Quality Assessment Of Andrographis Paniculata (Burm. F.) Wall. Ex Nees WSR To Geographical Region.

Abstract


Introduction:
Andrographispaniculata (Burm.F.) Wall.Ex Nees (AP) also called Kalmegh or "King of Bitters" belongs to family Acanthaceae.It has been used for centuries in Asia to treat gastro-intestinal tract and upper respiratory infections, fever, herpes, sore throat, and a variety of other chronic and infectious diseases 1,2 .Indian Pharmacopoeia narrates that it is a predominant constituent of at least 26 Ayurvedic formulations.In Traditional Chinese Medicine (TCM), Andrographis is considered as the herb possessing an important "cold property" useful to treat the heat of body in fevers, and to dispel toxins from the body.In Scandinavian countries, it is commonly used to prevent and treat common colds 3 .Habitat: It grows abundantly in south-eastern Asia: India (andSri Lanka), Pakistan and Indonesia but it is cultivatedextensively in China and Thailand, the East and West Indies,and Mauritius.AP is normally grown from seeds ubiquitously in its native areas where it grows in pine, evergreen anddeciduous forest areas, and along roads and in villages 4 .

Collection of plant parts
Roots of selected plant were collected as per Guidelines of National Medicinal Plants Board.The roots of selected plants were collected in Ghrishmarutu i.e. in the month of May roots were collected with minimum required digging with use of suitable tools.Collected roots thoroughly washed and cut into appropriate sizes and dried under sunlight 5,6 .

Quality assessment of plant parts:
1 Identification and authentication Plants and collected plants parts were indentified and authenticated by experts, Central Research Facility, ASU Drug testing Lab.Shri.B.M. K. Ayu.College of KAHER`s Belagavi.Voucher no: CRF/auth/260-74/2016. Plants and their parts were identified and authenticated on the basis of pharmacopeia standards, floras and Dravyaguna text book. 2 Macroscopic study: The macroscopic study refers to the physical evaluation of the plant materials in terms of colour, odour, shape, surface, fracture, etc. Analysis was done at natural remedies Bangalore.The alcohol extract of collectedsampleswere applied band wise usingLinomat V applicator (CAMAG, Muttenz, Switzerland) on a commercial 20 cm × 10 cm precoated HPTLC plate Silica gel 60 F254 (Merck).The application conditions were: carrier gas, nitrogen; syringe delivery speed, 10 s/μL; application volume, 10 μL; bandwidth, 8 mm; space between two bands, 20 mm; distance from bottom, 10 mm. 15 ml of mobile phase consisting of with mobile phase Toluene:ethylacatate:methanol(7:3:1).The result was examined under UV 254 nm& 366 nm by using a UV viewer cabinet (CAMAG).

5) Assay of Total Tannins & Flavonoids by UV Spectroscopy 9 : a) Determination of Total Phenols
Total phenols content of plant was determined by with the Folin-Ciocalteucolorimetric method, Gallic acid was used as standard.

Brief protocol:
Gallic acid was used to make the calibration curve by dissolved it in methanol and then diluted to 6.25 -100 μg/ml of serial concentrations.Stock Solution of Extracts (1mg/ml) was also prepared with methanol.Reaction Solutions of 10ml contained: Sample extract stock solution / (Gallic acid standard) was mixed with 10 fold dilute Folin-Ciocalteureagentand 7.5% sodium carbonate.The tubeswere covered with Parafilm and allowed to stand for 30 minutes at room temperature before andthe absorbance was at read at 760 nm.Blank was prepared in similar way by replacing sample or standard with methanol.Sample and standard absorbance was measured at 760 nm with a Shimadzu UV-1800 spectrophotometer.Calibration curve using ABSORBANCE vs CONCENTRATION of gallic acid standard was prepared and the concentration of total flavonoid in the sample was determined by using a slope equation that was obtained from the standard graph and results for total phenols was expressed as mg of gallic acid equivalent /gm dried extract.

b)Determination of total flavonoids
Total flavonoid content of plant was determined by Aluminium chloride colorimetric method, quercetin was used as standard.

Brief protocol:
Quercetin was used to make the calibration curve by dissolved in methanol/Ethanol and then diluted to 6.25-200 μg/ml of serial concentrations.Stock Solution of Extracts(1mg/ml) was prepared with methanol/Ethanol.Reaction Solutions of 10 ml contained: Sample extract stock solution / (quercetin standard) to which Methanol, 10% Aluminium Chloride, 1MPotassium Acetate solution and distilled water and were added and mixed well.Sample blank was prepared in similar way by replacing Aluminium chloride with distilled water.Sample and standard absorbance was measured at 420 nm with a Shimadzu UV-1800 spectrophotometer.Calibration curve using ABSORBANCE vs CONCENTRATION of Quercetin standard was prepared and the concentration of total flavonoid in the sample was determined by using aslopeequation that was obtained from the standard graph and results for total flavonoids was expressed as mg of quercetin equivalent /gm dried extract.

DISCUSSION
In Ayurveda system of Medicine, kalmegh Andrographispaniculata (Burm.F.) Wall.Ex Nees is one of the main ingredients used in many formulations like Bhunimbadi Kashaya etc. Medicinal plant quality influenced due to collection of geographical area in respect of physico-chemical & chemical strength 1,2 .In this present study it reveals that the Kalmegh collected from the Belgaum region showedphysicochemical strength, HPTLC finger printing showed more separation of phytochemicals and assays of Total phenols &flavoinds comparative than Koppal & Kerala collected samples.

Conclusion:
The study reveals that the geographical collection required consider along with SOP of collection of medicinal plants.There is need to be focus on cultivation practice in respect of habitat of respective medicinal plants.