Compounds isolated from Harpalyce brasiliana Benth and their pharmacological properties

Harpalyce brasiliana Benth, known as snakeroot, is one of the most popular herbal medicines against snakebite in South America. A hydroalcoholic solution is traditionally prepared from the roots of H. brasiliana. In the last two decades, understanding the pharmacological properties and the possible medicinal applications of H. brasiliana has increased considerably. H. brasiliana has antivenom activity and anticarcinogenic, antimicrobial and antioxidant properties. Various in vitro and in vivo studies have shown H. brasiliana’s diverse biological properties and its potential for disease treatment. The different biological effects of this plant may be attributable to the presence of secondary active metabolites such as pterocarpans, triterpenoids, chalcones and flavonoids. This overview presents different aspects of this plant and the pharmacological properties of its compounds through a review of the available literature. The results support the use of H. brasiliana in the treatment of snakebite and its potential for treatment of other diseases in folk medicine.


INTRODUCTION
Harpalyce brasiliana Benth (family Leguminosae), commonly known as snakeroot, is a shrub used in South America, particularly in northeastern Brazil to treat snake bite poisoning.The local people have reported that hydroalcoholic solution from the roots of H. brasiliana is an antidote against snakebite (Silva et al., 1999).Much of the activity of H. brasiliana is because of pterocarpans, which are major compounds present in the plant.Pterocarpans play an important role as phytoalexins and have significant pharmacological applications.Pterocarpans are present in the roots and leaves (Jiménez-González et al., 2008).
Here, an overview of the available literature on the main properties of this plant and the compounds isolated from this plant has been presented.

PTEROCARPANS
Pterocarpans are phytoalexins, that is substances produced de novo by plants to provide resistance against stress factors such as microbial infections or stress of abiotic origin.The second largest class of natural isoflavonoids found in the Leguminosae family is characterized by the presence of a tetracyclic system of benzofuranbenzopyran rings with two chiral centers in the 6a and 11a positions in H. brasiliana (Jiménez-González et al., 2008).All natural pterocarpans have a junction of cis-fused rings.Computational studies have shown that the trans isomers are energetically unfavorable, which explains the reason for all pterocarpans of natural origin possessing the 6a, 11a-cis configuration (Van Aardt et al., 2001).
The main bioactivities of pterocarpans (Table 2) included anti-inflammatory, antifungal, antimicrobial, antiparasitic, antimitotic, anticarcinogenic and antiophidic activities (Nakagawa et al., 1982;Jiménez-González et al., 2008;Zhou et al., 2009;Harinantenaina et al., 2010;Dey and De, 2012).Cabenegrins A-I and A-II were isolated initially by Nakagawa et al. (1982) from "Específico Pessôa," a northeast Brazilian folk medicine used against snakebite and spider venom.According to the authors, it was a hydroethanolic extract from the root of the plant popularly known as "black head".Cabenegrin A-I and cabenegrin A-II showed antiophidic activity at a dose of 2.0 to 2.8 mg/kg in mice and at a dose of 1.0 mg/kg in beagle dogs previously envenomed using the venom of Bothrops atrox (Viperidae).Further, on the basis of spectroscopic analysis of cabenegrins, other pterocarpans were obtained by synthesis.The compound 4′dehydroxycabenegrin A-I obtained by synthesis showed antimyotoxic, antiproteolytic and PLA2 inhibitor activities against Bothrops jararacussu (Viperidae) crude venom in vitro (Ishiguro et al., 1982;Silva et al., 2004).
The pterocarpan medicarpin is naturally found in dietary legumes and has also been synthesized for pharmacological studies.Studies on rat liver microsomes showed that medicarpin can inhibit mixed-function oxidase (P450) activity, particularly the cytochrome P-450-dependent aryl hydrocarbon hydroxylase (AHH) and ethoxycoumarin deethylase (ECD) activities (Friedman et al., 1985).Recent studies have shown that medicarpin affects osteoclastogenesis.Tyagi et al. (2010) showed that medicarpin at a dose of 10 -10 mol/L suppresses osteoclastogenesis in bone marrow cells and induces apoptosis of mature osteoclasts in ovariectomized mice.The mechanism of action appeared to be independent of estrogen receptor activation.Medicarpin acted as an estrogen receptor agonist; however it showed no uterine estrogenicity.
Medicarpin stimulates osteoblast differentiation and promotes achievement of peak bone mass in rats (Bhargavan et al., 2012).
Various studies have shown that maackiain inhibits different forms of aryl hydrocarbon hydroxylase in human and rat livers (Gelboin et al., 1981), besides its inhibitory effect on the growth of human lymphoblastoid cell line I (Skinnider and Stoessl, 1986) and induction of apoptosis in the DNA of human promyelocytic leukemia HL-60 cells (Aratanechemuge et al., 2004).Leiocarpin C, isolated from the stem bark of the Goniothalamus leiocarpus (Annonaceae), also possesses cytotoxic activity against several human tumor cell lines (Yadav et al., 2008).Studies on three human cells lines, namely HL-60 (leukemia cells), HCT-8 (colon cancer cells) and MDA-MB-435 (melanoma cells) showed that leiocarpin is cytotoxic and its half-maximal inhibitory concentration (IC 50 ) values are 5.5, 6.9 and 13.7 µg/ml, respectively (Militão et al., 2007).Studies on understanding the synthesis of this compound have been reported (Yadav et al., 2008;Krishna and Alivelu, 2011).

TRITERPENOIDS
Triterpenoids are natural products, with thirty carbons, derived from a squalene skeleton or enzymatic reaction by cyclization products of squalene, oxidosqualene and bis-oxidosqualene.They are found either in the acyclic or cyclic forms and tetracyclic and pentacyclic triterpenoids the most common forms of triterpenoids (Xu et al., 2004).Triterpenoids from natural origin and their synthetic derivatives are pharmacologically active (Table 2) and have anti-inflammatory and anticarcinogenic activities (Bishayee et al., 2011).The triterpenoid 3β-hydroxy-20(29)-lupen-28-oic acid, betulinic acid inhibits the growth of multiple tumors, including human melanomas.In an experimental model of prostate cancer, betulinic acid treatment decreased the growth of the cancer cells and its IC 50 value was 1 to 5 µmol/L (Chintharlapalli et al., 2007).In addition, the anticarcinogenic effect of betulinic acid on colon cancer cells has also been investigated.The results showed that the betulinic acid at a concentration of ≥ 5 µmol/L inhibited the growth of cancer cells and down-regulated the specificity protein (Sp) transcription factors (Chintharlapalli et al., 2011).

CHALCONES
Chalcones, 1,3-diphenyl-2-propenones, considered as precursors of flavonoids and isoflavonoids, are also known as benzalacetophenone or benzylidene acetophenone.They play an important role in plants for the pigmentation flowers, and act against pathogens and insects (Batovska and Todorova, 2010).Studies with chalcones and their analogues have shown several biological activities such as anti-inflammatory, antimicrobial, antioxidant and anticancer activities (Table 2) (Hijova, 2006;Perjési and Rozmer, 2011).A previous study showed that diets containing 500 ppm of chalcone and 2hydroxychalcone showed an inhibitory effect on oral carcinogenesis induced using 4-nitroquinoline-1-oxide in F344 rats (Makita et al., 1996).Another study showed that chalcones had a preventive effect on diabetic complications.The effect of several chalcones at a dose of 25 mg/kg on glycogen contents in rats showed marked inhibition of aldose reductase, which affected the rate of glycogenolysis by decreasing the glycogen content of the liver (Jamal et al., 2009).These data indicate the importance of chalcones and their analogues as an emerging class of preventive and therapeutic agents for diabetes.

FLAVONOIDS
Flavonoids are a class of polyphenolic compounds synthesized in plants.In general, flavonoids are compounds characterized by three rings; two aromatic rings linked to one heterocyclic ring.Flavonoids can be divided into flavonols, flavones, flavanones, isoflavonoids, catechins and anthocyanins (Hollman and Katan, 1997;Peterson and Dwyer, 1998).These compounds generally are considered safe because of favorable pharmacological activity and low toxicity (Table 2).Many compounds possess anti-inflammatory, antioxidant, anticarcinogenic and antiallergenic activities (Khairullina et al., 2010).In addition, the antiproliferative effect of the analogue 4′-chloroflavanone was observed in MCF-7 and MDA-MB-453 cells.The synthetic flavanone at a concentration of > 50 µmol/L inhibits cell proliferation through disruption of the G1/S phase and by induction of apoptosis (Choi et al., 2010).Additional biological activities were described for compounds belonging to the class of flavonoids.Recent studies with flavanone compounds showed antihyperglycemic action and an effect on bone metabolism.In an experimental model of diabetes, the flavanone naringenin (50 mg/kg) significantly decreased the levels of fasting blood glucose and glycosylated hemoglobin and increased the serum insulin levels.The same study showed an increase in the activities of the pancreatic enzymatic antioxidants and plasma non-enzymatic antioxidants (Annadurai et al., 2012).Another study showed favorable effects of naringenin and hesperidin on bone metabolism.Naringenin 0.5%, hesperidin 0.5% and naringenin 0.25% + hesperidin 0.25% were administered to male gonadintact senescent rats, and systemic parameters of bone metabolism were evaluated.The results showed that the flavanones exerted a protective effect on the bone in senescent male rats through inhibition of bone resorption (Habauzit et al., 2011).
In vitro assays showed that the antioxidant capacity of naringenin was higher than that of its glycoside naringin (Cavia-Saiz et al., 2010).In addition, a study on lipid peroxidation in mouse liver homogenates showed that both glycosides naringin and hesperidin had no antioxidant activity.On the other hand, quercetin at a concentration of 1 µmol/L showed antioxidant action (Day et al., 2000).Quercetin, the most abundant flavonoid, is recognized for its antioxidant action; however, several studies have also shown that quercetin has prooxidant activity (Constantin and Bracht, 2008).Moreover, similar to other flavonoids, quercetin showed reversible inhibition of butyrylcholinesterase, an enzyme linked to scavenging (Katalinić et al., 2010).

CONCLUSION
Plants extracts have been used for the treatment of several human diseases.Although few studies have been performed using H. brasiliana; studies show that this plant represents a particularly rich source of pharmacologically active compounds and possesses more than one bioactivity.This plant has anti-inflammatory, antivenom, antimicrobial, anticarcinogenic and antioxidant activities.Finally, further studies should be performed to identify the clinical applications of this plant for the benefit of mankind.

Table 2 .
Compounds and their pharmacological activities