In vitro xanthine oxidase inhibitory activity of the fractions of Erythrina stricta Roxb.

Aim of the study

To assay the in vitro xanthine oxidase inhibitory activity of the various fractions of the hydromethanolic extract of the leaves of Erythrina stricta and to determine its enzyme inhibition mechanism.

Materials and methods

Xanthine oxidase inhibitory activity was assayed spectrophotometrically under aerobic conditions and the degree of enzyme inhibition was determined by measuring the increase in absorbance at 295 nm associated with uric acid formation. Enzyme kinetics was carried out using Lineweaver-Burk plots using xanthine as the substrate.

Results

Among the fractions tested, the chloroform fraction exhibited highest potency (IC₅₀ 21.2 ± 1.6 μg/ml) followed by the pet–ether (IC₅₀ 30.2 ± 2.2 μg/ml), ethyl acetate (IC₅₀ 44.9 ± 1.4 μg/ml) and residual (IC₅₀ 100 ± 3.3 μg/ml) fractions. The IC₅₀ value of allopurinol used, as the standard was 6.1 ± 0.3 μg/ml. Enzyme inhibition mechanism indicated that the mode of inhibition was of a mixed type.

Conclusion

These results suggest that the use of Erythrina stricta for the treatment of gout could be attributed to its xanthine oxidase inhibitory activity.     

α-Glucosidase inhibitory activity of selected Philippine plants

Ethnopharmacological relevance

Antidesma bunius Spreng. (Phyllantaceae), Averrhoa bilimbi L. (Oxalidaceae), Biophytum sensitivum (L.) DC. (Oxalidaceae), Ceriops tagal (Perr.) C.B. Rob. (Rhizophoraceae), Kyllinga monocephala Rottb. (Cyperaceae), and Rhizophora mucronata Lam. (Rhizophoraceae) are used as remedies to control diabetes. In the present study, these plants were screened for their potential α-glucosidase inhibitory activity.

Materials and methods

The 80% aqueous ethanolic extracts were screened for their α-glucosidase enzyme inhibitory activity using yeast alpha glucosidase enzyme.

Results

Except for A. bilimbi with IC₅₀ at 519.86±3.07, all manifested a significant enzyme inhibitory activity. R. mucronata manifested the highest activity with IC₅₀ at 0.08±1.82 μg mL⁻¹, followed by C. tagal with IC₅₀ at 0.85±1.46 μg mL⁻¹ and B. sensitivum with IC₅₀ at 2.24±1.58 μg mL⁻¹.

Conclusion

This is the first report on the α-glucosidase inhibitory effect of the six Philippine plants; thus, partly defining the mechanism on why these medicinal plants possess antidiabetic properties.     

MAO-A inhibitory activity of quercetin from Calluna vulgaris (L.) Hull.

Aim of the study

This study investigated MAO-A inhibitory activity of methanol extract of Calluna vulgaris (L.) Hull., which traditionally has been used as a nerve calming remedy.

Materials and methods

A methanolic extract of Calluna vulgaris was partitioned against heptane, ethyl acetate and water. The three fractions were tested in a photometric peroxidase linked MAO-A bioassay. The ethyl acetate phase showed the highest MAO-A inhibitory activity. Quercetin was isolated by VLC through bioassay-guided fractionation and purified by re-crystallisation. The structure was elucidated by LC–MS and ¹H NMR.

Results

The IC₅₀-value for MAO-A inhibition by quercetin was 18 ± 0.2 μM in an assay where the IC₅₀-value for MAO-A inhibition by clorgylin was 0.2 ± 0.02 μM.

Conclusion

The content of quercetin in Calluna vulgaris might explain the reported nerve calming effect of the plant.     

Ethnopharmacology guided screening of traditional Indian herbs for selective inhibition of Plasmodium specific lactate dehydrogenase

Ethnopharmacological relevance

Medicinal plants traditionally used to treat malaria can provide quality leads towards identifying novel anti-malarial drugs. Here we combined this approach with target based drug discovery and explored Plasmodium specific lactate dehydrogenase (LDH) inhibitory activity of 8 Indian plants which are ethnically used to treat malaria.

Methods

LDH from Indian Plasmodium falciparum and Plasmodium vivax strains, were cloned and expressed in Escherichia coli, followed by purification of recombinant enzymes (rPfLDH and rPvLDH respectively). Extracts of 8 plants in different organic and aqueous solvents, were screened for their inhibitory activity on rPfLDH, rPvLDH and mammalian LDHs. Phyllanthus amarus aqueous extract was further tested for in vitro parasiticidal activity.

Results

Aqueous extract of Phyllanthus amarus Schum. and Thonn. and chloroform extract of Murraya koenigii (L.) Spreng. exhibited profound and exclusive inhibitory effect on Plasmodium falciparum LDH (IC₅₀=11.2 μg/ml±0.4) and Plasmodium vivax LDH (IC₅₀=6.0 μg/ml±0.6) respectively. Moreover, Phyllanthus amarus aqueous extract also demonstrated antiplasmodial activity in vitro, on Chloroquine sensitive and resistant strains of Plasmodium falciparum (IC₅₀=7.1 μg/ml±0.5 and 6.9 μg/ml±0.7 respectively).

Conclusion

Target specific screening of traditional herbs used in malaria treatment has proffered Phyllanthus amarus and Murraya koenigii extracts as hits which can optimistically provide novel antimalarial drugs.     

Active constituents from Sophora japonica exhibiting cellular tyrosinase inhibition in human epidermal melanocytes

Aim of the Study

There is greater consumer awareness of plant-based skin-care products. Sophora japonica L. (Fabaceae) has been used traditionally as a hemostatic agent and also has skin-care and whitening benefits. The effect of the isolated active compounds of Sophora japonica L. (Fabaceae) that inhibits tyrosinase activity in human epidermal melanocytes (HEMn) was examined.

Materials and methods

We used the mushroom tyrosinase inhibitory assay to isolate active constituents from the extracts. The structures of these constituents were characterized by physical and spectroscopic analyses. Cellular tyrosinase kinetics were analyzed and showed by Lineweaver–Burk plot.

Results

A new compound, N-feruloyl-N′-cis-feruloyl-putrescine (8), together with four flavonoids and three putrescine derivatives were obtained after assay-guided isolation of S. japonica. In HEMn, compound 8 was minimally cytotoxic (cell viability >90% at 100 μM) and the IC₅₀ value for suppression of cellular tyrosinase activity was estimated as 85.0 μM. Zymography analysis demonstrated the compound's concentration-dependent effects and the kinetic analysis indicated the compound's mixed-inhibitory action.

Conclusions

We concluded that the new compound 8 is the most potent component of S. japonica yet discovered. Its pigment inhibition activity may be exploitable cosmetically.     

Protein tyrosine phosphatase 1B inhibitory by dammaranes from Vietnamese Giao-Co-Lam tea

Ethnopharmacological relevance

Gynostemma pentaphyllum (Thunb.) tea was used in Vietnamese folk medicine as anti-diabetic agent.

Aim of the study

This study was aimed to investigate the inhibitory activities of fractions and constituents isolated from Gynostemma pentaphyllum on protein tyrosine phosphatase 1B (PTP1B) since it has been proposed as a treatment therapy for type 2 diabetes and obesity.

Materials and methods

The 70% EtOH extract, CHCl₃ fraction, EtOAc fraction, BuOH fraction, and seven isolated dammarane triterpenes were evaluated for their inhibitory activity in protein phosphatase enzymes (PTP1B and VHR).

Results

CHCl₃-soluble fraction showed a dose-dependent inhibitory activity of the PTP1B enzyme with the IC₅₀ value of 30.5 μg/mL. Among seven tested compounds, compounds 6 showed the most potent PTP1B inhibitory activity with IC₅₀ value of 5.3 ± 0.4 μM compared to a range 15.7–28.5 μM for the other six compounds. The inhibition mode of 6 was competitive toward p-NPP with a K_i value of 2.8 μM.

Conclusion

These study results suggested that the PTP1B inhibitory activity of these dammaranes may enable this plant to play an important role in the treatment of diabetes.     

Insulin releasing and alpha-glucosidase inhibitory activity of ethyl acetate fraction of Acorus calamus in vitro and in vivo

Ethnopharmacological relevance

The radix of Acorus calamus L. (AC) is widely used in the therapy of diabetes in traditional folk medicine of America and Indonesia, and we previously reported the insulin sensitizing activity of the ethyl acetate fraction of AC (ACE).

Aim of the study

To investigate the insulin releasing and alpha-glucosidase inhibitory activity of ACE in vitro and in vivo.

Materials and methods

Insulin releasing and alpha-glucosidase inhibitory effects of different fractions from AC were detected in vitro using HIT-T15 cell line and alpha-glucosidase enzyme. Furthermore, effects of ACE orally on serum glucose were detected in fasted and glucose/amylum challenged normal mice.

Results

AC and ACE increased insulin secretion in HIT-T15 cells as gliclazide did. As in vivo results, ACE (400 and 800 mg/kg) significantly decreased fasting serum glucose, and suppressed the increase of blood glucose levels after 2 g/kg glucose loading in normal mice. In addition, ACE as a mixed-type inhibitor inhibited alpha-glucosidase activity in vitro with an IC₅₀ of 0.41 μg/ml, and 100 mg/kg of it clearly reduced the increase of blood glucose levels after 5 g/kg amylum loading in normal mice.

Conclusions

Apart from its insulin sensitizing effect, ACE may have hypoglycemic effects via mechanisms of insulin releasing and alpha-glucosidase inhibition, and thus improves postprandial hyperglycemia and cardiovascular complications.     

In vitro biological activities of niloticane,a new bioactive cassane diterpene from the bark of Acacia nilotica subsp. kraussiana

Ethnopharmacological relevance

Acacia nilotica subsp. kraussiana was reported in African traditional medicine for the treatment of various ailments. Isolation of an active compound in this study from the bark extract may lead to the validation of its efficiency as a traditional crude drug.

Aims of the study

This study aimed to isolate active compound(s) from an ethyl acetate bark extract of Acacia nilotica subsp. kraussiana and to investigate some of its biological activity.

Materials and methods

The isolation process was carried out using bioassay-guided fractionation. The isolated compound was tested for antibacterial activity using the micro-dilution assay; anti-inflammatory activity using the COX-1 and COX-2 assays and investigated for inhibitory effect against acetylcholinesterase using the microplate assay.

Results

A new bioactive compound was isolated and identified as a cassane diterpene, niloticane. Niloticane showed antibacterial activity against Gram-positive bacteria Bacillus subtilis and Staphylococcus aureus with MIC values of 4 and 8 μg/mL, respectively. With Gram-negative bacteria, niloticane showed weak activity. MIC values obtained were 16 and 33 μg/mL against Klebsiella pneumonia and Escherichia coli, respectively. In the cyclooxygenase test, niloticane possessed activity with IC₅₀ values of 28 and 210 μM against COX-1 and COX-2, respectively. IC₅₀ values observed with indomethacin (positive control) were 3.6 μM for COX-1 and 189 μM for COX-2. In the acetylcholinesterase test, niloticane showed anti-cholinesterase activity with an IC₅₀ value of 4 μM. IC₅₀ values obtained by the galanthamine (positive control) was 2.0 μM.

Conclusion

The results obtained support the traditional uses of the bark of Acacia nilotica subsp. kraussiana in African traditional medicine for the treatment of some ailments that relate to microbial diseases, inflammation and central nervous system disorders.     

Free radical scavenging activities and inhibition of inflammatory enzymes of phenolics isolated from Tripodanthus acutifolius

Ethnopharmacological relevance

Leaf extracts from Tripodanthus acutifolius (Ruiz and Pavón) Van Tieghem have long been used in argentinean traditional medicine as anti-inflammatory, however, there is no scientific evidence which supports this use in the literature.

Aim of the study

The present study was conducted to evaluate the ability of five phenolic compounds purified from infusion prepared from Tripodanthus acutifolius leaves to inhibit key enzymes in inflammatory processes. As anti-inflammatory compounds frequently possess free radical scavenging activities, purified substances were comparatively evaluated to asses their free radical scavenging properties. Genotoxic effects were also evaluated.

Materials and Methods

Compounds were evaluated on their ability to inhibit hyaluronidase and cyclooxygenase-2 (COX-2) activities to assess their anti-inflammatory capacities. Free radical scavenging activity was assessed by: 1,1-diphenyl-2-picryl-hydrazyl radical (DPPH), superoxide anion assay and the inhibition on lipid peroxidation. Genotoxicity was evaluated by Bacillus subtilis rec assay.

Results

Fractionation of Tripodanthus acutifolius infusion yielded a novel phenylbutanoid derivative (tripodantoside) and four known flavonoid glycosides (rutin, nicotiflorin, hyperoside and isoquercitrin). Flavonoids produced higher inhibition on hyluronidase activity (IC₅₀ ≈ 1.7 mM) than tripodantoside (IC₅₀ = 27.90 mM). A similar COX-2 inhibition activity was exerted by tripodantoside and monoglycosilated flavonoids (IC₅₀ ∼ 50 μM). Compounds were strong radical scavengers, with effective concentration 50 (EC₅₀) values for DPPH in the range of 2.7–6.3 μg/mL, and for superoxide anion in the range of 3.9–8.7 μg/mL. All compounds scavenged peroxyl radicals in the lipid peroxidation assay. The substances showed no genotoxic effects.

Conclusions

The anti-inflammatory effects, free radical scavenging activities and lack of genotoxicity of purified compounds may support the folk use of infusion from Tripodanthus acutifolius leaves as anti-inflammatory.