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ORIGINAL ARTICLE
Year : 2015  |  Volume : 11  |  Issue : 44  |  Page : 434-438  

The isolation, characterization and preclinical studies of metal complex of Thespesia populnea for the potential peroxisome proliferator-activated receptors-γ agonist activity


1 Department of Pharmacognosy, Modern College of Pharmacy, Nigdi, India
2 Department of Pharmacognosy, Maratha Mitramandal College of Pharmacy, Kalewadi, Pune, Maharashtra, India
3 Department of Pharmaceutics, Mallareddy Institute of Pharmaceutical Sciences, Hyderabad, Telangana, India

Date of Web Publication16-Nov-2015

Correspondence Address:
Mohini Ashok Phanse
Department of Pharmacognosy, Modern College of Pharmacy, Nigdi, Pune, Maharashtra and PhD research scholar JNTUH, Hyderabad
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0973-1296.168984

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   Abstract 


Background: Diabetes mellitus is an international public health problem since ancient days. The condition is predominantly more severe in developing countries like India where, life is more sedentary due to the even changing lifestyles in this fast-paced global scenario. Thespesia populnea is widely used in the ayurvedic system of medicine for treatment of diabetes mellitus in India for years. The aim of this work is to explore the anti-diabetic activity of the isolated compound. Materials and Methods: The sesquiterpene isolated from hexane fraction of bark of T. populnea modified synthetically then identified by using analytical techniques such as electron paramagnetic resonance spectra for confirmation and the anti-diabetic activity was evaluated by anti-hyperglycemic, hypoglycemic potential. Result: In the present work, we have studied the anti-hyperglycemic and hypoglycemic activity of the vanadium complex in glucose loaded and normal animals were shown significantly decreased in plasma blood glucose level. The results derived from preclinical studies confirm the potential of new sesquiterpene. Conclusion: The findings could provide evidence regarding the anti-diabetic potential of T. populnea by lowering blood glucose level.

Keywords: Antihyperglycemic and hypoglycemic activity, diabetes mellitus, Thespesia populnea


How to cite this article:
Phanse MA, Patil MJ, Abbulu K. The isolation, characterization and preclinical studies of metal complex of Thespesia populnea for the potential peroxisome proliferator-activated receptors-γ agonist activity. Phcog Mag 2015;11, Suppl S3:434-8

How to cite this URL:
Phanse MA, Patil MJ, Abbulu K. The isolation, characterization and preclinical studies of metal complex of Thespesia populnea for the potential peroxisome proliferator-activated receptors-γ agonist activity. Phcog Mag [serial online] 2015 [cited 2019 Aug 19];11, Suppl S3:434-8. Available from: http://www.phcog.com/text.asp?2015/11/44/434/168984

SUMMARY

  • Thespesia populnea is widely used in the ayurvedic system of medicine for treatment of diabetes in India. Present study aimed to explore the anti diabetic potential of isolated compound. Isolation of sesquiterpene from hexane fraction of bark of Thespesia populnea and modified synthetically then authenticated by using analytical techniques such as electron paramagnetic resonance spectra for confirmation. The modified complex was further assessed for its anti diabetic property in glucose loaded rats. Vanadium complex demonstrated significant reduction in plasma blood glucose level in glucose loaded animals. The results derived from preclinical studies confirm the potential of new sesquiterpene. The present findings conclude that anti diabetic potential of Thespesia populnea could be due to lowering blood glucose level by acting on PPAR-? receptor.





   Introduction Top


Diabetes mellitus is a chronic metabolic disorder characterized by insulin resistance, hyperglycemia and associated with impaired lipid profile.[1] Considering the high incidence of diabetes mellitus, between 2010 and 2030, there will be a 69% raise in numbers of adults through diabetes in developing country and a 20% raise in developed countries. As a result, there is an increase in demand for a novel drug with a lesser side effect.[2] The main aim of organic and medicinal chemistry is to design, synthesize and produce molecules which act as important human curative agents. The compounds obtain from synthesis or plant source containing heterocyclic ring are of great significance receiving special attention as they belong to a class of compounds with proven utility in medicinal chemistry.[3]

In 1982, thiazolidinediones were intensively studied for their anti-hyperglycemic activity.[3] Thiazolidinediones are known to be insulin sensitizers and have been clinically used as anti-diabetic agents. The maleate of rosiglitazone, a medicine of the thiazolidinediones class, showed considerable clinical efficacy against diabetes mellitus.[1] Peroxisome proliferator-activated receptors (PPARs) include three isoforms: α, β/δ, and γ. PPARγ-is the most abundant isoform in adipose tissue, macrophages, monocytes, intestinal cells, skeletal muscle, and endothelium. It plays an important role in the regulation of insulin sensitivity, lipid metabolism, adipogenesis and glucose homeostasis.[4] The target of the thiazolidinediones has been identified as the PPARγ and the glucose-lowering activities of the thiazolidinediones were shown to be closely related to their PPARγ agonistic activity.[5] Still the herbal supplementation and other substitute medicine have gradually increased to use for management of the diabetic disorder.

Thespesia populnea of Malvaceae family is widely employed in the management of diseases. Various experimental findings showed that T. populnea bring wide spectrum of activities such as anti-bacterial activity, anti-hepatotoxic activity, anti-steroidogenic activity, anti-implantation activity, cytotoxicity and superoxide anion generation, anti-nociceptive and anti-inflammatory, anti-psoriatic activity, wound healing activity, dermatitis, anti-oxidant activity, Alzheimer's disease, anti-diabetic activity, synergistic activity, immunomodulatory activity, anti-inflammatory, α-amylase inhibitory activity, anti-ulcer activity, anti-oxidant and anti-inflammatory, memory-enhancing activity.[6],[7],[8],[9],[10],[11],[12],[13],[14],[15],[16],[17],[18],[19],[20],[21],[22],[23],[24],[25]

Vanadium is a trace mineral that is nearby present in various foods and may be essential in minute amounts in the body. High dose of vanadium has been experienced as an aid to controlling blood sugar level in the crowd with diabetes. It exists in oxidation states of − I, 0, +II, +III, +IV, and + V; the latter two are stable solution structures at physiological pH: Vanadyl (+IV) and vanadate (+V). The oral administrations of inorganic vanadium (IV, V) salts have revealed anti-diabetic activity. Vanadium compounds come first in an insulin-mimetic approach both in vitro and in vivo has been sound recognized. Both inorganic and organic vanadium compounds have been revealed to lower plasma glucose levels, raise peripheral glucose uptake, progress insulin sensitivity, decline plasma lipid levels and return to normal liver enzyme activities in an array of animal models of both type I and type II diabetes.[26] Vanadyl sulfate is one of the component vanadium's colorful forms and it is sometimes called a vanadium salt. Vanadyl sulfate is the form of preference for the dealing. Vanadyl sulfate management of diabetes is correlated to their anti-hyperglycemic, hyperinsulinemic and antihyperlipidemic effects.[27]

The objective of this study was to ameliorate the effect of T. populnea and its modified compound against diabetes mellitus. The findings could provide evidence to support the use of T. populnea as an anti-hyperglycemic and hypoglycemic plant. The modified compound characterized by electron paramagnetic resonance (EPR) spectra at low concentration also showed the anti-hyperglycemic and hypoglycemic effect.


   Materials and Methods Top


Drug and chemicals

The Glimepiride were purchased from Aventis Pharma Ltd., (Batch No: 20120701). Vanadyl sulfate were purchased from Loba Chemie Lab Reagent Pvt. Ltd., Biochemical estimation of plasma glucose was done by glucose oxidase/peroxidase (GOD/POD) method respectively using standard diagnostic kits from Biolabs India Ltd., India.

Plant material

The dry bark of T. populnea was collected from Hilly area of Dehu 15 km from Pune, Maharashtra, India. The plant material were authenticated by Botanical Survey of India with voucher specimen is conserved under reference number BSI/WRC/Herbarium/2011.

Preparation of hexane extract, isolation, and modification of isolated compound

The air dried bark and finely powdered bark was extracted by routine process with 90% hexane, then isolation of the sesquiterpene by using column chromatography with elution of chloroform: Methanol (8:2) solvent system and characterized. This isolated compound complexed with vanadium in alkaline condition with methanolic solutions vanadium sulfate and the respective ligand in 1:2 metal: Ligand. The reaction mixtures were maintained in alkaline condition and stirred 6 h at room temperature. The precipitating metal complexes were collected by filtration, washed with cold methanol.

Characterization by electron paramagnetic resonance spectra

The EPR spectra of the vanadium complex provided information on metal ion environment and the electron delocalization. The X-band EPR spectra of vanadium complex, recorded in DMSO glass at 77 K.

Experimental animals

Male Wistar rats weighing approximately 140–160 g were used. They were housed in an air-conditioned room (approximately 22°C ± 2°C and during the entire experiments). The animals were maintained with pelleted food while tap water was available ad libitum. All the animals were monitored carefully and maintained in accordance with the ethical recommendations of animal experimentation protocol (MCP/IAEC/46/2011).

Determination of LD50 values for vanadium complex

Albino mice of both sex and of 25–30 g weights were divided into six groups, each containing six animals for the purpose of determining the LD50 value of a vanadium metal complex. Each group was caged separately. Seven different doses of 1.75, 5.5, 17.5, 55, 175, 550, and 2000 mg/kg body weight were employed for each test drug. Each animal in every group was administered with a predetermined dose orally. After 24 h, the numbers of dead animals in groups were recorded. The data were tabulated. The toxicological effect was assessed on the basis of mortality, which was expressed as an LD50 value. In the groups with survived animals and in the groups with dead animals, the obtained percentages were corrected using the following formula:

  • Correction formula for 0% dead group ¼ 100 (0.25/n)
  • • Correction formula for 100% dead group = 100 [n – 0.25/n].

    Where, n represents the number of animals in the group.

    After correction, the percentages were converted into probit. The values thus obtained were plotted against log dose. The LD50 value was determined.[28],[29]

    Anti-hyperglycemic study in normal rats

    • The Wistar albino rats weighing 140–160 g of either sex were used. The overnight fasted animals were divided into groups (n = 6). The animals were received 10% glucose solution (2 g/kg, p.o.) 30 min after drug administration
    • The animals of Group I, II, III, IV, V, VI received (control) 1% gum acacia (1 ml/kg, p.o.), glimepride (0.09 mg/kg, p.o.), ligand vanadium (LVa) complex (5 mg/kg, p.o.), LVa complex (10 mg/kg, p.o, LVa complex (20 mg/kg, p.o.), vanadyl sulfate (0.2 mM/kg, p.o.), respectively.


    Biochemical analysis

    At the 0th, 30th, 90th, and 150th min (fixed interval of times) after the glucose load, blood was collected by puncturing retro-orbital plexus by using fine glass capillary, in tubes containing an anticoagulant. Plasma was separated by centrifugation. Plasma glucose levels were determined with a standard diagnostic kit by GOD/POD method[Figure 2].[30],[31]

    Hypoglycemic study in normal rats

    • The Wistar albino rats weighing 140–180 g were used. The overnight fasted animals were divided into groups (n = 6)
    • The animals of Group I, II, III, IV, V, and VI received 1% gum acacia (1 ml/kg, p.o.), glimepiride (0.09 mg/kg, p.o.), received LVa complex (5 mg/kg, p.o.), LVa complex (10 mg/kg, p.o.), LVa complex (20 mg/kg, p.o.), vanadium sulfate (0.2 mM/kg, p.o.).


    Biochemical analysis

    At the 0th, 60th, 120th, and 180th min after drug administration blood was collected by puncturing retro-orbital plexus by using fine glass capillary, in eppendorf tubes containing anti-coagulant. Plasma was separated by centrifugation. Plasma glucose levels were determined with a standard diagnostic kit by GOD/POD Method.[30],[31]


       Results Top


    Plant material

    The extraction methods by means of organic solvents are normally used for the pharmacological study. However, the extraction methods might not reflect the real characteristic of traditional Indian medicine; because most traditional Indian medicine remedies are prepared in the form of infusion, decoctions, and tinctures. Boiling water extraction and organic solvent extractions result in different phytochemical profile in the extracts. The present study used the solvent (hexane) extraction process to investigate the real active components that exist in the clinical active extract the plant constituent was isolated by routine process [Figure 1] and incorporated with vanadium sulfate which was thought for the T. populnea profound pharmacological activity.
    Figure 1: The structures of the compound identified in Thespesia populnea

    Click here to view


    Characterization by electron paramagnetic resonance spectra

    The four well-resolved metal hyperfines in the low field region are indicative of monomeric Vanadium complex gives conformation about the complex.

    Determination of LD50 values for vanadium complex

    The LD50 for the vanadium complex was found to be 100 mg/kg during a 24 h period were hardly perceptible.

    Anti-hyperglycemic study in normal rats

    During the anti-hyperglycemic study, treatment with vanadium complex showed a significant decrease in plasma glucose level at the dose of LVa-10 and LVa-20 ANOVA followed by Dunnett test. *P < 0.05, **P < 0.01 when compared with control group. Group treated with Vanadium complex shows dose-dependent anti-hyperglycemic activity [Figure 2].
    Figure 2: Effect of vanadium complex on plasma glucose in glucose loaded rats (anti--hyperglycemic effect). Values are expressed as mean ± standard error of the mean (n = 6), ANOVA followed by Dunnett test (*P < 0.05, **P < 0.01 when compared with control)

    Click here to view


    Hypoglycemic study in normal rats

    In the hypoglycemic studies, Group treated with Vanadium complex at the dose of LVa-10 and LVa-20 showed a significant decrease in plasma glucose level when compared with the control group [Figure 3].
    Figure 3: Effect of vanadium complex on plasma glucose in normal rats (hypoglycemic effect). Values are expressed as mean ± standard error of the mean (n = 6), when compared with control


    Click here to view



       Discussion Top


    The dose of a chemical that has been calculated to cause death in 50%, of a defined experimental animal is LD50. The effects of vanadium complex in p.o doses ranging between 10 mg/kg and 20 mg/kg.

    As expected, after starting the glucose tolerance test (anti-hyperglycemic study) in normal rats overloaded with glucose, the plasma glucose concentration was significantly increased when compared with the zero time of this group. Treatment with Vanadium complex showed a significant decrease in plasma glucose level when compared with control group. Pharmacological uses of vanadium consist of lower of triglycerides cholesterol, and glucose levels, diuretic, anti-carcinogenic effect, contraction of blood vessels, enhancement of oxygen affinity of hemoglobin and myoglobin.[32],[33],[34]

    The anti-hyperglycemic activity of vanadium complex in glucose loaded animals is likely to be mediated by an increase in insulin release from pancreatic β-cells or may be due to increased insulin sensitivity. It is also possible that such effects may also be the result of inhibition of the intestinal absorption of glucose or by stimulating glucose uptake and use in the peripheral tissues or by increasing insulin from pancreatic β-cells.

    In the subsequent hypoglycemic studies, it was observed that vanadium complex proved the significant and dose-dependent hypoglycemic effect in normoglycemic animals. This study possesses that Vanadium complex show significant glucose lowering effect in normal rats. The plants showed hypoglycemic activity through the stimulation of insulin release.[35]


       Conclusion Top


    From the above result it was concluded that these effects may also be the due to of inhibition of the intestinal absorption of glucose or by stimulating glucose uptake and use in the peripheral tissues or by increasing insulin from pancreatic β-cells by acting on PPAR-γ receptors. These results signify innovatively incorporated ligand benefits beyond their nutritional properties. Group treated with Vanadium complex shows dose dependent anti-hyperglycemic and hypoglycemic effect. Thus ligand is considered for developing into a potent anti-diabetic moiety.

    Acknowledgments

    Authors are indebted to Director JNTU, Hyderabad and Dr. P. D. Chaudhari Dean, Pune University and Principal, Modern College of Pharmacy, Pune for their constant guidance, motivation and being supportive during this course of investigation.

    Financial support and sponsorship

    Nil.

    Conflicts of interest

    There are no conflicts of interest.



     
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       Authors Top


    Mohini Ashok Phanse, Assistant professor, Department of Pharmacognosy, Modern College of Pharmacy, Nigdi, Pune, India. PhD research scholar at JNTU, Hyderabad, Telangana, India.







    Manohar Janardhan Patil, Principal, Department of Pharmacognosy, Maratha Mitramandal College of Pharmacy, Kalewadi, Pune, Maharashtra, India.







    Konde Abbulu, Principal, Department of Pharmaceutics, Mallareddy Institute of Pharmaceutical Sciences, Hyderabad, Telangana, India.


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