinaleand juiceofCitrusaurantifoliaanddrunk

Ruysschaertetal.(2009)

Leaves Headache,pain,depurativeifhigh dose–emetic;malaire

Crushedandrubbedontheforehead;

squeezed,thejuiceobtainedisdrunk;slightly boiled,fordrinkingandbathing(forchildren)

Sanz-Bisetetal.(2009)

Leaves Lumbarpain Innatura–plaster Garciaetal.(2010)

Leaves Cancer,inﬂammation Infusion KambojandSaluja(2009)

Leaves Migraine,headache Juicewithcoconutorandirobaoil KambojandSaluja(2009)

Leaves Furuncle,skinulcers Heattheleavesandusethemtopically KambojandSaluja(2009)

Seed Styedisease Juiceobtainedfromcrushedseedsisappliedto

eyes(1–2drops,2–3timesdaily)

Rahmatullahetal.(2010)

Leaves Headache Crumpledleafapplication Boulogneetal.(2011)

Leaves Liverproblems Innature CostaandMayworm(2011)

Leaves Bonepain,inﬂammation,

muscular-skeletalsystemdisorders (brokenbone)

Crushedandcompressed Panyaphuetal.(2011)

Leaves Bonefracture,constipationand

kidneystoneproblem

Paste(topical)andwaterdecoctionwithsugar (oral)

Tangjangetal.(2011)

Leaves Antidiabetic Juiceofboiledleaves(100g)isprescribed

orallytwiceaday

Taraketal.(2011)

Leaves Cuts – Bhatetal.(2012)

Leaves Stomachulcer Rawleaveseatendailyinemptystomach BoscoandArumugam(2012)

Leaves,roots Hypertension Decoction,infusion,mixture Gbolade(2012)

Leaves Wart.Skindiseases,wounds,

scabies,stopbleeding

Prepareapoulticeandapplyonwart NunkooandMahomoodally (2012)

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Box4:Continued

Leaves ScorpionstingVenomousinsect

bite,rheumatism,stiffjoints,kidney stones

Paste(external) PrabuandKumuthakalavalli

(2012),Bahekaretal.(2012)

Leaves TreatingamenorrheaTreating

morningsickness

Hotinfusion Srithietal.(2012)

Leaves Sexualtransmittedinfections

(gonorrhea)

Amixtureofchoppedleaves,choppedOpuntia strictastemandEuphorbiahypericifolia(whole plant)allboiledin2lofwaterandadministered onceadayasanenema

Wetetal.(2012)

– Gastrointestinaldisorders Thedoseis45–180grainsmixedwithtwiceits

amountofmeltedbutter

Kadiretal.(2013)

Leaves Removekidneystone,bladder,

pancreatic,emollient

Extract,paste Mahmoodetal.(2013)

Aerialherbalmaterial Guineaworm Poulticeasapasteanddecoction Agyareetal.(2014)

Leaves Eczema,pruritus – Bhatetal.(2014)

Leaves,ﬂower Diabetes,hypertension,analgesic, inﬂammation,woundulcers, anti-parasitic,insectbites, anti-cancer,cough,diarrhea, sedative,diuretic,anti-microbial, convulsions

Juiceextract EzuruikeandPrieto(2014)

Leaves,root Cholera,diarrhea,anddysentery, ulcer,urinarydiseases, gastrointestinaldisorders

Juice,decoction

Leafjuiceanddecoctionofroot

Islametal.(2014)

Leaves Kidneystone 1cup(50ml)freshjuicethreetimesadayfor

5–10daysortillremovalofstones

Choudhuryetal.(2014) Leaves Coughing,whoopingcough 2–3spoonfulofleafpasteistakenasdrink2–3

timesdailyfor3weeks

Kadiretal.(2014)

Leaves Paininthefeet,diabetic

neuropathy(diabetesandrelated complications)

Prepareadecoctionwiththeleavesanduseit asafootbath

Mootoosamyand Mahomoodally(2014)

Leaves Snakebite 1–2spoonsofleafdecoctionaregivenevery1h

aftersnakebite

Félix-Silvaetal.(2017),Sarkhel (2014),SikdarandDutta(2008)

Leaves Legspain,bodyache,

antimicrobial,anti-ulcer, antinociceptive,anti-inﬂammatory, antidiabetic,neurosedativeand musclerelaxant,hepatoprotective, jointpain(rheumatism)

Decoction

Heatedoilisappliedevenlyonleafwhichis carefullytiedtopainsite.Bathwithdecoction ofleavesforlegspainandbodyache

Sreekeesoonand Mahomoodally(2014)

Leaves Stonedisease,kidneystone – AgarwalandVarma(2015)

Leaves Allergy,dysentery Crushedextract Choudhuryetal.(2015a)

Leaves Digestivesystemdisorder Extractfromthecrushedleaves Choudhuryetal.(2015b)

Leaves Malaria,fever Leavesjuice(orallyadministered)formalaria

andfever

Frausinetal.(2015)

Leaves Cold Decoction,juice Pickingetal.(2015)

Wholeplant Diabetes Freshjuice Tarafdaretal.(2015)

Leaves Menstrualpain,kidneystone,

wormsinstomach

– Xavieretal.(2015)

Stem,leaves Antitumoractivity – Teneetal.(2007)apud

Bailon-Moscosoetal.(2015)

Leaves Sprain Leavesdecoctedorgrilledonﬁre.Appliedthe

leavesontheaffectedlimbforfewhours

Chassagneetal.(2016)

Several non-clinical studies have investigated the numerous phar- macological activities of K. laciniata. Summaries of the non-clinical in vitro and in vivo studies are presented in

Boxes5and6

, respec- tively.

Regarding the non-clinical in vivo studies, anti-inﬂammatory and immunomodulatory have been the most frequently investi- gated activities so far. This fact is probably due to the popular use of K. laciniata for treating inﬂammatory disorders. Besides these activities, K. laciniata have been investigated to treat against snake bites by Bothrops species, B. jararaca and B. arternus, where it showed great potential to ameliorate the local effects induced by the snake venom, especially the hemorrhagic reaction. Finally, although often recommended by folk medicine practitioners to use K. laciniata for gastroprotection, there is only one article that inves- tigated its potential use to treat gastric ulcer. Therefore, there are so many pharmacological activities that still lack investigation on this species.

Bryophyllum pinnatum

Several non-clinical in vitro and in vivo studies have been reported for B. pinnatum as summarized in

Boxes7and8

, respec- tively. In contrast to the K. laciniata species where most studies investigated its anti-inﬂammatory activity, the studies performed for B. pinnatum evaluated its leishmanicidal, anti-diabetic, anti- inﬂammatory, immunomodulatory and anti-cancer properties.

B. pinnatum is used in folk medicine to treat various diseases. For

this reason, several in vitro studies were carried out in order to ver-

ify the pharmacological properties of those species, which includes

hepatoprotective, leishmanicide, immunomodulatory, antimicro-

bial, antioxidant, anticancer, and antiurolithiatic activities. Studies

have shown the potential activity of B. pinnatum against hemato-

logical parasites such as Leishmania, Plasmodium and Trypanossoma,

whose properties are important due to the very limited pharma-

cological alternatives for treating these neglected diseases, where

researches in this area are of outmost importance.

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Box5:Non-clinicalinvitrostudiesperformedforKalanchoelaciniata.

Plantpart Extract/fraction/compound Method Result Reference

Reversibleandtime-dependentinhibitoryeffectsonCYP2C19andCYP3A4activities Wholeherb Methanolextract Crudemethanolextractand

fractionsofK.crenatawere incubatedand

preincubatedwith recombinanthuman CYP2C19andCYP3A4.

Asigniﬁcanttime-dependentinhibitionof testedsamplesonCYP3A4withcrude methanolandfractionswereobserved.

Awortweetal.

(2015)

Leishmanicidalactivity

Aerialparts Mixtureofsolvents consistingofmethanoland methylenechloride(v/v)

Antileishmanialactivity againstpromastigotes formsofL.donavani–cell viability.

TheK.crenatashowednoactivityagainst promastigotesformsofL.donavani.

Hubertetal.(2013)

Antibacterialactivity Aerialparts (leavesandstems), essentialoil

Hydroalcoholicextract (leaves),thealcohol (stems),essentialoil (leaves)

Diffusioninsolidmedium methodtodeterminationof theminimuminhibitory concentration(MIC).

Onlytheessentialoilshowedactivity againstmethicilinresistantStaphylococcus aureus(MRSA).Thisactionwasconsidered bacteriostaticwiththereductiontoone log10CFU/mlafter6hofexhibitionatthe concentrations4and8%.

Silvaetal.(2009)

Antiviralactivity

Leaves Aqueousextract Antiviralactivityagainst

herpesvirusbovineHBV typeI.

Theextractshowedreasonableantiviral activity,withinhibitionindex10⁻^4.48and hightherapeuticindex(88.16%).

Schiavo(2005)

Acetylcholinesteraseinhibition

Leaves Hexane,methanol,ethyl acetateextract

Microplateassaywiththe enzyme

acetylcholinesterase (AchE).

Themethanolandethylacetateshowed 100%inhibitionattheconcentration 2mg/ml.TheethylacetatehadIC500.16mg.

Trevisanetal.

(2006)

Leaves Ethylacetate,methanol extracts

Microplateassaybasedon Ellman’smethodandTLC (thin-layer

chromatography)assay.

Inthemicroplateassay,theethylacetate andmethanolextractsshowed100%

inhibition.Theethylacetateextract presentedIC500.16mg/ml.IntheTLCassay, theethylacetateextractshowed

positive/false-positiveresultandmethanol extractshowednegativeresult.

Feitosaetal.(2011)

Thyroidperoxidaseinhibition

Leaves Aqueousextract TPO(thyroidperoxidase)of

tissuesamplesobtained duringthyroidectomy.

Atthedose2␮M,theextractcaused signiﬁcantinhibitionofTPO iodine-oxidationactivity.

Ferreiraetal.(2000)

Larvicideactivity

Leaves Hexane,methanol,ethyl acetateextract

Larvicideactivityagainst Aedesaegyptignatlarvae.

Significantlarvicideactivitytohexaneand ethylacetateextracts,butnottoisolated flavonoid(3-O-␣-LL-rhamnopiranosil- 3,3,4,5,7-pentahydroxi-8-metoxiflavon)at theconcentrationof500ppm.

Trevisanetal.

(2006)

Antihistaminicactivity

Leaves Juice(withoutsolvents), aqueous,butanolfractions

AssayswithGuineapig ileum.

Thejuiceandfractionschangedresponse curvetohistamine,displayingan antihistamineeffect.

Almeidaetal.

(1997)

Lymphoproliferativeactivity

Leaves Juice(withoutsolvents) andisolatedcompounds (Box1compound1–7)

Lymphoproliferative activityinhuman lymphocites.

Theconcentrationofﬂavonoids2 (0.25␮g/ml)and1(0.5␮g/ml)thatinhibited thelymphocyteproliferationwas80and40 times,respectively,greaterthanthejuice (20␮g/ml).

Costaetal.(1994)

Anti-inﬂammatoryactivity

Leaves Juice(withoutsolvents) Inﬂammationinducedby Zymozan.

Theinhibitionoflymphocytesproliferation wasdose-dependent,withtheinhibitory concentrationof50%at50␮g/ml.

Ibrahimetal.(2002)

Antioxidantactivity

Wholeplant Methanolextract TBARS(thiobarbituric acid-reactivesubstances), MDA(malondialdehyde) activity,SOD(superoxide dismutase)activityandCAT (catalase)activity.

Treatmentoftheratswiththeextractforsix weeksloweredMDAlevelby34–44%,but increasedCATlevelsby78–176%,andSOD levelsby116–257%.

Fondjoetal.(2012)

PhospholipaseA2activity

Leaves Hydroethanolicextract PhospholipaseA2activity wasdetermined turbidimetricallyin96-well microplatesusinganegg yolksuspension.

Theextractshowedsigniﬁcantinhibitory activityofPLA2.

Fernandesetal.

(2016)

(12)

Box6:Non-clinicalinvivostudiesperformedforKalanchoelaciniata.

Plantpart Extract/fraction/

compound

Doseandrout Method Animalmodel Result Reference

Anti-inﬂammatoryactivity

Leaves Aqueous1(before

ﬂowering)Aqueous 2(afterﬂowering)

0.25,0.5,1.0, 2.0g/kg,i.p.

Pawedema inducedby carrageenan

Wistarrats Allconcentrationsofextract1 inhibitedthepawedema4h aftercarrageenaninjection, whiletheextract2showedno anti-inﬂammatoryactivity.

Mourãoetal.

(1999)

Aerialparts Juiceandaproduct ofjuicepuriﬁcation (KMC)

480mg/kg(juice), i.p.;240mg/kg (kalanchosine dimalate–KMC–in water),i.p.

Pawedema inducedby zymosan

C57B110male mice

Thedose240mg/kg,i.p.,KMC signiﬁcantlyreducedtherats popliteallymphonodes increasing.Thedose 480mg/kg,i.p.,ofjuiceofK.

laciniataobtainedthesimilar result,indicatingthatKMCisat leasttwicemoreactivethenthe juice.

Costaetal.

(2006)

Leaves Methanolextract 300,600mg/kg, oral

Pawedema inducedby carrageenan

Wistarrats Thedose600mg/kgexhibited themaximum

anti-inﬂammatoryeffect (43.47%)in30min.

Dimoetal.

(2006)

Adjuvanttreatmenttopoisoning(localanti-inﬂammatoryactivity) Aerialparts Aqueousextract Extractandtopic formulation(3:7– extract:lanette cream)

Hemorrhagic activityinduced andbyBhotrops alternus

Adultmices Theextractandtheformulation showedsigniﬁcantresultsin reducingedema,hemorrhagic haloandnecrosisprevention inducedbyBothropsalternatus venom.

Fonsecaetal.

(2004)

Leaves Aqueousextract Preincubationof venomwiththe extract

Hemorrhagicassay inducedbyB.

jararaca

Swissmice Intheconcentration1:48the extractshowedasigniﬁcant reductioninhemorrhage(57%).

Mouraetal.

(2015)

Leaves Hydroethanolic

extract

125,250, 500mg/kg,i.p.

Pawedemaand hemorrhagic activityinducedby Bhotropsjararaca

Swissalbino mice

Inpre-treatmentprotocol,the extractreducedthe hemorrhagicactivityreaching about40%,andinthe post-treatmentprotocol,itdid notshowactivity.Inthe antiendematogenicactivity,K.

laciniatadidnotshowany antiendematogenicactivityin bothtreatmentprotocols.

Fernandesetal.

(2016)

Immunomodulatoryactivity

Leaves Aqueousextract 200␮l,oral;160, 320,480or 960mg/kg,i.p.

Pawedema inducedby zymosan

C57B110male mice

Theinﬂammationwasreduced after7daysoftreatment.

Ibrahimetal.

(2002) Aerialparts Productofjuice

puriﬁcation(w/o useofsolvents):

KMC

480mg/kgofjuice and160mg/kgof KMC(kalanchosine dimalate),i.p.

Administrationof juiceandKMCat themicewith posteriorremoval ofcellsforanalysis

C57BL/10and C57BL/10ScCr malemice

KMCpromotedselective inhibitionofB-cell lymphopoiesis,byinhibiting IL-7.

Paivaetal.

(2008)

Anti-dyslipidemicpotential

Wholeplant Methanolicextract 50,68mg/kg,oral Diabetesand nephropathywere inducedintherats withstreptozotocin

MaleWistar rats

Aftersixweeksoftreatment, thedosesreducedtheblood leveloftheglucose,the triacylglycerides,thetotal cholesterolandtheLDL cholesterol.TheHDLlevelwas enhancedandhencedecreased theatherogenicindex.The extractreducedtheglucosuria andproteinuria.

Fondjoetal.

(2012)

Antihyperglycaemicpotential Wholeplant Hydroethanolic

extract

135,200mg/kg,oral Diabeteswas inducedby hypercaloric sucrosedietover4 months

Adultmale Wistarrats

Bothdosesreducedtheblood glucoselevelsinnormaland diabeticratswithoutreal dose-dependanteffect,6hafter asingleoraladministration.

Kamgangetal.

(2008)

Analgesicactivity

Leaves Methylene

chloride/methanol (1:1)extract

150,300mg/kg,oral Writhingtest, Formalintestand Analgesymetertest

AdultSwiss albinomice andadult Wistarrats

Theextractsigniﬁcantly reducedthewrithingreaction inducedbyaceticacid,reduced thelickingtimeattheﬁrst phaseofobservation,inthe secondphase,animalstreated showednosignofpainand reducedtheanimal’ssensitivity topain.

Nguelefack etal.(2006)

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Box6:Continued

Plantpart Extract/fraction/compound Doseandrout Method Animalmodel Result Reference

Anticonvulsantactivity

Leaves Methylene

chloride/methanol (1:1)extract

150,300, 600mg/kg,oral

Pentylenetetrazol, Thiosemicarbazide andStrychnine sulphate-induced seizures

AdultSwiss albinomice andadult Wistarrats

Theextractsincreasedthe latencyperiodandreducedthe durationofseizuresinadose dependentmanner.Thedose 600mg/kgdelayedtheonsetof convulsionandreducedthe durationofconvulsion.

Cardiovasculareffects

Leaves n-Butanolextract 2,5,10mg/kg,via cannulaintheright femoralvein

Effectofextracton bloodpressure, heartrate,ECG

Guineapigs Theslowadministrationofthe n-butanolextractresultedina signiﬁcanttransientfallin bloodpressurethatlastedfor 4min,inducedadose dependentfallinheartrateand amodiﬁcationofthe electrocardiogramcurve.

Antitumoractivity

Leaves Hydroethanolic

extract

62.5,250mg/kg,i.p. Implantinmice tumorcells:Ehrlich carcinomaand sarcoma-180

Femalealbino Swissmice

Bothdosesshowedanupper inhibitionto50%for sarcoma-180.ForEhrlich Carcinoma,thehighest inhibitionwas66.59%forthe higherdose.

Silva(2007)

Leaves Aqueousextract 50mg/kg,i.p. Antitumoractivity inmiceinfected withsarcoma-180

Femalealbino Swissmice

Theextractshowedinhibitory effectoftumorgrowingagainst sarcoma-180,withtumormass reduction52.8%.

Machadoand Melo-Júnior (2009)

Gastroprotective

Leaves Aqueousextract 125,250,

500mg/kg,oral

Gastricslesions inducedbyethanol andindomethacin

FemaleWistar rats

Thepre-treatmentprotectsthe mucosaofratsagainstthe gastricdamageand signiﬁcantlyreduceddamage byimprovingparameters relatedtooxidativestressand inﬂammationonmucosal structures.

Araújoetal.

(2018)

The potential use of B. pinnatum against Leishmania amazonensis has been investigated in vitro (Muzitano

etal.,2006a,b)

and in vivo (Muzitano

etal.,2009),

where the authors demonstrated such activ- ity with the leaves extract (320 mg/kg body weight), as well as with the isolated ﬂavonoids, 3-O-

␣

-

l

-arabinopyranosyl (1

→

2)-

␣

-

l

-rhamnopyranoside and quercetrin (16 mg/kg body weight), with both being able to signiﬁcantly reduce parasite load (Muzitano

etal.,2009).

In the same article,

Muzitanoetal.(2009)

studied the oral metabolism of flavonoids from B. pinnatum in a murine model of cutaneous leishmaniasis. In addition, they performed another study investigating the influence of cultivation conditions, season of collection and extraction method on the content of antileishma- nial flavonoids, where they demonstrated that active flavonoids were more abundant when the leaves were collected during the summer season and after aqueous extraction at 50

^◦

C. Regarding the in vivo studies, the most investigated activ- ities are leishmanicidal, hepatoprotective, immunoprotective, anti-inﬂammatory, anti-ulcer, antihypertensive, antinociceptive, wound healing, anti-asthmatic, antitussive, antidiabetic and anti- convulsant. Although these studies have demonstrated the several pharmacological activities attributed to B. pinnatum, it seems that its full potential is far from being proved and additional studies are needed in order to fully investigate its pharmacological properties, its mechanisms of action and its pharmacokinetics.

Another interesting area of research is the anti snake-bite activ- ity. Snake-related accidents are a serious public health problem and have been included on the WHO’s List of Neglected Tropical Dis- eases since 2009 (Gutiérrez

etal.,2013).Fernandesetal.(2016)

investigated the activity of B. pinnatum against the local effects induced the venom of B. jararaca, where they showed that the

extract of this plant was able to antagonize the hemorrhage and edema as well as to inhibit the phospholipase A2 from the venom.

The extract of B. pinnatum was active in pre- and post-treatment protocols, indicating the potential antiophidic activity of Kalanchoe species against local effects induced by B. jararaca snake, suggesting their potential use as a new source of bioactive molecules against bothropic venom.

Unlike K. laciniata, the antiulcer activity of B. pinnatum has been studied extensively in recent years. The work published by

Araújoetal.(2018)

showed that the pre-treatment with B. pinna- tum juice protects the mucosa of rats against the gastric damage of indomethacin and ethanol-induced gastric lesions and reduced damage by improving parameters related to oxidative stress and inﬂammation on mucosal structures. However, additional stud- ies are necessary in order to investigate what active components are responsible for such activity and the mechanisms of action involved. In addition, further clinical studies are necessary to prove the gastroprotective activity of B. pinnatum in humans.

In this current literature review, only one clinical study was

found that evaluated the efﬁcacy and safety of capsules contain-

ing B. pinnatum extract in the treatment of overactive bladder

syndrome. The clinical study of phase II, prospective, multicenter,

double-blind randomized, placebo-controlled was conducted with

twenty female patients and suggests that this species might have

potential use in the treatment of overactive bladder (Betschart

etal., 2013).

In addiction to this article, the same research group investi-

gated the effects of the leaves juice, fractions enriched in ﬂavonoids

and bufadienolides as well as a ﬂavonoid aglycone mixture and

individual aglycones on detrusor contractility as a major target in

overactive bladder treatment, where the authors found that several

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Box7:Non-clinicalinvitrostudiesperformedforBryophyllumpinnatum.

Anti-helminticactivity

Aerialherbalmaterial Hydroethanolic(50:50, v/v)extract

Anti-helminticactivity againstthefree-living modelnematode Caenorhabditiselegans

NoactivitywasobservedfortheextractofB.pinnatum. Agyareetal.(2014)

Antibacterialandantifungalactivities

Leaves Methanolicextract

(60%)

Agar-welldiffusion method

Fiveofthetestedbacteria(Bacillussubtilis,Escherichia coli,Proteusvulgaris,ShigelladysenteriaeandS.

aureus)weresensitivetoextractat25mg/ml.K.

pneumoniae,P.aeruginosaandC.albicanswere resistanttotheextract.

Akinpelu(2000)

Leaves Ethanolicextract(70%) Agar-welldiffusion method

Broad-spectrumantimicrobialactivitywasdetectedin crudeextracts,beingactiveagainstGram-positiveand Gram-negativebacteria,yeastandﬁlamentousfungi (C.albicans,Rhizoctoniabataticola,A.nigerand Alternariaalternate).

AqilandAhmad(2003)

Leaves Aqueous,methanolic

extracts,extractwith Palmwineandwithgin andwithbrewcorn

Agar-welldiffusion, MICandminimum bactericidal concentration(MBC)

Themethanolicextractwasmostactive,inhibitingS.

aureus,Enterococcusfaecalis,B.subtiliseP.

aeruginosa.Theotherextractsshowedmoderate activity.

Akinsulireetal.(2007)

Leaves Alcoholic(90%),

aqueousextract

CupPlateMethod (ZoneofInhibition)

Themostactiveextractwastheaqueousonefollowed bythealcoholicextract.Thesusceptibilitydecreasing orderfortheaqueousextract:Enterobacter aerogenes>E.coli>Shigelladysenteriae>Salmonella entericavar.Typhi>S.aureus>B.

subtilis>Staphylococcusepidermidis>Micrococcus luteus.Andfortheethanolicextract:S.aureus>S.

entericavar.Typhi>S.dysenteriae>E.aerogenes>E.

coli>B.subtilis.

Jainetal.(2010)

Stem Methanolic,aqueous

extracts

Agar-diffusionmethod Themethanolextracthadsigniﬁcantantibacterial actionagainstB.subtilisandS.aureusat100,50and 25mg/ml.Also,theaqueousextractpresented antibacterialeffectagainstS.entericavar.TyphiandB.

subtilisatthesameconcentrations.

Nwadinigwe(2011)

Leaves,stems Petroleumether, aqueousextracts

Agar-discdiffusion method

Bothextractsshowedmoderateactivityagainstall testedfungalstrains(A.niger,Blastomyces dermatitides,C.albicans,Pityrosporumovale, Trichophytonspp.andMicrosporumspp.).The petroleumetherextractwasmoreeffectiveagainst Microsporumspp.,whereasC.albicanswasmore susceptibletoaqueousextract.

Chowdhuryetal.(2011)

Leaves Chloroformextract Agar-discdiffusion method

Theextractexhibitedlowlevelofantibacterialactivity againstB.subtilis,B.megaterium,S.aureus,E.coli,P.

aureginosa,S.entericavar.TyphiandS.dysenteriae.

ThehighestinhibitionwasagainstE.coli,whileno antibacterialactivitywasfoundagainstV.cholerae.

Biswasetal.(2012)

Wholeplant Methanolicextract Disk-diffusionmethod Themethanolexhibitedweakantimicrobialeffect againstB.cereus,Bacillusmegaterium,B.subtilis, Sarcinalutea,S.aureus,E.coli,Salmonellaentérica var.Typhi,Salmonellaparatyphi,Shigellaboydii,S.

dysenteriae,P.aeruginosa,Vibriomimicus,Vibrio parahaemolyticus,Aspergillusniger,C.albicansand Sacharomycescerevisiae.

Sharkeretal.(2012)

Wholeplant Methanolicextract MIC Theextractpresentedantibacterialandantifungal activitieswithMICvaluesrangingfrom32to512␮g/ml.

ThemicroorganismstestedwereS.aureus,P.

aeruginosa,Salmonellaentericavar.Typhi,C.albicans, CandidaparapsilosisandCryptococcusneoformans.

Tatsimoetal.(2012)

Root,stem,leafand wholeplant

Methanolic,aqueous extracts

MIC,agar-welldiffusion method

Thestemmethanolicextractshowedhigher antibacterialactivitybeingeffectiveagainst Corynebacteriumdiphtheriae,Micrococcusluteus,B.

subtilis,Alcaligenesfaecalis,Bordetellabronchiseptica andSerratiamarcescens.Aqueousextractofleafwas onlyactiveagainstB.subtilisandA.faecalis,while methanolicextracthadnoactivity.

Sharmaetal.(2014)

Leaves Methanolicandethyl

acetateextracts

Anti-Helicobacter activityfor determinationofMIC andMBC

Methanolextractshowedasigniﬁcantanti-Helicobacter activitywithMICandMBCvaluesof32and256␮g/ml, respectively

Mabekuetal.(2017)

Antitrypanosomalactivity

Leaves Aqueousextract Microtiterplate

method

Noobservablereductioninmotilityat10mg/ml, motilityreducedslightlyat20mg/mlandhighly reducedmotilityat40mg/ml.

Alhajietal.(2014)

(15)

Box7:Continued

Anti-hepatitisCvirusactivity

Leaves Methanolicextract HUH7it-1cells(human hepatocellular carcinoma)were infectedwiththeHCV genotype2astrain JFH1inthepresenceof crudemethanol extracts

Thecrudemethanolextractexhibitedanti-HCVactivity withaIC5017.2␮g/ml.

Aokietal.(2014)

Antiplasmodialactivity

Leaves Ethanolicextract SYBRgreenI-based

ﬂuorescenceassay

Goodantiplasmodialactivity(IC5011–20␮g/ml)was observedinleafethanolextract.

Singhetal.(2015)

Leishmanicideactivity

Leaves Aqueousextract,

isolatedﬂavonoid

Antileishmaniaactivity againstLeishmania amazonensis (antiamastigoteand antipromastigote)

Theaqueousextractandtheﬂavonoidisolatedshowed activityagainstLeishmaniaamazonensis.

Muzitanoetal.(2006a)

Antioxidantactivity

Leaves Aqueousextract DPPHandoxideradical

scavenging,reducing power,antilipidic peroxidation

Theextracthadsigniﬁcantantioxidantandoxidative radicalscavengingactivities.

Harlalkaetal.(2007)

Leaves Methanolicextract DPPHradical

scavenging

Adosedependentradicalscavengingactivitywas observedwiththemaximumactivity(63.97%)atthe highestdosecomparabletothestandardBHT.

Guptaetal.(2009)

Leaves Alcoholic,aqueous

extract

DPPHscavenging method

Theaqueousextractpresentedmoreantioxidant activitythenthealcoholicone.Aqueousandalcoholic extractsandascorbicacidexhibited74.7,58.4and 88.6%inhibition,respectively,andtheEC50(␮g/ml) –144.23,117.42and96.15␮g/ml,respectively.

Jainetal.(2010)

Roots Aqueous,ether,

chloroformic, methanolicextracts

DPPHandoxideradical scavengingmethods, reducingpower

Themethanolicextracthadmoreeffectiveantioxidant activitythentheotherextracts.

Majazetal.(2011)

Wholeplant Methanolicextract Methodof Brand-Williams

Asigniﬁcantantioxidantactivitywasveriﬁed. Sharkeretal.(2012) Wholeplant Methanolicextract DPPHradical

scavengingmethod

Themethanolicextractshowedantioxidantactivity withanIC50of52.48␮g/ml.

Tatsimoetal.(2012)

Leaves Aqueousextracts DPPHradical

scavengingmethod

Supplementalbluelightimprovedtheantioxidant activity.

Nascimentoetal.(2015)

Leaves Aqueousextract Thiobarbituricacid

(TBA)test,reducing power,DPPHand hydrogenperoxide

Whencomparedwithascorbicacid(standard),the extractshowedlowantioxidantactivity.

Alhajietal.(2014)

Root,stem,leaf, wholeplant

Methanolic,aqueous extract

Hydroxyland superoxideradical scavenging,ferrous chelatingactivities

Aqueousandmethanolicextractsofroot,stem,leaf andwholeplantshowedantioxidantactivity.

Sharmaetal.(2014)

Leaves Methanolicextract Phosphomolybdenum

assay,ferricreducing ability,hydroxylradical scavenging,

thiobarbituricacid reactivesubstance assay,metal ion-chelatingcapacity

Theextractexhibitedsigniﬁcantantioxidantandfree radical-scavengingactivity.

PhatakandHendre (2015a)

Leaves Aqueous,methanolic

andmethanolic50%

extracts

DPPH,

␤-carotene-linoleate method,metal(ferrous ion)chelatingmethod, lipidperoxidation inhibition;electron paramagnetic resonance

spectroscopy,inhibition ofDNAbreakage activity,browning potential,tyrosinase inhibitionassay

Themethanolic50%extractwasthemostactive.The studyindicatedthattheextractscanbeapotent antioxidant.Andtheywereactiveagainstthe tyrosinase.

Guptaetal.(2015)

Leaves Methanolicextract DPPHscavenging,

reducingpower,and hydroxylradical scavengingassay

DPPHradical,hydroxylradicalandreducingpower assaysshowedIC50valuesof25.31±0.34,55.94±0.68 and11.18±0.74␮g/ml,respectively.

Mabekuetal.(2017)

Leaves Hydroethanolic70% DPPHscavenging

method

Theextractpresentedantioxidantactivity. Haraetal.(2018)

(16)

Box7:Continued

Immunomodulatoryactivity

Leaves Aqueousextract Mastcelldegranulation

inmesenteryand histaminereleaseassay

Theextractpreventedthemastcelldegranulation antigen-induced,at100␮g/mlfor30min,andthe histaminereleaseat0.5mg/mlfor1h.

Cruzetal.(2008)

Leaves Aqueousextract,

isolatedcompounds

EffectofKp,ﬂavonoid quercetin(QE), quercitrin(QI)onmast cellactivation

TreatmentswithKpandQEinhibiteddegranulationand cytokineproductionofbonemarrow-derivedmastcells followingIgE/Fc␧RIcrosslinking.TreatmentwithKp signiﬁcantlyreducedlevelsofTNFinsupernatant.

Cruzetal.(2012)

Antiproliferativeactivity

Leaves Ethanolicextract,acid fraction

Evaluationin lymphocytesofmice BALB/cinguinal lynphonodesand humanperipheral blood

Theacidfractionwas20timesmoreeffectivethenthe ethanolicextractininhibitinglymphocyteproliferation.

Almeidaetal.(2000)

Thombolyticactivity

Wholeplant Methanolicextract MethodofPrasad Theextractpresentedmoderatethrombolyticactivities (16.41%).

Sharkeretal.(2012)

Treatmentoftheoveractivebladdersyndrome

Leaves Juice Stripsofporcine

detrusorwereprepared inKrebssolutionand contractilitywas measured

Thejuiceextractinhibitedcontractionsinducedby electricalﬁeldstimulationandrelaxes

carbachol-inducedcontractions.

Schuleretal.(2012)

Leaves Methanolicextract Detrusormusclestrips werepreparedfrom porcinebladdersand, theelectricallyinduced musclecontractility measured

At10%concentration,theextractreducedthe contractilityofthedetrusorto58.6±13.3%after74min, afteradose-independentinitialincreaseincontractility duringtheﬁrst40min.

Füreretal.(2015)

Leaves Juice Detrusormusclestrips

werepreparedfrom porcineurinary bladders

Thepretreatmentincreasedcontractionstrengthof porcinedetrusorstripsrelativetothenegativecontrol.

Bachamannetal.(2017)

Antiurolithicactivity

– Ethanolicextract Crystallizationof

calciumoxalatecrystals

Theextracthasantiurolithicactivityandhastheability inreducedthesizeofcrystals.

YasirandWaqar(2011)

Leaves Aqueousextract Nucleationassay

(turbiditymethod), aggregationassay

Theextracthadgreatercapacitytodissolvecalcium oxalate.Theextractinhibitedthecrystallization.The extractwasslightlybetterincomparisontoCystone (standard)ininhibitingtheformationofCODcrystals.

PhatakandHendre (2015b)

Tocolysisactivity

Leaves Aqueousextract Contractilitywas

measuredinstripsof termmyometrium exposedtoincreasing concentrationsofB.

pinnatum

Inhibitionofspontaneouscontractionwas concentration-dependent.Theextractincreased contractionfrequencyby91%andinhibited oxytocin-stimulatedcontractionsby20%withslightly decreasedfrequency.

Gwehenbergeretal.

(2004)

– Juice Stimulationbyoxytocin

activity

Thejuicepreventedtheoxytocin-inducedincreasein [Ca²⁺]iinhumanmyometrialcellsinadose-dependent manner,reachingaca.80%inhibitionata2%

concentration.

Simões-Wüstetal.

(2010)

Leaves Leafpressjuice(BPJ) Repeatedadditionof BPJinseveraldilutions (undiluted,1–10%)on myometriumstrips

TheBPJdecreasedamplitudeandinhibited contractilitysigniﬁcantlyfasterandincreased frequencysigniﬁcantlyfasterthenthecontrol.

Wächteretal.(2011)

Anticanceractivity

Leaves Bufadienolidesof

methanolicextract

Inhibitoryeffecton earlyantigenof Epstein–Barrvirusin theinductionRajucell activationbyatumor promotinggene

Allbufadienolidesshowedinhibitoryactivityand brioﬁlinAexhibitedthehighestactivity(IC50=0.4␮M) amongthecompounds.Thus,bufadienolidesare potentialcancerchemopreventiveagents.

Supratmanetal.(2001)

Leaves Chloroformextract MTT,electrophoretic mobilityshift,northern blottingandassaysin cervicalcancercells

Theextractinhibitedcervicalcancercellgrowthby 30%.Resultsshowndepictadose-dependentdecrease inthelevelofHPV18transcriptsincellstreatedwith crudeextract.

Mahataetal.(2012)

Antimutagenicactivity

Leaves Ethylacetate,

methanol,petroleum etherextracts

Antimutagenicactivity againstEMS(ethyl methanosulfonate)- inducedreversion mutationsinS.

typhimurium

Theethylacetateandpetroleumetherextracts exhibitedpotentantimutagenicactivitiesatthe non-toxicconcentrationsof200and400␮g/plate.

Obaseiki-Eboretal.

(1993)

(17)

Box7:Continued

Leaves Juice Salmonella/mammalian

microsomeassay (Amestest)

Atadose0.25mg/ml,thejuicecausedastatistically signiﬁcantreductioninthemutagenicityof2AA (p≤0.05)andthepercentofinhibitionwasmorethen 90%athigherdosesinallconditionstested.

Umbuzeiro-Valentetal.

(1999)

Hepatoprotectiveactivity

Leaves Juice,ethanolicextract Isolationofhepatocytes andexaminationofthe effectoftoxicantsalong withthetestsamples

Theconcentrateandethanolicextractsigniﬁcantly decreasedtheGOT(glutamyloxalaceticacid transaminase)levelby55.55and36.50%andGPT (glutamylpyruvatetransaminase)levelby69.57and 38.61%,respectively.

YadavandDixit(2003)

Acetylcholinesteraseactivity

Leaves Ethylacetate,methanol extracts

Microplateassayto AChEinhibitoryactivity andpositiveandfalse positiveactivityinTLC

Theethylacetateextractshowedastronginhibitionof acetylcholinesteraseactivity.

Feitosaetal.(2011)

Antidiabeticactivity

Stalk Aqueous,ethanolic

extracts

␣-Amylaseinhibition Theethanolextractshowedsigniﬁcantinhibitory activityof␣-amylaseenzymeinrelationtotheaqueous extract.

Matthewetal.(2013a)

PhospholipaseA2activity

Leaves Hydroethanolic50%

extract

PhospholipaseA2

activitywasdetermined turbidimetricallyin 96-wellmicroplates usinganeggyolk suspension

Theextractshowedsigniﬁcantinhibitoryactivityof PLA2.

Fernandesetal.(2016)

Box8:Non-clinicalinvivostudiesperformedforBryophyllumpinnatum.

Plantpart Extract/fraction/

compound

Doseandroute Method Animalmodel Result Reference

Gastroprotectiveactivity

Leaves Methanolicextract 100,300mg/kg,i.p. Ulcerinducedby indomethacin, serotonin,reserpine, aceticacidethanoland stress;Lesiongastricin pylorus-ligatedinduced byacetylsalicylicacid, Duodenalulcers inducedbyhistamine

Charles-Fosterrats, albinoguineapigs

Theextractexhibiteda signiﬁcantinhibitoryeffecton aspirin-inducedulcers.

Pretreatmentat300mg/kg inhibitedtheformationof indomethacin-inducedgastric ulcers.Therewasasigniﬁcant inhibitoryeffectonulcer formationbyserotoninand reserpine.Theextractprotected againstofulcersbystress.The extractreducedtheseverityof ulcersandcausedasigniﬁcant reductionofulcerindexinthe ethanol-inducedulcers.

PalandChaudhuri (1991)

Leaves Methanolicextract 10,20,40mg/kgof aqueousextract

Indomethacininduced gastriculceration

Adultmalealbino Wistarrats

Theextracthada dose-dependent gastro-protectiveeffecton indomethacininduced ulceration.Withresults,the extractcouldprobablybemore potentthenpropranololinthe measuredvariables.

Adesanwoetal.

(2007)

Leaves Aqueousextract 1and2g/kg,oral Gastriclesioninduced byindomethacin

MaleWistarrats Theaqueousextractshowed signiﬁcantanti-ulcerogenic effectwhencomparedwiththe negativestandard.The ranitidineandaqueousextract at1and2g/kgreducedthe ulcerationin45.49%,49.51%, respectively.

Brazetal.(2013)

Wholeplant Aqueousextract andmucilage

500and750mg/kg Ulcerinducedby ethanol

FemaleWistarrats Theextractatdoseof 750mg/kgp.o.andmucilageat doseof500mg/kgp.o.

markedlydecreasethe incidenceofulcersinrats.

Therewasadecreaseinthe gastricvolume,freeandtotal acidityandulcerativeindex was72.69forextractand 69.65%formucilage.

Sharmaetal.(2014)