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Bufadienolides of Kalanchoe species: an overview of chemical structure, biological activity and prospects for pharmacological use


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Toad venom is regarded as the main source of bufadienolides; however, synthesis of these substances takes also place in a variety of other animal and plant organisms, including ethnomedicinal plants of the Kalanchoe genus. Chemically, bufadienolides are a group of polyhydroxy C-24 steroids and their glycosides, containing a six-membered lactone (α-pyrone) ring at the C-17β position. From the pharmacological point of view, bufadienolides might be a promising group of steroid hormones with cardioactive properties and anticancer activity. Most of the literature concerns bufadienolides of animal origin; however, the medicinal use of these compounds remains limited by their narrow therapeutic index and the risk of development of cardiotoxic effects. On the other hand, plants such as Kalanchoe are also a source of bufadienolides. Kalanchoe pinnata (life plant, air plant, cathedral bells), Kalanchoe daigremontiana (mother of thousands) and other Kalanchoe species are valuable herbs in traditional medicine of Asia and Africa. The present review focuses on the available data on chemical structures of 31 compounds, biological properties and prospects for therapeutic use of bufadienolides from Kalanchoe species. Furthermore, it presents some new investigational trends in research on curative uses of these substances.
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Bufadienolides of Kalanchoe species: an overview
of chemical structure, biological activity and prospects
for pharmacological use
Joanna Kolodziejczyk-Czepas .Anna Stochmal
Received: 15 January 2017 / Accepted: 26 July 2017 / Published online: 2 August 2017
ÓThe Author(s) 2017. This article is an open access publication
Abstract Toad venom is regarded as the main
source of bufadienolides; however, synthesis of these
substances takes also place in a variety of other animal
and plant organisms, including ethnomedicinal plants
of the Kalanchoe genus. Chemically, bufadienolides
are a group of polyhydroxy C-24 steroids and their
glycosides, containing a six-membered lactone (a-
pyrone) ring at the C-17bposition. From the pharma-
cological point of view, bufadienolides might be a
promising group of steroid hormones with cardioac-
tive properties and anticancer activity. Most of the
literature concerns bufadienolides of animal origin;
however, the medicinal use of these compounds
remains limited by their narrow therapeutic index
and the risk of development of cardiotoxic effects. On
the other hand, plants such as Kalanchoe are also a
source of bufadienolides. Kalanchoe pinnata (life
plant, air plant, cathedral bells), Kalanchoe daigre-
montiana (mother of thousands) and other Kalanchoe
species are valuable herbs in traditional medicine of
Asia and Africa. The present review focuses on the
available data on chemical structures of 31 com-
pounds, biological properties and prospects for ther-
apeutic use of bufadienolides from Kalanchoe species.
Furthermore, it presents some new investigational
trends in research on curative uses of these substances.
Keywords Bufadienolide Kalanchoe
Cytotoxicity Cancer therapy Ethnomedicine
Bufadienolides are a group of polyhydroxy C-24
steroids and their glycosides. The first described
bufadienolide was scillaren A, identified in Egyptian
squill (Scilla maritima) (Stoll et al. 1933). The term
‘bufadienolides’’ originates from the genus Bufo
toads, which venom (a skin secretion) contains these
compounds. Both animals (toads, snakes) and plants
(Crassulaceae and Hyacinthaceae, in particular) syn-
thesize bufadienolides, while the bufadienolide
orthoesters were found only in several plant species:
Kalanchoe daigremontiana Raym.-Hamet & H. Per-
rier, Kalanchoe tubiflora (Harv.) Raym.-Hamet, the
hybrid Kalanchoe daigremontiana 9tubiflora,Kalan-
choe pinnata (Lam.) Pers., as well as in Melianthus
comosus Vahl and Bersama abyssinica Fresen
(Melianthaceae family).
The range of biological properties of bufadieno-
lides includes cytotoxic, antitumor and cardiotonic
activities (Gao et al. 2011), however, uncontrolled
J. Kolodziejczyk-Czepas (&)
Department of General Biochemistry, Faculty of Biology
and Environmental Protection, University of Lodz,
Pomorska 141/143, 90-236 Lodz, Poland
A. Stochmal
Department of Biochemistry, Institute of Soil Science and
Plant Cultivation, State Research Institute, Czartoryskich
8, 24-100 Pulawy, Poland
Phytochem Rev (2017) 16:1155–1171
DOI 10.1007/s11101-017-9525-1
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
administration of these substances may induce the
occurrence of side effects (Puschett et al. 2010). Most
of the literature concerning the chemical characteris-
tics, biological properties and possible therapeutic
effects of bufadienolides includes data derived from
studies on substances of animal origin. Chemistry and
biological activities of bufadienolides synthesized by
Kalanchoe plants are less known. Members of the
Kalanchoe genus (Crassulaceae) are native for sub-
tropical and tropical regions of Asia, Africa and
America as well as for Australia and Madagascar. In
Europe, K. pinnata and K. daigremontiana are mainly
grown as house ornamental plants; however, their
remedial properties are also known. Furthermore, both
these and other Kalanchoe species are popular
medicinal herbs in different regions of the world
(Table 1). Traditional recommendations for using
these plants include a wide range of diseases, includ-
ing gastric ulcers, kidney stones, rheumatoid arthritis,
bacterial and viral infections, skin diseases, cold as
well as other disorders (e.g. Fu
¨rer et al. 2016; Kawade
et al. 2014; Pattewar 2012; Rajsekhar et al. 2016).
Ethnomedicinal uses of Kalanchoe-derived prepara-
tions are mostly based on internal or external admin-
istration of crude extracts or plant juice. There is no
data on traditional uses of purified bufadienolides or
semi-purified bufadienolide-rich preparations. How-
ever, available findings suggest that therapeutic
activities (anti-cancer action, in particular) of Kalan-
choe-derived medicines may be partly dependent on
the presence of bufadienolides. Studies on these
compounds, originated from various sources, revealed
their anti-inflammatory, anti-cancer, anti-viral and
other beneficial activities (Kamboj et al. 2013).
Different research groups demonstrated anti-cancer
properties of bufadienolides synthetized by Kalanchoe
plants (e.g. Deng et al. 2014; Huang et al. 2013;Wu
et al. 2006; Yamagishi et al. 1989). Daigremontianin
and bersaldegenin-1,3,5-orthoacetate are listed in
literature as sedative substances and natural adaman-
tane derivatives (‘‘trioxaadamantanes’’) that may
possess anti-influenza activity (Wanka et al. 2013).
Additionally, the analysis of existing ethnomedicinal
evidence (e.g. Botha 2016; Lans 2006;Su
¨sskind et al.
2012), followed by studies with contemporary (bio)-
chemical and other scientific methods, may provide
new data on safety or possible risk of using of
Kalanchoe bufadienolide-containing extracts and
preparations in humans.
This work reviews the available data on Kalanchoe
species as a source of bufadienolides. Chemistry,
biological activities and prospects for possibility of
therapeutic use of Kalanchoe plant-derived bufadieno-
lides have been presented. Some information on
possible side effects of bufadienolides have been also
included. The current review comprises data (to May,
2017) from journals recorded in international data-
bases (Medline/Pubmed, Scopus, ScienceDirect/Else-
vier, Springer Link/ICM) and other scientific journals,
non-indexed in these databases.
Bufadienolide structures and their concentration
in Kalanchoe plants
The Kalanchoe species are succulent plants. Their
aerial parts were reported to contain not only steroid
compounds, but also some flavonoids, phenolic acids,
anthocyanins, alkaloids, saponins and tannins (El
Abdellaoui et al. 2010; Chowdhury et al. 2011). The
polyhydroxy C-24 structure of bufadienolides is based
on a six-membered lactone (a-pyrone) ring, located at
position C-17b. Some of these compounds have been
isolated from Kalanchoe plants, and their structures
have been established by spectral techniques. The
available structures of these compounds are presented
in the Figs. 1,2,3and 4. Supratman et al. (2000) have
identified two compounds including bryophyllin A (5)
and bryophyllin C (7) from K. pinnata, and in 2001
five next compounds: bersaldegenin-3-acetate (4),
bersaldegenin-1,3,5-orthoacetate (2), daigremontianin
(9), bersaldegenin-1-acetate (3) and methyl daigre-
monate (30) in study on bufadienolides of K. daigre-
montiana 9tubiflora (Fig. 1). Eight bufadienolides
were identified by Wu et al. (2006), in the extract of
aerial parts of K. gracilis, which included kalancho-
sides A (12), B (13) and C (14), thesiuside (31),
hellebrigenin (10), hellebrigenin-3-acetate (11), bryo-
phyllins A (5) and B (6). The systematic names of
these compounds are shown in the Table 2. From roots
of K. daigremontiana, eight new bufadienolides
named as kalandaigremoside A (15), B (16), C (17),
D(18), E (19), F (20), G (21) and H (22) were isolated
and characterized by Moniuszko-Szajwaj et al. (2016).
The presence of daigredorigenin-3-O-acetate (8)in
K. daigremontiana was reported as early as in the 80s
of the twentieth century (Wagner et al. 1985). Next
compounds were isolated in 2008 by Kuo et al. from
1156 Phytochem Rev (2017) 16:1155–1171
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Table 1 Kalanchoe species in ethnomedicine (a compilation of data)
Species (with English names) Traditional uses and geographical region and/or country
Kalanchoe crenata (Andrews) Haw. (Never-die) Medicinal plant, used during pregnancy by Anyi-Ndenye women (Eastern Ivory Coast, Africa)
(Malan and Neuba 2011)
Leaves are recommended to heal umbilical cord wounds in newborns (Mabira Central Forest
Reserve, Uganda) (Tugume et al. 2016)
Kalanchoe daigremontiana Raym.-Hamet & H. Perrier, syn. Bryophyllum
daigremontianum Raym.-Hamet & H. Perrier. (Mother of Thousands)
One of the most frequently prescribed anthroposophic medications, administered against
psychic agitation, restlessness, and anxiety—studies conducted at Hospital Havelhoehe,
Germany (Su
¨sskind et al. 2012)
Kalanchoe densiflora Rolfe For the treatment of wounds (Samburu of Mt. Nyiru, South Turkana, Kenya) (Bussmann 2006)
Kalanchoe germanae Raym.-Hamet ex Raadts (Air plant) Removal of ganglion—the pound leaves are used on ganglion area (Kenya) (Kipkore et al.
Kalanchoe glaucescens Britten Leaves are used to treat cough (Mabira Central Forest Reserve, Uganda) (Tugume et al. 2016)
Kalanchoe gracilis Hance, syn. Kalanchoe ceratophylla Haw. To cure injuries, pain, fever and inflammation (Taiwan) (Lai et al. 2010)
Kalanchoe laciniata L. (Christmastree plant) Juice from the leaves is used externally for joint pain (Southern India) (Karuppuswamy 2007)
Powdered leaves are administered to alleviate cough, to cure colds and inflammation and for
healing of boils and wounds (Southern India, Malaysia)
Headache (Philippines)
Crushed leaves are applied externally to decrease body temperature and to heal ulcers
(Cambodia, Laos, Vietnam)
To cure wounds, inflammation and diabetes (India) (Deb and Dash 2013)
Kalanchoe lanceolata (Forsk.) Pers. Anti-malarial remedy (Kenya) (Njoroge and Bussmann 2006)
The leaf juice is administered during dysentery (India) (Bapuji and Ratnam 2009)
Kalanchoe marmorata Bak. Boiled juice is used as eye drops for treatment of eye infections (eastern Ethiopia) (Belayneh
and Bussa 2014)
Kalanchoe petitiana A. Rich. Leaf juice is applied on the fractured for bone setting (Ethiopia) (Ragunathan and Abay 2009)
Phytochem Rev (2017) 16:1155–1171 1157
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Table 1 continued
Species (with English names) Traditional uses and geographical region and/or country
Kalanchoe pinnata (Lam.) Pers., syn. Bryophyllum pinnatum Lam., Bryophyllum
calycinum Salisb. (Life plant, air plant, love plant, Canterbury bells, Cathedral
In the treatment of urinary bladder stones (India, Trinidad and Tobago) (Lans 2006; Sen
et al. 2008)
Leaf extract is used to cure amoebic dysentery (North Bengal) (Mitra and Mukherjee
Wounds, bruises, swellings and insect bite (Himalaya) (Hussain and Hore 2007)
Diarrhea (India) (Dash and Padhy 2006)
Antibacterial and anti-inflammatory remedy (Vietnam) (Nguyen et al. 2004)
Internally: to cure acute and chronic bronchitis, pneumonia and others respiratory tract
infections, fever; externally: to treat dermatomycosis (Nigieria) (Okwu and Nnamdi
Leaves are recommended for treatment of cough in adults and children (Kibale National
Park, Uganda) (Namukobe et al. 2011)
Inflammation, dermatosis, skin problems, wound healing, arthritis, asthma, bruises,
diabetes, infections, tumours and ulcers—worlwide (Quazi Majaz et al. 2011a,b)
Paste from macerated leaves is used externally for muscle and joint pain (Bangaldesh)
(Tumpa et al. 2014)
Preparations from leaves are used to treat digestive disorders (India) (Barukial and
Sarmah 2011)
Decoction from leaves is administered to remove kidney stones (Bangladesh) (Afroz
et al. 2013)
Leaves are chewed with salt as a remedy for dissolving of gall bladder stones
(Bangladesh) (Rahmatullah et al. 2011)
Herbal preparation from roots and leaves is administered to women for recovering after
childbirth (West Java) (Sihotang 2011)
Leaf paste is applied externally to treat scorpion bite (India) (Vaidyanathan et al. 2013)
Leaf juice is recommended to treat cholera, diarrhea and dysentery (Bangladesh) (Khan
et al. 2015)
Leaves are used to treat urinary problems, incl. kidney and gall bladder stones
(Bangladesh) (Bhowmik et al. 2014)
Raw laves are chewed with sugar to treat dysentery and diarrhea; leaf juice is
recommended to cure jaundice; leaf paste is used externally to heal skin infections and
pimples (Bangladesh) (Das and Choudhury 2012)
Kalanchoe tubiflora Raym.-Hamet, syn. Bryophyllum delagoense (Eckl. & Zeyh.)
Druce (Chandelier plant)
One of the most common medicinal plants used for wound healing (Brazil) (Hsieh et al.
1158 Phytochem Rev (2017) 16:1155–1171
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15 H2HCH
16 H2HCH
Fig. 1 Structures of the
compounds 3,4,8,1022,
2629 and 31
Phytochem Rev (2017) 16:1155–1171 1159
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the extract of K. hybrid: kalanhybrin A (23), B (24)
and C (25). The only one new compound was isolated
from K. tomentosa and it was 3b-(40,60-dideoxy-b-
droxy-19-oxobufa-20,22-dienolide (1) (Rasoanaivo
et al. 1993). Few years ago, two bufadienolide
glycosides, i.e. kalantuboside A (26) and kalantu-
boside B (27) were found in the extract from the whole
plant of K. tubiflora (Huang et al. 2013). Furthermore,
it has been also established that K. lanceolata (Forssk.)
Pers. synthesizes 5-O-acetylhellebrigenin glycosides,
i.e. lanceotoxin A (5-O-acetylhellebrigenin 3-O-a-L-
rhamnoate) (28) and lanceotoxin B (5-O-acetylhelle-
brigenin 3-O-a-L-rhamnopyranoside) (29) (Anderson
et al. 1984). In the flower heads, leaves and stems
extract of K. tubiflora and in the roots of the hybrid K.
tubiflora 9pinnata, three bryotoxin A, B and C were
detected. In the extract from flower heads, leaves and
stems of the hybrid, K. daigremontiana 9pinnata,
only bryotoxins B and C were found. No bryotoxins
were detected in extract from Kalanchoe fedtschenkoi
Raym.-Hamet & H. Perrier (McKenzie et al. 1987).
Quantification of bufadienolides in leaves of K.
pinnata grown in Brazil and Germany (Oufir et al.
2015) revealed that bryophyllin A (5), bersaldegenin-
3-acetate (4), bersaldegenin-1,3,5-orthoacetate (2) and
bersaldegenin-1-acetate (3) are main bufadienolide
components of these plant organs. In plants grown in
Brazil, the total bufadienolide concentrations ranged
from 16.28 to 40.50 mg/100 g of dry weight. The total
content of bufadienolides in plant material from
Germany was lower and attained from 3.78 to
12.49 mg/100 g of dry weight. Additional analyses
of other species indicated that in leaves of K.
daigremontiana and in stems of K. tubiflora, bersalde-
genin-1,3,5-orthoacetate (4) was the predominant
bufadienolide compound. Contrary to K. pinnata, the
leaves of K. tubiflora contained very low amounts of
bryophyllin A (5), bersaldegenin-3-acetate (4), ber-
saldegenin-1,3,5-orthoacetate (2) and bersaldegenin-
1-acetate (3).
On the other hand, there are significant gaps in the
available literature on the presence of bufadienolides
in Kalanchoe species and their distribution in different
organs of these plants. Our preliminary studies
(unpublished data) suggested that the total content of
bufadienolides varied in different plant parts. While
bufadienolides content per gram of dried stems and
roots K. daigremontiana was 65 and 395 lg, respec-
tively, there was no occurrence of these compounds in
the leaves. This distribution in the plant was quite
unusual and probably reflected physiological and
ecological function of these compounds. It is assumed
that bufadienolides, similarly to other secondary
metabolites, are involved in chemical plant protection
against pathogenic microorganisms and herbivores.
They are also recognized as precursors of hormonal
substances and participate in the formation of mem-
branous structures.
Pharmacological actions of bufadienolides
of various origins
The therapeutic effects of bufadienolide-containing
preparations have been known from the ancient times.
A bufadienolide-rich plant Scilla maritima was used
by Egyptians to cure heart diseases. Bufadienolides
28 H2H CHO H OAc
29 H2H CHO H OAc
31 H2H CHO H OAc
Fig. 1 continued
1160 Phytochem Rev (2017) 16:1155–1171
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are also the principal bioactive ingredient of a
traditional Chinese drug Ch’an Su, containing the
skin secretions of toads such as Bufo gargarizans
Cantor and Bufo melanostictus Schneider. Currently,
the most investigated pharmacological activities of
bufadienolides of various origins are cardiotonic and
anticancer properties. Other physiological actions of
bufadienolides include blood pressure stimulating,
antiangiogenic, antiviral, immunomodulatory and
antibacterial activities (Gao et al. 2011; Kamboj
et al. 2013; Wei et al. 2017). Biological activity of
Kalanchoe-derived bufadienolides is a relatively new
Fig. 2 Structures of
compounds 2,5,7and 9
Fig. 3 Structures of the
compounds 23,24 and 25
Fig. 4 Structures of the
compounds 6and 30
Phytochem Rev (2017) 16:1155–1171 1161
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issue. For that reason, a number of reports directly
related to molecular background of pharmacological
action of bufadienolides isolated from Kalanchoe
plants is limited. Contemporary literature mostly
provides data on pharmacological actions and possible
therapeutic significance of animal bufadienolides,
however, some information on compounds that are
also present Kalanchoe species is available.
The main molecular mechanism of pharmacolog-
ical action of bufadienolides and their derivatives
involves the induction of a local increase of Na
as a
result of inhibition of a carrier enzyme: Na
ATPase (EC; the sodium pump), commonly
described as a ‘‘digitalis-like’’ effect. Na
ATPase is responsible for maintaining of electro-
chemical gradient of Na
and K
through the cell
membrane. The keeping of low Na
and high K
intracellular concentrations and membrane potential is
critical for excitability of nerves and muscle cells
(including cardiomyocytes) as well as for the sec-
ondary active transport. Bufadienolides have the
ability to alter myocardial ion balance resulting in an
increase of intracellular Ca
concentration ([Ca
via a backward-running of Na
exchanger, and
as a consequence, leading to contractions of cardiac
and arterial myocytes (Melero et al. 2000; Schoner and
Scheiner-Bobis 2007). Additionally, studies on
numerous cell lines confirmed the anticancer proper-
ties of different bufadienolides (Kamboj et al. 2013)
and provided some information on anticancer mech-
anisms and selective toxicity of bufadienolides
towards malignant cells. Studies on human liver
microsomes (HLMs) indicated that hydroxylation
and dehydrogenation might be the major metabolic
pathways of bufadienolides (Han et al. 2016). Molec-
ular mechanisms of anticancer activities of hellebrin
and its aglycone hellebrigenin (compounds that were
also found in Kalanchoe plants) were described by
Moreno et al. (2013). According to those authors, both
compounds are able to bind to the alpha subunits of the
-ATPase and display similar growth inhibi-
tory effects in different cancer cell lines, i.e. A549
(lung cancer), U373 (glioblastoma astrocytoma),
Hs683 (glioma), T98G (glioblastoma), MCF-7 (breast
adenocarcionoma), SKMEL-28 (melanoma), PC-3
(prostate cancer) and HT-29 (colorectal cancer). For
hellebrin, the growth inhibitory concentrations at 50%
) were estimated as 6–58 nM, while the IC
hellebrigenin ranged from 3 to 42 nM. Other
experiments (Yuan et al. 2016) conducted on human
glioblastoma U-87 cell line and a pancreatic SW1990
cancer cell line demonstrated that gamabufotalin and
arenobufagin (bufadienolides of animal origin) pos-
sessed selective cytotoxic activity against tumour cells
rather than normal cells (peripheral blood mononu-
clear cells, PBMCs). Both bufadienolides (at the final
concentrations of 1.6, 8, 40, 200 and 1000 ng/ml)
displayed dose-dependent anticancer effects, when
compared to control (untreated) cells. For gamabufo-
talin, IC
values were 16.8 ±6.5 and 8.1 ±1.5 ng/
ml in the U-87 and SW1990 cells, respectively.
Arenobufagin action was characterized by IC
10.3 ±3.3 and 9.9 ±2.2 ng/ml, in the U-87 and
SW1990 cells, respectively. Moreover, the authors
suggested that gamabufotalin might be a promising
candidate for using as an adjuvant therapeutic agent.
This opinion was based on data originated from
experiments on PBMCs treated with bufadienolides at
nontoxic concentrations, which resulted in a modula-
tion of fractions of CD4 ?CD25 ?Foxp3 ?regu-
lator T (Treg) cells in mitogen-activated PBMCs. In
pathophysiology of cancer and haematologic malig-
nancies, Treg cells were found to play a critical role in
development of tumour immunotolerance by sup-
pressing the host response to tumour immunity. Thus,
by decreasing the amount and activity of these cells,
gamabufotalin may enhance the efficiency of conven-
tional anticancer drugs (Yuan et al. 2016). Studies of
Zhang et al. (2016) revealed that the treatment of A549
cell line with gamabufotalin (5–500 nM) significantly
reduced viability of the cells, when compared to the
control (untreated A549 cells). For the 48 h-treatment,
the IC
value was 48.4 ±2.5 nM. Moreover, no
cytotoxicity was found in analogous experiments on
human normal lung cell line (HLF cells). Molecular
mechanisms of cytotoxic action of the bufadienolide
involved the G2/M cell cycle arrest and induction of
apoptosis in A549 cells. In vivo, gamabufotalin (10 or
20 mg/kg of body weight) was able to down-regulate
the protein level of Hsp90 in tumor tissues of the
xenograft mice, when compared to control animals
(treated with phosphate-buffered saline) (Zhang et al.,
2016). Furthermore, in studies of other scientists (Yu
et al. 2014), gamabufotalin (10, 50 and 100 nM)
suppressed the expression of cyclooxygenase 2 (COX-
2) in lung cancer cells, in comparison to the dimethyl
sulfoxide (DMSO) vehicle control group. Biochemi-
cal mechanisms of this anti-inflammatory action of
1162 Phytochem Rev (2017) 16:1155–1171
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gamabufotalin involve the inhibition of the phospho-
rylation of inhibitor of nuclear factor kappa-B (IjB),
which prevents the translocation of nuclear factor
kappa B (NF-jB) to nucleus and, in consequence,
halts the recruitment of NF-jB and p300 on COX-2
promoter (Yu et al. 2014). As another potential
mechanism of antitumour activity of gamabufotalin,
the inhibition of the vascular endothelial growth factor
(VEGF)-induced angiogenesis by suppressing vascu-
lar endothelial growth factor receptor 2 (VEGFR-2)
signaling pathway has been also suggested (Tang et al.
Recently published data (Bachmann et al. 2017)
suggest that the bufadienolide-enriched fraction from
K. pinnata leaf juice (containing bersaldegenin-1-
acetate, bryophyllin A, bersaldegenin-3-acetate, ber-
saldegenin-1,3,5-orthoacetate as well as two uniden-
tified compounds: flavonoid (m/z 581, [M ?H]
303 (aglycone)) and bufadienolide m/z 477
([M ?H]
) display biological activity that might be
useful in the treatment of overactive bladder. The
examined fraction (0.01–1 mg/ml) had the inhibitory
effect on detrusor contractility in vitro. The inhibition
was dose-dependent, and no such effects were found
for flavonoid fraction isolated from the leaf juice.
Biological activity of Kalanchoe species-derived
Anticancer effects
The existing evidence of anticancer properties of
bufadienolides originates mostly from research on
compounds isolated from animal sources, particularly
of toad venom (Takai et al. 2012). However, reports
indicating on the chemopreventive effects of Kalan-
choe bufadienlides are also available. Bryophyllin B,
isolated from Bryophyllum pinnatum (Lam.) Oken (K.
pinnata) was shown to be a potent cytotoxic agent
against the KB cell line, with the ED
value\80 ng/
ml (Yamagishi et al. 1989). Studies on 8 bufadieno-
lides, including kalanchosides A–C, isolated from the
aerial parts of K. gracilis Hance revealed considerable
cytotoxic/anticancer activities of all isolated com-
pounds against several human tumour cell lines such
as nasopharyngeal (KB) and its MDR variant (KB-
VIN), lung (A549), ovarian (1A9), prostate (PC-3),
ileocecal (HCT-8), and epidermoid (A431) cells.
Mostly, effectiveness of the examined bufadienolides
was higher than the effect of etoposide (a reference
cytostatic/anticancer drug) and attained the nanomolar
range of their concentrations (Wu et al. 2006).
Furthermore, bryophyllin B was able to inhibit the
replication of HIV in H9 lymphocytes, at the ED
value of \0.25 lg/ml and therapeutic index of
[6.27 lg/ml. Additionally, Huang et al. (2013)
demonstrated that bufadienolide glycosides isolated
from K. tubiflora displayed strong cytotoxicity against
four human cancer cell lines: A549, Cal-27 (oral
adenosquamous carcinoma), A2058 (melanoma) and
HL-60 (promyelocytic leukemia). Bufadienolide
effects were assessed in comparison with positive
controls, i.e. mitomycin-C and cycloheximide, while
0.05% DMSO-treated samples were used as vehicle
controls. IC
values for the examined glycosides
ranged from 0.01 to 10.66 lM. For mitomycin-C, IC
ranged from 4.63 to 9.34 lM, while IC
for cyclo-
heximide was detectable only in experiments on HL-
60 cells and attained 40.60 lM (Huang et al. 2013).
The cytotoxic effect against tumour cell lines was also
found in experiments with bufadienolides isolated
from the crude methanol extract of K. hybrida Desf. ex
Steud. Anticancer activity of the isolated compounds
(4 and 20 lg/ml) was evaluated in experimental
models of three cancer cell lines, i.e. MCF-7, NCI-
H460 (large cell lung cancer), and SF-268 (anaplastic
astrocytoma), using actinomycin D (10 mM) and
DMSO (0.3%) as positive and vehicle controls,
respectively. The strongest cytotoxic effects (even
up to 100% of growth inhibition) towards the exam-
ined cells were found for bersaldegenin 3-acetate and
daigredorigenin 3-acetate (Kuo et al. 2008).
Cardiotonic effects
The cardiac glycoside-like effects of a bufadienolide
compound, extracted from K. daigremontiana, were
demonstrated by Scholtysik et al. (1986). During the
studies on animals, the authors observed pharmaco-
logical effects similar to those evoked by digitalis
glycosides. The IC
for Na
-ATPase activity
in vitro was estimated as 1.4 910
M, while for
ouabain (a reference compound) this parameter was
M. Intravenous infusion of the examined
bufadienolide to guinea-pigs with a rate of 20 lg/kg/
min resulted in ventricular arrhythmias and death after
accumulated doses of about 760 and 860 lg/kg of
Phytochem Rev (2017) 16:1155–1171 1163
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Table 2 The systematic names of compounds 1–31
No. Name Systematic name Species and plant organs References
K. tomentosa (leaves) Rasoanaivo et al.
2 Bersaldegenin-1,3,5-
orthoacetate =Melianthugenin
(1b,3b,5b)-1,3,5-[(1R)-ethylidynetris(oxy)]-14-hydroxy-19-oxobufa-20,22-dienolide K. daigremontiana 9
tubiflora (leaves)
Supratman et al.
3 Bersaldegenin-1-acetate 3-acetoxy-1,5,14-trihydroxy-19-oxobufa-20,22-dienolide K. daigremontiana 9
tubiflora (leaves)
Supratman et al.
4 Bersaldegenin-3-acetate (1b,3b,5b)-3-(acetyloxy)-1,5,14-trihydroxy-19-oxobufa-20,22-dienolide K. daigremontiana 9
tubiflora (leaves)
Supratman et al.
5 Bryophyllin A =Bryotoxin C [1b(R),3b,5b,11a]-1,3,5-ethylidynetris(oxy)-11,14-dihydroxy-19-oxo-bufa-20,22-
K. pinnata (leaves) Supratman et al.
6 Bryophyllin B (1b,3b,5b,8n,9n,10n,11a,19R)-1-acetoxy-3,5,14,19-tetrahydroxy-11,19-epoxybufa-
K. gracilis (aerial parts) Wu al et. (2006)
7 Bryophyllin C [1b(R),3b,5b,11a]-1,3,5-ethylidynetris (oxy)-11,14,19-trihydroxybufa-20,22-dienolide K. pinnata (leaves) Supratman et al.
8 Daigredorigenin-3-O-acetate 3-(acetyloxy)-1,5,14-trihydroxy-, (1b,3b,5b)-bufa-20,22-dienolide K. daigremontiana
(aerial parts and roots)
Wagner et al.
9 Daigremontianin (1b,3b,5b,11a)-1,3,5-ethylidynetris(oxy)-11,14-dihydroxy-12,19-dioxobufa-20,22-
K. daigremontiana 9
tubiflora (leaves)
Supratman et al.
10 Hellebrigenin (3b,5b)-3,5,14-trihydroxy-19-oxobufa-20,22-dienolide K. gracilis (aerial parts) Wu et al. (2006)
11 Hellebrigenin-3-acetate (3b,5b)-3-acetoxy-5,14-dihydroxy-19-oxobufa-20,22-dienolide K. gracilis (aerial parts) Wu et al. (2006)
12 Kalanchoside A (3b,5b)-3-[(6-deoxy-a-D-glucopyranosyl)oxy]-5,14-dihydroxy-19-oxobufa-20,22-
K. gracilis (aerial parts) Wu et al. (2006)
13 Kalanchoside B (3b,5b)-3-[(6-deoxy-a-L-galactopyranosyl)oxy]-5,14-dihydroxy-19-oxobufa-20,22-
K. gracilis (aerial parts) Wu et al. (2006)
14 Kalanchoside C 12-oxohellebrigenin-3-O-4,6-dideoxy-a-ribo-hexopyranoside K. gracilis (aerial parts) Wu et al. (2006)
15 Kalandaigremoside A 1b,3b,5b,14b,19-pentahydroxybufa-20,22-dienolide K. daigremontiana
Szajwaj et al.
16 Kalandaigremoside B 19-(acetyloxy)-1b,3b,5b,14-tetrahydroxybufa-20,22-dienolide K. daigremontiana
Szajwaj et al.
17 Kalandaigremoside C 3b-(O-a-L-rhamnopyranosyl)-5b,11a,14,19-tetrahydroxybufa-20,22-dienolide K. daigremontiana
Szajwaj et al.
18 Kalandaigremoside D 19-(acetyloxy)-3b,5b,11a,14-tetrahydroxybufa-20,22-dienolide K. daigremontiana
Szajwaj et al.
1164 Phytochem Rev (2017) 16:1155–1171
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
Table 2 continued
No. Name Systematic name Species and plant organs References
19 Kalandaigremoside E 3b,5b,11a,14b,19-pentahydroxy-12-oxo-bufa-20,22-dienolide K. daigremontiana
Szajwaj et al.
20 Kalandaigremoside F 19-(acetyloxy)-3b,5b,11a,14b-tetrahydroxy-12-oxo-bufa-20,22-dienolide K. daigremontiana
Szajwaj et al.
21 Kalandaigremoside G 19-(acetyloxy)-1b,3b,5b,11a,14b-pentahydroxy-12-oxo-bufa-20,22-dienolide K. daigremontiana
Szajwaj et al.
22 Kalandaigremoside H 1b-(acetyloxy)-3b,5b,11a,14b,19-pentahydroxy-12-oxo-bufa-20,22-dienolide K. daigremontiana
Szajwaj et al.
23 Kalanhybrin A Chol-22-ene-19,24-dial, 3-(acetyloxy)-14,21-epoxy-1,5,22-trihydroxy-21-methoxy-,
K. hybrida (whole plant) Kuo et al. (2008)
24 Kalanhybrin B Chol-22-ene-19,24-dial, 1-(acetyloxy)-14,21-epoxy-3,5,22-trihydroxy-21-methoxy-,
K. hybrida (whole plant) Kuo et al. (2008)
25 Kalanhybrin C Chol-22-en-24-al, 3-(acetyloxy)-14,21-epoxy-1,5,22-trihydroxy-21-methoxy-,
K. hybrida (whole plant) Kuo et al. (2008)
26 Kalantuboside A Bufa-20,22-dienolide, 3-[(3-O-acetyl-4,6-dideoxy-a-L-ribo-hexopyranosyl)oxy]-
5,11,14-trihydroxy-12,19-dioxo-, (3b,5b,11a)-
K. tubiflora (whole
Huang et al.
27 Kalantuboside B Bufa-20,22-dienolide, 3-[(4,6-dideoxy-a-L-ribo-hexopyranosyl)oxy]-5,11,14-
trihydroxy-12,19-dioxo-, (3b,5b,11a)-
K. tubiflora (whole
Huang et al.
28 Lanceotoxin A [(3S,5S,8R,9S,10S,13R,14S,,17R)-5-acetyloxy-10-formyl-14-hydroxy-13-methyl-17-
K. lanceolate (whole
Anderson et al.
29 Lanceotoxin B [(3S,5S,8R,9S,10S,13R,14S,17R)-10-formyl-14-hydroxy-13-methyl-17-(6-oxopyran-
K. lanceolata (whole
Anderson et al.
30 Methyl daigremonate Methyl[1b,3b,5b,11a,12a]-(22E)-1,3,5-ethylidynetris(oxy)-14,21-epoxy-11,12-
K. daigremontiana 9
tubiflora (leaves)
Supratman et al.
31 Thesiuside 5-O-acetylhellebrigenin 3-O-b-D-glucopyranoside K. gracilis (aerial parts) Wu et al. (2006)
Phytochem Rev (2017) 16:1155–1171 1165
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body weight, respectively. However, in general, the
examined bufadienolide was less toxic than ouabain.
Anti-viral activity
Bufadienolides isolated from leaves of K. pinnata and
K. daigremontiana 9tubiflora are able to inhibit the
activation of Epstein-Barr virus early antigen (EBV-
EA) in Raji cells, induced by 12-O-tetradecanoylphor-
bol-13-acetate. Bryophyllin A had the strongest
inhibitory effect (IC
=0.4 lM), while compounds
lacking the orthoacetate moiety such as bryophyllin C
and bersaldegenin-3-acetate possessed significantly
lower activities (IC
=1.6 and 3 lM, respectively),
when compared to control samples (untreated with the
bufadienolide) (Supratman et al. 2001a,b).
Inhibition of serine proteinases
So far, research on bufadienolides and enzyme
interactions has been focused only on the inhibition
of ATP-ase activity. The issue of inhibition of other
groups of enzymatic proteins by bufadienolides has
appeared in the literature within last 2 years. In 2015,
Shibao and co-authors published results from studies
on a serine proteinase inhibitor, isolated from Rhinella
schneideri (Schneider’s toad) poison. The inhibitor
was identified as lithocholic acid, a biosynthetic
precursor of bufadienolide. In spite of the fact that
the study was conducted on animal-derived prepara-
tion, it should be mentioned as the first report
confirming that bufadienolide-type compound might
suppress the enzymatic activity of serine proteinase.
Thus, the influence of Kalanchoe-derived bufadieno-
lides on enzymatic properties of serine proteinases still
is very poorly evidenced. Inhibitory action of bufa-
dienolide-rich fraction from K. daigremontiana on
enzymatic activity of thrombin has been recently
described by Kolodziejczyk-Czepas et al. (2017).
Native (untreated with the examined fraction) throm-
bin was used as a control sample. A serine proteinase
enzyme—thrombin (plasma coagulation factor II), is
responsible for the formation of fibrin clot, and thus,
for the prevention of uncontrolled blood loss after
injury of blood vessel. In the above in vitro study,
bufadienolide-rich fraction inhibited enzymatic activ-
ity of thrombin with IC
=2.79 lg/ml. The efficacy
of a reference compound (direct inhibitor of throm-
bin)—argatroban (anti-thrombotic drug) was charac-
terized by IC
=0.78 lg/ml. On the other hand,
analysis of kinetic parameters of the reaction indicated
that K. daigremontiana fraction contains compounds
with diverse inhibitory mechanisms, when compared
to argatroban. Components of the investigated frac-
tions were uncompetitive inhibitors of thrombin. In
silico studies on interactions of the most common
compounds, identified in the examined bufadienolide-
rich fraction to crystal structure of thrombin were also
conducted. The obtained results indicated that for the
inhibitory effect of K. daigremontiana fraction, most
likely the presence of compounds such as bersalde-
genin-1,3,5-orthoacetate, bersaldegenin-1-acetate,
bersaldegenin, hovetrichoside C, deigredorigenin-3-
acetate is responsible.
Bufadienolides as antioxidants?
Due to hydrophobic, steroid structure of bufadieno-
lide-type compounds, antioxidant properties of those
substances are considered to be weak. However,
existing evidence indicated that bufadienolides pos-
sess some antioxidant potential. Moreover, this group
of compounds may be a base for development of new
derivatives with enhanced antioxidant properties and
decreased toxicity (obtained by chemical alterations of
the pyrone moiety) (Aucamp 2014). Recent studies on
bufadienolide-rich fraction of Kalanchoe daigremon-
tiana roots demonstrated that its DPPH
ability was characterized by EC
=21.80 lg/ml
(Kolodziejczyk-Czepas et al. 2016). Under the same
experimental conditions, for the reference compounds,
i.e. Trolox and (-)-epicatechin, EC
values were 4.64
and 3.30 lg/ml, respectively. It should be emphasized
that earlier results, obtained by other authors in
analogous experiments on different Kalanchoe plants
indicated on significantly lower antioxidant efficacy of
extracts, originated from different organs of Kalan-
choe species (Sharker et al. 2012; Quazi Majaz et al.
2011a,b). Furthermore, antioxidant action of the
mentioned bufadienolide-rich extract of Kalanchoe
daigremontiana was also confirmed using an experi-
mental model of blood plasma exposed to peroxyni-
trite-induced oxidative stress (Kolodziejczyk-Czepas
et al. 2016).
1166 Phytochem Rev (2017) 16:1155–1171
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
Other biological actions of Kalanchoe
Toxic (insecticidal) action of daigremontianin and
bersaldegenin-1,3,5-orthoacetate, isolated from the
leaves of K. daigremontiana, was demonstrated using
an experimental system of larvae of Bombyx mori
(Maharani et al. 2008).
The risk of side effects and prospects
for pharmacological use of bufadienolides
occurring in Kalanchoe species
During the last 10 years, a growing interest in the
evaluation of the metabolome of Kalanchoe plants and
biological activities of Kalanchoe-derived extracts
and substances, including bufadienolides has been
observed. For instance, from the total amount of 483
publications containing the name ‘Kalanchoe’’ avail-
able in the Medline/Pubmed database, over 150
records derive from the last 10 years (data from 30
March, 2017; search criteria ‘Kalanchoe’’). On the
other hand, contrary to in vivo studies, confirming
pharmacological effects of composed extracts isolated
from different Kalanchoe species, physiological
effects of bufadienolides extracted from these plants
and their safety have been poorly described. The vast
majority of reports on pharmacological activity of
different Kalanchoe-based drugs still derive from
traditional medicine and concern preparations based
on crude extracts. However, studies on standardized
preparations from Kalanchoe species also are avail-
able. According to data from a service of the U.S.
National Institutes of Health ‘‘’’
(, data from 30
March, 2017; search criteria ‘‘Kalanchoe’’ or
‘‘ Bryophyllum’), four clinical studies on Bryophyllum
pinnatum/K. pinnata have been recorded. Further-
more, after using a word ‘‘bufadienolide’’ four another
results have been appeared, while a combination of
‘‘ Kalanchoe’ and ‘bufadienolide’ have not provided
any results. No information on animal or clinical
studies on therapeutic effects of bufadienolides iso-
lated from Kalanchoe was found in Medline/Pubmed,
Scopus, ScienceDirect/Elsevier and Springer Link/
ICM databases (data from 30 March, 2017). Biological
activities of bufadienolides that are synthetized by
these plants are very promising from a
pharmacological point of view, however, they have
been mostly studied in vitro. Therefore, nowadays,
only a preliminary indication of the most promising
prospects for pharmaceutical uses of bufadienolides is
The therapeutic use of most bufadienolides is
limited by to a narrow therapeutic index and risk of
development of cardiotoxicity (Cheng 2001; Pamnani
et al. 1994). A risk of toxicity of bufadienolide-
containing plant extracts is inadequately evaluated.
For example, no toxicity of the bufadienolide-rich K.
daigremontiana fraction on blood platelets was found
in vitro (Kolodziejczyk-Czepas et al. 2016). On the
other hand, studies in South Africa indicated that
ingestion of cumulative neurotoxic various plant-
derived bufadienolides such as cotyledoside, tyledo-
sides, orbicusides and lanceotoxins is a potential risk
to humans (Botha 2016). Hence, numerous investiga-
tions have been developed to generate chemical and
biotransformed bufadienolide derivatives or ana-
logues with effective therapeutic action and consider-
ably reduced toxicity. The in vitro biotransformations
of natural bufadienolides have been conducted in
various systems—in plant cell suspension cultures,
fungi and bacteria (Gao et al. 2011). Some of these
modified bufadienolides were able to selectively kill
malignant cells. Studies of Daniel et al. (2003) showed
this preferential cytotoxic action towards malignant
cells for both a natural cardioactive bufadienolide—
hellebrin (0.1–100 lM) as well as for its three
derivatives (100 lM), lacked the cardioactive proper-
ties. Medium for the controls was supplemented with
corresponding amounts of the used bufadienolide
vehicle. While normal peripheral blood mononuclear
cells were affected to a minimal extent, the examined
substances induced the caspase-dependent pathway
and initiated apoptosis in Jurkat T lymphoblasts. Since
the therapeutic use of bufadienolides in anti-cancer
therapy is limited by their influence on heart physi-
ology, a considerable potential of using in cancer
therapy may have compounds possessing a tumour-
specific cytotoxicity with simultaneous lack of cardiac
activity (Daniel et al. 2003). Current research of
bufadienolides (hellebrigenin, among others) also
covers some pharmacokinetic aspects of their interac-
tions with human serum albumin (HSA), the main
carrier of various drugs. In vitro and in silico analyses
indicated that the binding affinity for HSA of various
bufadienolides is considerably related to differences in
Phytochem Rev (2017) 16:1155–1171 1167
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
their structures. The presence of C=O bond at the C12
position decreased the binding affinity for HSA, while
other polar groups increased the bufadienolide affinity
to HSA. In particular, the presence of 11-OH or
16-OAc groups may be important for anchoring
bufadienolides within site I of the HSA pocket. The
11-OH or 16-OAc-mediated interactions of bufa-
dienolide and HSA involve the hydrogen bonding
(H-bonding) with protein Tyr150 or Lys199 groups,
respectively (Zhou et al. 2015).
Another way of the enhancing the therapeutic effect
and reducing the toxicity of anticancer drugs such as
bufadienolides may be preparation of long-circulating,
poloxamer-modified liposomes. According to Hu et al.
(2011), these liposomes have significantly prolonged
retention time, when compared to bufadienolide
solutions and unmodified liposomes. The LD
of modified liposomes was about 3.5 times higher than
the LD
recorded for bufadienolide solution (i.e. 4.48
and 1.28 mg/kg, respectively). The use of bufadieno-
lide liposomes resulted in a considerable increase of
anti-tumour efficiency both in mice bearing H22 liver
cancer cells and Lewis pulmonary cancer cells (2.15
and 2.96 times, respectively), compared to the anti-
cancer effects observed in animals treated with
bufadienolide solution. Promising results have been
obtained by Mexican scientists (Alvarado-Palacios
et al. 2015) in experiments on using nanocapsules
containing the aquoethanolic extract from K. daigre-
montiana as selective anticancer agents. The nanocap-
suled extract was characterized by higher cytotoxic
efficacy (IC
=48.53 lg/ml) towards MDA-MB-
231 metastatic breast cancer cell line, when compared
to the non-encapsulated aquoethanolic extract
=61.29 lg/ml). Moreover, studies on non-
cancerous breast cell line MCF 10A revealed no
cytotoxic effect of the nanocapsules containing the
aquoethanolic extract of K. daigremontiana (at con-
centrations B200 lg/ml), whereas the non-encapsu-
lated extract displayed significant cytotoxic effect
=100.2 lg/ml).
Ethnomedicinal plants of the Kalanchoe genus may be
regarded as a new source of bufadienolides, since
synthesis of these substances has been confirmed for
these species. At present, however, toad venom
remains the main source of these compounds. On the
other hand, a growing number of reports have
confirmed that Kalanchoe-derived bufadienolides dis-
play a wide range of biological actions, including
cardiotonic, anticancer, anti-viral and other properties.
Despite these promising findings, the therapeutic use
of Kalanchoe plants is considerably limited by the lack
of clinical evidence. Therefore, further studies on
medicinal applications of bufadienolides and extracts
of Kalanchoe species origin are required.
Acknowledgements This work was supported by Grants
506/1136 (from University of Lodz, Poland) and (2012/05/B/
NZ9/00812 from the National Science Centre, Poland). We
thank Łukasz Pecio for the preparing structures of compounds.
Additionally, the authors would to thank Prof. Pawel Nowak for
helpful suggestions, and Prof. Beata Olas for reading the
preliminary outline of this work in 2013.
Open Access This article is distributed under the terms of the
Creative Commons Attribution 4.0 International License (http://, which permits unre-
stricted use, distribution, and reproduction in any medium,
provided you give appropriate credit to the original
author(s) and the source, provide a link to the Creative Com-
mons license, and indicate if changes were made.
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... These five compounds can be seen in the UV chromatogram ( Fig 1A; peaks 1-5, blue line). Through extracted ion chromatogram (EIC) analysis, it was also possible to annotate three bufadienolides (6)(7)(8): bryophillin A (6), bersaldegenin-3-acetate (7), and bersaldegenin-1,3,5-orthoacetate (8) (Fig 1B), consistent with previous literature [22]. ...
... Bufadienolides, which were discovered in trace amounts in our analysis, are considered toxic owing to their cardiotonic effects [36]. However, studies have provided evidence that they possess anticancer, antiviral, and antioxidant properties [22]. It would be interesting to further investigate the therapeutic and toxicological effects of the isolated compounds in ZF embryos. ...
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Bryophyllum pinnatum (Lam.) Oken (BP) is a plant that is used worldwide to treat inflammation, infections, anxiety, restlessness, and sleep disorders. While it is known that BP leaves are rich in flavonoids, the extent of the beneficial and toxic effects of its crude extracts remains unclear. Although some neurobehavioral studies using leaf extracts have been conducted, none has examined the effects of water-extracted leaf samples. The zebrafish is a powerful animal model used to gain insights into the efficacy and toxicity profiles of this plant due to its high fecundity, external development, and ease of performing behavioral assays. In this study, we performed behavioral testing after acute exposure to different concentrations of aqueous extract from leaves of B . pinnatum (LABP) on larval zebrafish, investigating light/dark preference, thigmotaxis, and locomotor activity parameters under both normal and stressed conditions. LABP demonstrated dose-and time-dependent biphasic effects on larval behavior. Acute exposure (25 min) to 500 mg/L LABP resulted in decreased locomotor activity. Exposure to 300 mg/L LABP during the sleep cycle decreased dark avoidance and thigmotaxis while increasing swimming velocity. After sleep deprivation, the group treated with 100 mg/L LABP showed decreased dark avoidance and increased velocity. After a heating stressor, the 30 mg/L and 300 mg/L LABP-treated groups showed decreased dark avoidance. These results suggest both anxiolytic and psychoactive effects of LABP in a dose-dependent manner in a larval zebrafish model. These findings provide a better understanding of the mechanisms underlying relevant behavioral effects, consequently supporting the safe and effective use of LABP for the treatment of mood disorders.
... Many species are popular in other regions, including in Europe, due to their pro-health properties. Among the biological activities of Kalanchoe plants, the most well-known are their anti-inflammatory, antioxidative, analgesic, antimicrobial, antidiabetic, antimalarial, and antiallergic properties [1][2][3][4][5][6][7][8][9][10]. An anticancer action has also been reported; however, cytotoxic studies on these plants are still being ...
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Kalanchoe species are succulents with anti-inflammatory, antioxidant, and analgesic properties, as well as cytotoxic activity. One of the most popular species cultivated in Europe is Kalanchoe daigremontiana Raym.-Hamet and H. Perrier. In our study, we analyzed the phytochemical composition of K. daigremontiana water extract using UHPLC-QTOF-MS and estimated the cytotoxic activity of the extract on human ovarian cancer SKOV-3 cells by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, flow cytometry, luminometric, and fluorescent microscopy techniques. The expression levels of 92 genes associated with cell death were estimated via real-time PCR. The antioxidant activity was assessed via flow cytometry on human keratinocyte HaCaT cell line. The DPPH (2,2-diphenyl-1-picrylhydrazyl) radical and FRAP (ferric-reducing antioxidant power) assays were also applied. We identified twenty bufadienolide compounds in the water extract and quantified eleven. Bersaldegenin-1,3,5-orthoacetate and bryophyllin A were present in the highest amounts (757.4 ± 18.7 and 573.5 ± 27.2 ng/mg dry weight, respectively). The extract showed significant antiproliferative and cytotoxic activity, induced depolarization of the mitochondrial membrane, and significantly arrested cell cycle in the S and G2/M phases of SKOV-3 cells. Caspases-3, 7, 8, and 9 were not activated during the treatment, which indicated non-apoptotic cell death triggered by the extract. Additionally, the extract increased the level of oxidative stress in the cancer cell line. In keratinocytes treated with menadione, the extract moderately reduced the level of oxidative stress. This antioxidant activity was confirmed by the DPPH and FRAP assays, where the obtained IC50 values were 1750 ± 140 and 1271.82 ± 53.25 μg/mL, respectively. The real-time PCR analysis revealed that the extract may induce cell death via TNF receptor (tumor necrosis factor receptor) superfamily members 6 and 10.
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The genus Kalanchoe, mostly indigenous from Madagascar and Tropical Africa, is widely traded for ornamental value. In this study, we provided an updated list of wild and cultivated Kalanchoe taxa in Ecuador; we analyzed the temporal–spatial pattern of their records, and we categorized the invasion status for each taxon and its environment preferences. The records of any taxa belonging to this genus were compiled from an extensive search using various information sources. Our results confirmed the presence of 16 taxa of Kalanchoe in the country. Seven species and a hybrid were detected in the wild. Kalanchoe densiflora, K. laxiflora, K. pinnata, K. tubiflora, and K. ×houghtonii were categorized as invasive. We detected invasive records of some of these plants in protected areas. Almost all taxa had at least one record as cultivated, suggesting that the invasion pathway is ornamental trade. Kalanchoe pinnata individuals in the wild were recorded in the four biogeographic regions of Ecuador, which could be associated with the wide range of precipitations and temperatures in which the species may dwell. Our study highlights the importance of reducing the ornamental value and limiting the use of Kalanchoe taxa with invasive potential in horticulture and promoting, instead, the use of indigenous species.
2H-pyran-2-one is six-membered oxygen heterocycle having one oxygen in the ring and oxo group at position 2 of the ring and is also known as cyclic unsaturated ester. Two double bonds present are in the conjugation of the ring oxygen. 2-Pyranone-based compounds are well known for more than a century, but the chemistry of these compounds was unexplored and recognized after 1960. Various plants, animals, marine organisms, insects, and bacteria contain pyranone-based compounds in their biological system and participate in various biological processes such as defense mechanism, biosynthetic intermediates, etc. They are reported as HIV protease inhibitors, antifungals, anticonvulsants, cardiotonics, pheromones, antimicrobials, natural pigments, antitumor, and plant growth regulators. In addition, they are synthetically very important precursors for the generation of large class of molecules, for example, use of triacetic acid lactone (I) and tetraacetic acid lactone (II) involved for the assembly of α-chymotrypsin, solanopyrones, elastase, pheromones, and coumarins. 6-Aryl-4-methylsulfanyl/sec.amino-2H-pyran-2-one-3-carbonitriles are used for the assembly of larger class of carbocycles and heterocycles.
Using nanocrystals (NCs) technology may be a promising drug delivery strategy for oral administration of multicomponent anticancer drugs. However, the intestinal epithelium and the mucus layer on the intestine extremely limited drug transport and absorption by orally. In this study, we selected multicomponent inartificial compound Bufadienolides (BU) with broad spectrum antitumor activity as the model drug to prepare BU NCs with different stabilizers by wet grinding, and explored the efficiency of penetrating through the mucus layer and transporting intestinal epithelial cells in vitro and ex vivo. Results revealed that BU NCs can dramatically improve dissolution behavior synergistically and the efficiency of mucus permeation. Besides, we found that BU NCs with different stabilizers enhanced cellular uptake, which was mainly attributed to increasing or changing the endocytosis pathway and plasma membrane/Endoplasmic reticulum (ER) pathway involved in the transmembrane transport of NCs. Furthermore, BU NCs could definitely improve intestinal absorption efficiency and change the absorption site of BU ex vivo. This multi-angle exploration will provide reference for the development of BU oral delivery formulations.
Objetivo: Evaluar la actividad antioxidante y antiinflamatoria en extractos hidroalcohólicos de las partes aéreas de Kalanchoe daigremontiana Raym. -Hamet & H. Perrier. Materiales y métodos: Se obtuvieron extractos hidroalcohólicos utilizando como solventes etanol (EtOH) y metanol (MeOH) en diferentes concentraciones. Los fenoles totales y los flavonoides se cuantificaron con el método de Folin Ciocalteu y el de complejación de aluminio respectivamente. Metodología: Para determinar la actividad antioxidante en los extractos se utilizó la técnica de decoloración del radical libre DPPH» (2,2-difenil-1-picrilhidracilo) y para la actividad antiinflamatoria se evaluó la capacidad inhibitoria de la enzima lipoxigenasa (LOX). Resultados: El extracto etanólico al 70% presentó mayor capacidad antioxidante frente a los radicales DPPH» y mayor inhibición sobre la enzima lipooxigenasa como respuesta antiinflamatoria, lo cual estaría relacionado con el contenido de fenoles y flavonoides. Conclusiones: Esto sugiere que el K. daigremontiana posee propiedades antioxidantes y antiinflamatorias.
Introduction : Kalanchoe laxiflora (Family Crassulaceae) contains different bioactive phytochemicals, which possess several biological activities. The present study aimed at assessing the antioxidant, cytotoxic, antidiabetic, and antibiofilm activities of Kalanchoe laxiflora flower extracts, as well as identification of their phytochemicals. Methods : The total phenolic and flavonoid contents of the tested extracts were established. The antioxidant, cytotoxic, antidiabetic and antibiofilm activities were also evaluated using different methods. The phytoconstituents of K. laxiflora flower extracts were profiled using LC-ESI(-ve)-MS analysis. Results : The phytochemical investigations showed that the n-BuOH fraction had the highest total phenolic content (397.25 ± 1.18 mg GAE / g ext.) and flavonoid content (99.71 ± 0.24 mg RE / g ext.). The n-BuOH fraction also possessed the most potent antioxidant activities (DPPH; SC50=18.72±1.81 µg/mL, RPA; 78.26±0.49 mg AAE /g ext.), and TAC; 446.37±0.23 mg AAE /g ext.) and cytotoxic potential in HepG2 cells with IC50= 9.15±0.33 μg/mL. Furthermore, it exhibited the strongest α-amylase and α-glucosidase inhibitory activities (IC50= 54.72±0.96 µg/mL and 89.02±0.99 µg/mL, respectively). Whereas, the 85% methanolic extract exhibited the highest antibiofilm activity against Escherichia coli (MBIC100= 15.63 µg/ml). Additionally, the LC-ESI(-ve)-MS analysis of K. laxiflora extracts led to the characterization of a total of 35 compounds classified to flavonoid glycosides, gallotannins, alkaloids and fatty acids, which are considered to be the major components. Conclusions : K. laxiflora extracts reduce ROS, improve antidiabetic responses and inhibit bacterial growth. The identification of K. laxiflora specific phytoconstituents will provide a starting point to carry out further in vivo studies on these extracts.
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Toxic metals such as aluminium accumulation in the brain have been associated with the pathophysiology of several neurodegenerative disorders. Bryophyllum pinnatum leaves contain a vast array of polyphenols, particularly flavonoids, that may play a role in the prevention of toxic and degenerative effects in the brain. This study assessed the neuro-restorative potential of leaves of B. pinnatum enriched flavonoid fraction (BPFRF) in aluminium-induced neurotoxicity in rats. Neurotoxicity was induced in male Wistar rats by oral administration of 150 mg/kg body weight of aluminium chloride (AlCl3) for 21 days. Rats were grouped into five (n = 6); Control (untreated), Rivastigmine group, AlCl3 group and BPFRF group (50 and 100 mg/kg b.wt.) for 21days. Neuronal changes in the hippocampus and cortex were biochemically and histologically evaluated. Expression patterns of acetylcholinesterase (AChE) mRNA were assessed using semi-quantitative reverse-transcription-polymerase chain reaction protocols. Molecular interactions of BPFRF compounds were investigated in silico. The results revealed that oral administration of BPFRF ameliorated oxidative imbalance by augmenting antioxidant systems and decreasing lipid peroxidation caused by AlCl3. BPFRF administration also contributed to the down-regulation of AChE mRNA transcripts and improved histological features in the hippocampus and cortex. Molecular docking studies revealed strong molecular interactions between BPFRF compounds, catalase, superoxide dismutase and glutathione peroxidase Overall, these findings suggest the neuroprotective effect of Bryophyllum pinnatum against aluminium-induced neurotoxicity.
p>Plak adalah pembentukan komunitas bakteri yang terorganisir pada permukaan gigi yang berupa lapisan tipis tidak berwarna. Pseudomonas aeruginosa adalah salah satu bakteri yang sangat berperan pada pembentukan plak. Metabolit sekunder yang terkandung dalam tanaman Kalanchoe tomentosa dapat digunakan sebagai antibakteri. Tahapan penelitian diawali dengan maserasi menggunakan pelarut n-heksan dan diklorometana. Ekstrak diklorometana dipisahkan menggunakan Kromatografi Cair Vakum (KCV) dengan n-heksana dan etil asetat sebagai pelarut bergradien selanjutnya direkristalisasi dengan n-heksan, hasil rekristalisasi didapatkan isolat 1. Isolat 1 dikarakterisasi dengan spektroskopi infra-red (IR), ultraviolet (UV) dan nuclear magnetic resonance hydrogen (<sup>1</sup>H NMR) serta dibandingkan dengan literatur. Isolat 1 diketahui merupakan stigmast-5-en-3ꞵ-ol atau ꞵ-sitosterol. Pada ekstrak diklorometana dan senyawa ꞵ-sitosterol kemudian ditentukan aktivitas antibakteri secara mikrodilusi didapatkan hasil bahwa ekstrak diklorometana dan senyawa ꞵ-sitosterol bukan hanya menunjukan sifat bakteriostatik tetapi juga bakterisid kuat terhadap bakteri Pseudomonas aeruginosadengan nilai Konsentrasi Hambat Minimum (KHM) <100 µg/mL. Steroid Compounds from Cocor Bebek (Kalanchoe tomentosa) as Antibacterial Pseudomonas aeruginosa. Plaque is a thin, colorless layer that adheres tightly to the surface of the teeth and contains a collection of bacteria. One of the bacteria that can form dental plaque is Porphyromonas aeruginosa. Kalanchoe tomentosa contain secondary metabolite which can be used as antibacterial. Stages of the study begin with maceration using n-hexane and dichloromethane. Dichloromethane extract was separated using Vacuum Liquid Chromatography (KCV) using n-hexane and ethyl acetate solvents and then recrystallized with n-hexane, the recrystallization results were obtained by isolate 1. Isolate 1 was marked by IR spectroscopy, and <sup>1</sup>HNMR and compared with literature. Isolate 1 is known as stigmast-5-en-3ꞵ-ol or ꞵ-sitosterol. In dichloromethane extracts and ꞵ-sitosterol is determined by microdilution by microdilution. The results showed that dichloromethane extract and ꞵ-sitosterol compound not only showed strong bacteriostatic but also bacterisid activity against P. aeruginosa with Minimum Inhibiroty Concentration (MIC) <100 μg / mL.</p
Full-text available
Background Cinobufacin injection, also known as huachansu, is a preparation form of Cinobufacini made from Cinobufacin extract liquid. Despite that Cinobufacin injection is shown to shrink liver and gastric tumors, improving patient survival and life quality, the effective components in Cinobufacin remain elusive. In this study, we aim to screen antitumor components from Cinobufacin injection to elucidate the most effective antitumor components for treatment of liver and gastric cancers. Materials and Methods High performance liquid chromatography (HPLC) and LC-MS/MS analysis were used to separate and determine the components in Cinobufacin injection. Inhibition rates of various components in Cinobufacin injection on liver and gastric cancer cells were determined with MTT assay; Hepatocellular carcinoma and gastric cancer models were used to assess the antitumor effect of the compounds in vivo. Results The major constituents in Cinobufacin injection include peptides, nucleic acids, tryptamines and bufotalins. MTT assay revealed that bufadienolides had the best antitumor activity, with peptides being the second most effective components. Bufadienolides showed significant inhibition rates on gastric and hepatocellular tumour growth in vivo. Conclusion Bufadienolides are the most effective components in Cinobufacini injection for the treatment of liver and gastric cancers. This discovery can greatly facilitate further research in improving the therapeutic effects of Cinobufacin injection, meanwhile reducing its adverse reaction.
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The growth inhibitory effects of bufadienolide compounds were investigated in two intractable cancer cells, a human glioblastoma cell line U-87 and a pancreatic cancer cell line SW1990. Among four bufadienolide compounds, a dose-dependent cytotoxicity was observed in these cancer cells after treatment with gamabufotalin and arenobufagin. The IC50 values of the two compounds were 3-5 times higher in normal peripheral blood mononuclear cells (PBMCs) than these values for both cancer cell lines. However, similar phenomena were not observed for two other bufadienolide compounds, telocinobufagin and bufalin. These results thus suggest that gamabufotalin and arenobufagin possess selective cytotoxic activity against tumor cells rather than normal cells. Moreover, a clear dose-dependent lactate dehydrogenase (LDH) release, a well-known hallmark of necrosis, was observed in both cancer cells treated with gamabufotalin, suggesting that gamabufotalin-mediated cell death is predominantly associated with a necrosis-like phenotype. Of most importance, treatment with as little as 8 ng/ml of gamabufotalin, even an almost non-toxic concentration to PBMCs, efficiently downregulated the percentages of CD4+CD25+Foxp3+ regulator T (Treg) cells in mitogen-activated PBMCs. Given that Treg cells play a critical role in tumor immunotolerance by suppressing antitumor immunity, these results suggest that gamabufotalin may serve as a promising candidate, as an adjuvant therapeutic agent by manipulating Treg cells to enhance the efficacy of conventional anticancer drugs and lessen their side-effects. These findings provide insights into the clinical application of gamabufotalin for cancer patients with glioblastoma/pancreatic cancer based on its cytocidal effect against tumor cells as well as its depletion of Treg cells.
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Gamabufotalin (CS-6) is a major bufadienolide of Chansu, which shows desirable metabolic stability and less adverse effect in cancer therapy. CS-6 treatment inhibited the proliferation of NSCLC in a nanomolar range. And CS-6 could induce G2/M cell cycle arrest and apoptosis in A549 cells. However, its molecular mechanism in antitumor activity remains poorly understood. We employed a quantitative proteomics approach to identify the potential cellular targets of CS-6, and found 38 possible target-related proteins. Among them, 31 proteins were closely related in the protein-protein interaction network. One of the regulatory nodes in key pathways was occupied by Hsp90. Molecular docking revealed that CS-6 interacted with the ATP-binding sites of Hsp90. In addition, CS-6 inhibited the chaperone function of Hsp90 and reduced expression of Hsp90-dependent client proteins. Moreover, CS-6 markedly down-regulated the protein level of Hsp90 in tumor tissues of the xenograft mice. Taken together, our results suggest that CS-6 might be a novel inhibitor of Hsp90, and the possible network associated with CS-6 target-related proteins was constructed, which provided experimental evidence for the preclinical value of using CS-6 as an effective antitumor agent in treatment of NSCLC.
An earlier prospective, randomised, placebo-controlled clinical trial had suggested that Bryophyllum pinnatum might have potential in the treatment of overactive bladder. Here we investigated the effects of B. pinnatum leaf press juice, fractions enriched in flavonoids and bufadienolides, and a flavonoid aglycon mixture and individual aglycons on detrusor contractility as a major target in overactive bladder treatment. The strength of the detrusor contractions was investigated using porcine muscle strips stimulated with KCl. B. pinnatum leaf press juice increased the contraction force of muscle strips. Treatment with the flavonoid-enriched fraction had almost no effect on contractility, while the bufadienolide-enriched fraction and flavonoid aglycons led to a concentration-dependent lowering of the contraction force. The data indicate that several components of B. pinnatum leaf press juice may contribute to the inhibitory effect on detrusor contractility, which in turn provides support to overactive bladder treatment with B. pinnatum.
Thrombin is an active plasma coagulation factor II, critical for the formation of fibrin clot during blood coagulation. For that reason, this protein is also a crucial target for different anti-thrombotic therapies. The work is based on in vitro evaluation of the inhibitory effect of bufadienolide-rich fraction, isolated from roots of Kalanchoe daigremontiana (1–50 μg/ml) on enzymatic properties of a serine proteinase − thrombin. The efficacy of the inhibition of amidolytic activity of thrombin (measured as a hydrolysis of the chromogenic substrate S-2238, Chromogenix) attained about 10 and 66%, respectively. The IC50, established for the examined bufadienolide fraction was 2.79 μg/ml, while the IC50 calculated for argatroban (reference compound) was 0.78 μg/ml. Linearization conducted using Lineweaver-Burk plot indicated that the K. daigremontiana fraction contains compounds that are uncompetitive inhibitors of thrombin. K. daigremontiana fraction was also able to reduce the proteolytic activity of thrombin towards its physiological substrate, i.e. fibrinogen. Additionally, this study is supported by in silico analysis of interactions of the most common compounds, identified in the examined in Kalanchoe extract to crystal structure of this enzyme.
Context: The main source of bufadienolides is toad venom; however, plants such as members of Kalanchoe Adans. (Crassulaceae) genus may also synthesize these bioactive substances. Objective: This is the first study on antioxidant effects and cytotoxicity of bufadienolide-rich fraction isolated from Kalanchoe daigremontiana Raym.-Hamet & H. Perrier. Materials and methods: The methanolic fraction was extracted from the plant roots and contained 0.48 mg bufadienolides/mg of dry mass (11α,19-dihydroksytelocinobufagin, bersaldegenin-1-acetate, bersaldegenin-1,3,5-orthoacetate, 19-(acetyloxy)-3β,5β,11α,14-tetrahydroxyl-12-oxo-bufa-20,22-dienolide and 19-(acetyloxy)-1β,3β,5β,14-tetrahydroxyl-bufa-20,22-dienolide, mainly). The cytotoxicity of K. daigremontiana fraction was evaluated in an in vitro experimental model of blood platelets. The viability of blood platelets was determined on the basis of a release of lactate dehydrogenase. Results: The fraction scavenged DPPH• radicals, with EC50 of 21.80 μg/mL. Studies on an experimental model of blood plasma under peroxynitrite-induced oxidative stress revealed that the plant preparation had moderate antioxidant properties. Levels of 3-nitrotyrosine and thiol groups indicated that the protective effect of K. daigremontiana was significant mainly for its concentration of 50 μg/mL. No effect was found in prevention of oxidation of low-molecular plasma thiols (glutathione, cysteine and cysteinylglycine). Simultaneously, measurements of lipid hydroperoxides and thiobarbituric acid-reactive substances (TBARS) indicated that the examined fraction might be effective antioxidant at broader concentration range, that is 1–5 and 25–50 μg/mL for hydroperoxides and TBARS generation, respectively. No cytotoxicity was observed at the concentration range of 1–50 μg/mL. Conclusions: Based on the obtained results, we suggest that antioxidant activity may additionally contribute to beneficial properties of K. daigremontiana-derived extracts.
Bryophyllum pinnatum (syn. Kalanchoe pinnata) is a succulent perennial plant native to Madagascar that was introduced in anthroposophic medicine in the early 20th century. In recent years, we conducted a large collaborative project to provide reliable data on the chemical composition, pharmacological properties, and clinical efficacy of Bryophyllum. Here, we comprehensively review the phytochemistry, as well as the pharmacological and clinical data. As to the pharmacology, special emphasis is given to properties related to the use in anthroposophic medicine as a treatment for "hyperactivity diseases", such as preterm labor, restlessness, and sleep disorders. Studies suggesting that B. pinnatum may become a new treatment option for overactive bladder syndrome are also reviewed. Tolerability is addressed, and toxicological data are discussed in conjunction with the presence of potentially toxic bufadienolides in Bryophyllum species. The few data available on two related species with medicinal uses, Bryophyllum daigremontianum and Bryophyllum delagoense, have also been included. Taken together, current data support the use of B. pinnatum for the mentioned indications, but further studies are needed to fully understand the modes of action, and to identify the pharmacologically active constituents. Georg Thieme Verlag KG Stuttgart · New York.
A bufadienolide (compound A2) was isolated from Kalanchoe Daigremontiana Hamet et Perr. by methanol extraction and purification by column chromatography. Its pharmacological effects were investigated. The force of contraction of rabbit papillary muscles and left guinea-pig atria was increased in a concentration dependent manner between 3 × 10−7 and 10−5 M. The effects were reversible upon washing. Rested state contractions were not increased and carbachol was ineffective in the presence of compound A2 in rabbit papillary muscle. Thus, cyclic AMP seems not to be involved in the mechanism of the positive inotropic effect. Concentrations of up to 10−5 M did not alter the spontaneous beating rate, functional refractory period or the contraction and relaxation time of right guinea-pig atria. Arrhythmias followed by tonic contracture occurred with concentrations above 10−5 M. Electrophysiological measurements in guinea-pig papillary muscle revealed a decrease in (18%) and overshoot (30%), associated with a shortening of action potential duration (14%) and increase in force of contraction (117%). Dog kidney Na+/K+-ATPase activity was inhibited with an IC50 of 1.4 × 10−7 M. Intravenous infusion of compound A2 at a rate of 20 µg/kg/min in anaesthetized guinea-pigs caused ventricular arrhythmias and death after accumulated doses of about 760 µg/kg and 860 µg/kg respectively. The results show that compound A2 has a pharmacological profile similar to that of digitalis glycosides.