Description:BRIEF DESCRIPTION OF THE DRAWINGS
[0020] It is to be noted, however, that the appended drawings illustrate only typical embodiments of the present subject matter and are therefore not to be considered for limiting of its scope, for the invention may admit to other equally effective embodiments. The detailed description is described with reference to the accompanying figures. Some embodiments of system or methods in accordance with embodiments of the present subject matter are now described, by way of example, and with reference to the accompanying figures, in which:
[0021] Fig. 1 illustrates the illustrates the In vitro a-amylase inhibition of Different Parts of Cassia tora (CT-1, CT-2, CT-3) leaves and stem, in accordance with an embodiment of the present invention;
[0022] Fig. 2 illustrates the In vitro a-amylase Activity of BUG, BUK and BUK, in accordance with an embodiment of the present invention;
[0023] The figure depicts embodiments of the present subject matter for the purposes of illustration only. A person skilled in the art will easily recognize from the following description that alternative embodiments of the structures and methods illustrated herein may be employed without departing from the principles of the disclosure described herein.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0024] While the embodiments of the disclosure are subject to various modifications and alternative forms, specific embodiment thereof have been shown by way of example in the figures and will be described below. It should be understood, however, that it is not intended to limit the disclosure to the particular forms disclosed, but on the contrary, the disclosure is to cover all modifications, equivalents, and alternative falling within the scope of the disclosure.
[0025] The terms “comprises”, “comprising”, or any other variations thereof used in the disclosure, are intended to cover a non-exclusive inclusion, such that a device, system, assembly that comprises a list of components does not include only those components but may include other components not expressly listed or inherent to such system, or assembly, or device. In other words, one or more elements in a system or device proceeded by “comprises… a” does not, without more constraints, preclude the existence of other elements or additional elements in the system or device.
[0026] The present subject matter relates to provide a herbal pharmaceutical composition of Cassia tora, Betula utilis, Justicia adhatoda and Sonchus arvensis to treat a variety of cardiovascular diseases, treatment or improvement of vasodilation and diabetes.
[0027] Reference may be made to Figure 1 illustrating the In vitro a-amylase inhibition of Different Parts of Cassia tora (CT-1, CT-2, CT-3) leaves and stem, in accordance with an embodiment of the present invention;
[0028] Reference may be made to Figure 2 illustrating In vitro a-amylase Activity of BUG, BUK and BUK, in accordance with an embodiment of the present invention;
[0029] In accordance with an embodiment of the present invention relates to a herbal pharmaceutical composition to treat a variety of cardiovascular diseases, treatment or improvement of vasodilation and diabetes. The methodology omprises of collection of plants from different location and Extraction and fractionation of the plant materials using different solvents viz methanol, ethanol, hexane. ethyl acetate, chloroform and water. Partitioning of extracts was done according to nature of compounds viz. terpenes, phenolics, glycosides, alkaloids etc. Chromatographic analysis was done by UHPLC ESI MS-MS, GC-MS. All selected plant extracts were evaluated for invitro TPC (Total phenolic content) and TFC (Total flavonoid content).
[0030] Methodology:
[0031] Example 1: Determination of Total Phenolic Content Total Phenolic content was determined by the folin-ciocalteu method and calculated using (GAE) Gallic acid equivalent. 20 µl, 40 µl, 60 µl, 80µl, 100 µl (1 mg/ml) of methanol, hexane, ethyl acetate, chloroform and water extract of CT-1, CT-2 and CT-3, 0.5ml of the folin-ciocalteu (50%) reagent was added while mixing gently. After 2 min, 1 ml of sodium carbonate (20% solution) and 12.5 ml of distilled water was added. The content were mixed and allowed to stand for 2 h. The optical density of the samples was measured at 720 nm and total phenolic content were expressed as equivalent to Gallic acid.
[0032] Example 2: Determination of Total Flavonoid Content Total flavonoid content was determined by and calculated using (QE) Quercetin equivalent. 20, 40, 60, 80 and 100 µl of 1mg/ml stock solution of methanol, hexane, ethyl acetate, chloroform and water extracts from CT-1, CT-2 and CT-3,1.5 ml of the sodium nitrite (5%) reagent was added while mixing gently. Afterthat 0.15 ml aluminium chloride (10%), 1 ml of sodium hydroxide (1 N solution) was added. The content were mixed and allowed to stand. The optical density of the samples was measured at 415 nm and total flavonoid content was expressed as (1mg/ml) rutin. All plant extracts are evaluated for in vitro Antioxidant activity by DPPH assay.
[0033] Example 3: Determination of DPPH Radical Scavenging Activity DPPH radical-scavenging activity was determined by Shimada, Fujikawa, Yahara, and Nakamura (Shimada et al. 1992) method. Briefly, a 2ml of DPPH methanol solution was added to sample solution (1mg/ml) at different volume- 20. 40 µl, 60 µl, 80 µl and 100 µl. The mixture was shaken vigorously and allowed to keep at room temperature in the dark for 30 min. Then the absorbance was measured at 517 nm wavelength by spectrophotometer. Lower absorbance of the test sample showed highest free-radical scavenging activity. The scavenging activity of sample was expressed as 50% effective concentration (EC50), whichrepresented the concentrated of sample having 50% of DPPH radical-scavenging effect. The inhibition of the free radicals was calculated by the given formula. Quercetin was used as a standard. Inhibition % = Ac-As×100Ac * Ac=Absorbance of the control *As=Absorbance of the sample
[0034] Example 4: Biological activity: Cytotoxicity Assay (SRB Assay) A standard colorimetric SRB (sulforhodamine B) assay was used for the measurement of cell viability. (Wichtl 2004) (Arya et al. 2015). Briefly, 10,000- 20,000 cells (depending on the doubling time of each cell type) were seeded toeach well of 96-well plate in 5% serum containing growth medium and incubatedovernight in CO2 incubator at 37° C. The cytotoxic effects of the fractions were calculated as % inhibition in cell growth as per the formula [100-(Absorbance of treated cells/ Absorbance ofvehicle treated cells)] X 100. During initial screening, the samples showing equalto or more than 75% growth inhibition of cancer cells at 10µm concentration wereconsidered as =Hits‘ and further screened at 2-fold serial dilutions against cancercell lines to calculated their half maximal inhibitory concentration (IC50) value.Doxorubicin was used as a standard.
[0035] a- Amylase Inhibition: All plant extracts were evaluated for in vitro a- amylase inhibition. Given following method used same in all selected plants??a- Amylase inhibitory activity assay was performed by using chromogenic method adopted from Sigma–Aldrich was described by (Ali et al. 2006). Crude a- amylase was dissolved in ice-cold distilled water to make concentration of 0.5µg/mL solution. Starch (0.1% w/v) in 20 mM phosphate buffer (pH 6.9)containing 6.7 mM sodium chloride, was used as a substrate solution. Experimentswere performed in triplicate. Plant extract of 40 µl (2 mg/mL in DMSO), 160 µLof distilled water and 500 µL of enzyme were added to the test tube, all tubes wereincubated at 250 C for 10 min. After incubation 500 µL of starch solution wereadded to the test tubes and again incubated at 250 C for 10 min. Now 1 ml coloringreagent were added to the test tubes and incubated at 800C for 5 min., then thetubes were removed from the water bath and put it in the crushed ice and 9 mL of distilled water is added. Acarbose was used as a standard. The absorbance wasrecorded at 540 nm and calculated by the following formula:
[0036] Example 5: Essential oil by Gas Chromatography-Mass Spectrometry (GC-MS)Using Thermo Fisher TRACE GC ULTRA coupled with DSQ II MassSpectrometer instrument, GC-MS analysis was performed using a TR 50MS column (30m x 0.25mm ID x 0.25 µm, film thickness); carrier gas, helium;temperature programming, 5 min. delay for solvent, at 500C temperature, holdtime 5.0 min, rising at 40C/min to 2500C and finally held isothermally for 5 min.The injector temperature was 2300C and carrier flow was constant flow 1 ml/min,in split mode (1:50) with injector volume 1 µl. The ion source temperature was setat 2200C, transfer line temperature was 3000C, and the ionization of the samplecomponents was performed in EI mode at an ionization voltage of 70eV. Massrange was used from m/z 50 to 650 amu.
[0037] Compound IdentificationConstituents of the essential oil and fatty acid were identified by comparing theirRetention time, and Retention indices, using n-alkanes homologous series (C8-C20 hydrocarbons), under same experimental conditions, co-injection withstandard authentic samples and MS Library search (NIST and WILEY) (Adams2007), (Budzikiewicz et al. 1964) (McLafferty 1962). The volatile components were identified by comparing their linear RI with those from literature (KubeczkaInhibition activity (%) = Abs (control) - Abs (extract) x 100Abs (control)
[0038] Results:In the present investigation, Results indicated that the highest total content of five phenolic compounds was detected in leaves extract of Cassia tora. Investigation revealed the newer source of beneficial saturated and unsaturated fatty acid of significant value. Fatty acids were rich in Palmitic, linoleic and linolenic acid content which is beneficial for lowering body cholesterol if taken on daily basis for dietary supplements(Table 1). Fatty acid enriched with of palmitic, oleic and stearic acid that enhanced the nutritional value and beneficial to the overall health. Moreover, our study undoubtedly confirms that the FAME extract of Cassia tora leaves and stem leaves contain higher relative percentage of the above mentioned fatty acids that has potential antibacterial principle for clinical application.(Figure 1)
[0039] Table 1: The transitions and optimized compound dependent MRM parameters: declustering potential (DP), entrance potential (EP), collision energy(CE) and cell exit potential (CXP) of 5 analytes

[0040] The present investigation revealed the newer source of beneficial saturated and unsaturated fatty acid of significant value from Betula utilis. The fatty acid composition of BUG, BUK and BUS bark has been analyzed by GCMS. Twenty two fatty acids were identified in BUG, BUK and BUS representing 76.65%, 64.57% and 69.75% respectively(table-2). The major fatty acids found were Palmitic acid, Linoleic acid, Linolenic acid, oleic acid. Fatty acids were rich in Palmitic, Linoleic and Oleic acid content which is beneficial for lowering body cholesterol if taken on daily basis for dietary supplements. The highest occurrence of Palmitic, Oleic and Linoleic acid increases the nutritional value and adds to the overall health benefits. Moreover, our study undoubtedly confirmed that Betula utilis bark contain higher relative percentage of the above mentioned fatty acids that has potential activities for clinical application (Figure 2).
[0041] Table 2: Fatty acid composition from the bark of BUG, BUK and BUS

[0042] Present study evaluated that Justicia adhatoda leaves essential oil contains an appreciable amount of Phytol (3,7,11,15-tetramethylhexadec-2-en-1-ol) which is a diterpene, and already reported to inhibit the growth of Staphylococcus aureus and to block the teratogenic effects of retinol, thus the essential oil may find extensive applications for antimicrobial activity. The presence of beneficiary fatty acids was richer in extract and may have significant value for various activities and also play an important role in pharmaceutical industries (Table 3).
[0043] Table 3: Fatty acid composition from the leaves of JA-L and JA-K

[0044] Sonchus arvensis root essential oil composition and its antimicrobial and antioxidant activities have been reported for the first time. The essential oil of Sonchus arvensis contains an appreciable amount of Dithiaanthracene. The high occurrence of palmitic, oleic and stearic acid enhanced the nutritional value and significant to overall health benefits of oil. Oil and fatty acid values analyzed meet the set standards and are significant in terms of nutritional value. The present investigation revealed that the Fatty acids from the root of Sonchus arvensis can be used in various pharmaceutical products as it contains different bioactive fatty acids.
[0045] Although embodiments for the present subject matter have been described in language specific to structural features, it is to be understood that the present subject matter is not necessarily limited to the specific features described. Rather, the specific features and methods are disclosed as embodiments for the present subject matter. Numerous modifications and adaptations of the system/component of the present invention will be apparent to those skilled in the art, and thus it is intended by the appended claims to cover all such modifications and adaptations which fall within the scope of the present subject matter.

Claims:We Claim:
1. A herbal pharmaceutical composition for prevention or treatment of metabolic disordersr comprising Cassia tora, Betula utilis, Justicia adhatoda and Sonchus arvensis extract as active ingredients..
2. The herbal composition as claimed in claim 1, wherein the composition comprises:
an extract of Cassia tora (leaves and stem);
an extract of Betula utilis (bark);
an extract of Justicia adhatoda (leaves); and
an extract of Sonchus arvensis (leaves)
3. The herbal composition as claimed in claim 1, wherein the plant parts are extracted with methanol by cold extraction process.
4. The herbal composition as claimed in claim 1, wherein extract of Cassia tora is extracted with 300 mL of petroleum ether (40-600C) in Soxhlet extraction for 10 hrs.
5. The herbal composition as claimed in claim 1, wherein the FAME extract of Cassia tora leaves and stem leaves contain higher percentage of fatty acids