Design and Optimization of Herbal Oral Liquid

8/14/2019 Design and Optimization of Herbal Oral Liquid

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DESIGN AND OPTIMIZATION OF HERBAL ORAL LIQUID


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1. INTRODUCTION

Medicinal plants constitute a source of raw materials for both traditional systems

of medicine (e.g. Ayurvedic, Chivuse, Unani, Homeopathy and Siddha) andmodern medicine. Now days plant materials are employed throughout theindustrialized and developing world as home remedies. Over the counter drugs,The ingredients for the pharmaceutical industry. As such they represent asubstantial proportion of the global drug market. Most rural populations,especially in the developing world, depend on medicinal herbs as their mainsource of primary health care. Although most medicinal herbs are not in theirnatural states fit for administration, preparations. Suitable for administration aremade according to pharmacopoeia directions. The therapeutic potential of aherbal drugs depends on its form whether part of a plant, or simple extract, orisolated active constituents, Herbal remedies consists of portions of plants or

unpurified plant extracts containing several constituents, which often worktogether synergistically.The herbal drug preparation in its entirely is regarded as the active

substance and the constituents are either of known therapeutic activity or arechemically defined substances or group of substance generally accepted tocontribute substantially to the therapeutic activity of the drug, phytochemicalscreening involves botanical identification, extraction with suitable solvents,purification, and characterization of the active constituents of pharmaceuticalimportance. Qualitative chemical examination employing different analyticaltechniques are conducted to detect and isolate the active constituents. Ingeneral, all medicines, whether they are synthetic or of plant origin, should fulfill

the basic requirements of being efficacious and safe, ultimate proof of these canonly be achieved by some form of clinical research. A defined and constantcomposition of the drug is therefore one of the most important prerequisites forany kind of clinical experiment.

Quality control for the efficacy and safety of herbal products is essential.The quality control of phytopharmaceuticals may be defined as the status of adrug which is determined either by identify, purity, content and other chemical,physical or biological properties, or by the manufacturing process. Comparedwith synthetic drugs, the criteria and approach for herbal drugs are much morecomplex.

Phytopharmaceuticals are always mixture of many constituents and aretherefore vary variable and difficult to characterize. The active principles inphytopharmaceuticals are not always known.

The quality criteria for herbal drug are based on a clear scientific definitionof the raw materials. Depending on the type of preparation, sensory properties,physical constants, moisture ash content, solvent residues and adulterationshave to be checked to prove identify and purity materials such as heavy metals,pesticide residues aflatoxins, and radioactivity, also need to be tested for. toprove the constant composition of herbal preparations appropriate analytical


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method shave to be applied and different concepts have to be used in order toestablish relevant criteria for uniformity.

In our country, one of the most appreciable efforts to brings out the herbalmonographs in hence with modern scientific testing has been doen by IDMA incoordination with the renowed Scientists in the field and herbal industry. The

pharmacopoeia fourses on microscopica analysis of herbas, apart from otherusual tets like physiochemical parometers and information like therapeuticcategory, pharmacology, safety and dosage form. In general, the monographscover almost all the distriable parmameters of quality assessment.

Parameters of quality assessment for standardization of herbal drugs areas follows:

Authentication

Foreign matter

Oraganoleptic evaluation

Macroscopy and microscopy

Volatile matter

Ash Value Extractive Value

Chromatographic profile

Marker component

Pesticide residue

Microbial Count.

Radioactive Contamination (mandatory, only for particular region).

In spite of a well established herbal system, its global acceptance is underquestion because of the stringent norms of the western countries, for example,recently in European countries our herbal products suffered from a major setbackbecause of the heavy metal toxicity, until now, the test for heavy metal was notmandatory in our country monographs, but this incidence has raised an alarmand our government has responded to it immediately and made heavy metal testmandatory for all herbal products and prescribed the limits for heavy metal.

Some specific quantitative parameters are as follows:

Volatile Content

Bitterness Value

Sweetening Index

Swelling Factor

Faming Index

Hemolytic Index.One of the best method of standardizing herbs and herbal formulations

based on modern scientific tools is chromatography. It not only helps inestablishing the correct botanical identify but also helps in regulating thechemical sanctity of herbs. One such technique involves marker compoundtesting and finger print analysis, which are used for the standardization of

Ashwagandha, Mentha, and Senna.


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Hence it is clear that incorporating modern methodology in traditionalmedicine is an uncharted field and will require correlation of traditionalknowledge with the latest Scientific knowledge, it can play a vital role instandardizing the herbs and herbal formulations more accurately and should beadopted by various laboratories and industrial houses to maintain the standard of

quality of the herbs.Therapeutic use of medicinal plants has gained considerable momentumin the world during the past decade. The oversees of synthetic drugs withimpurities resulting in higher incidence of adverse drug reactions in moreadvanced communities has motivated mankind to go back to nature for saferremedial. How ever, it should be ensured that commercial formulations based onmedicinal plants are safe effective and of standard quality.

In this process, the scientific knowledge of plants in medicinal use will beof great help to all those engaged in cultivating, harvesting, processing andpreparing the final products for human consumption.

Use of Stevia rebaudiona as natural sweetener and nutraceutical for the

development of some Ayurvedic properiatory medicines as chyawanprash,churna, arka, Oral liquids, Kayakalpa. Herbal granules and tablets needsstandardization or stability aspect. Study of the products for beneficial, harmful,toxic or neutral effect of stevia in such herbal products is must. Real timestability study of these products should be carried out by consideringstandardization parameters. The developed products are studied by applicableand critical parameters.

Modern technological advances have made it possible to control andregulate various aspects of processing during production so as to minimize oreven eliminate il effects of factors affecting shelf-life mentioned aboveTechnological advances also make it possible to control temperature and

humidity in the processing and manufacturing areas hereby building stability ofthe formulation during processing and paclling. Exposure to one or all of thefactors mentioned above which are determined to stability can be controlled,crreat advances in design and development of packaging technology hasdeveloped packaging materials, which have requisite barrier properties to buildstability of the formulations. Formulations need to be carefully studied to selectsuitable pack aging material so that the products are compatible with thematerials and retain properties changes in color, odor, physical structure, particlesize moisture, flowability, taste pH and other physiochemical parameters are theproperties, which give evidence of product retaining shell life.

Modern formulation technology also has mode it possible for use of inertexcipients like stabilizers and preservatives to improve and maintain the stabilityof the formulations.Real time studies:

Which the above would apply to studies done under acceleratedconditions, manufactures may also conduct real time studies by keeping the

packs of formulations at 300 20C C/65%. 155 RH for long periods andevaluate for them for per determined parameters. For such real time studies,samples may be evaluated at 0, 3, 6, 12, 18 and 24 months.


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Parameters to be evaluated:Following parameters which may be selected for the stability studies on

the formulations depending on the dosage forms, ingredients going into thesame, stabilizer and preservatives present absent and knowledge of formulationsand processing conditions. A thorough knowledge of the formulations would help

one to identify parameters that are likely to change or show changes due to illeffects of the factors affecting the stability of formulations.AppearanceColorOdor (off odor, bad odor or odor different from the natural odor of formulation)TasteParticle sizeFlow abilityViscosityClarityPH

Moisture contentSedimentationFlocculationEmulsion breakageFriabilityHardnessExtractive values (in selected solvents)Volatile matter contentFree fatty acids/acidityPeroxide valueMicrobiological parameters namely total viable count (TC) yeast and mold

counter (YMC) Coli form count and other pathogensSpecific parameters applicable to the formulation /dosage form


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2. OBJECTIVES

To design and optimize herbal formulations alternative to the modern

medicine. To overcome safety, efficacy and stability problems by critical parameters

associated with herbs.

To develop and standardize herbal formulation.

To provide a scientific data of final products that have significant impact oncommunity; this will provide a novel, efficacious formulation by consumercompliance


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3.PHARMACOGNOSTIC AND PHYTOCHEMICAL REVIEW

Ashwagandha(Withania somnifera)

Bramhi(Bacopa monniera)

Shankpusphi(Canscora decussate)

3.1Shankapushpi

Family Name: Gentianaceae

Botanical Name(s):Canscora Decussata

Popular Name(s):Sankh Pushpi, Shankhini, Kambumalini, Samkhapushpi,Sankaphuli

Parts Used: Entire plant and juice

Habitat:This herb is found in India and Burma

Chemical constituents: Drug is found to contain bitter substance and an oleo-resin. Two crystalline compounds have been isolated from the aqueous andalcoholic extracts of plant. Shankhpushpi is found to contain triterpenes, alkaloidsand xanthones.Xanthones are secondary metabolites commonly occurringin a few higher plant

families, fungi and lichen


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Description: Shankhapushpi is a morning glory like perennial herb. Thebranches of the Shankhapushpi are spread on the ground and can be more than30 cm long. The flowers are blue in color (5mm) and the leaves, which are ellipticin shape (2mm), are located at alternate positions with branches or flowers.

Uses:Primarily, Shankhapushpi is used as a brain tonic. Shankhapushpi is one of the

best and prominent natural medicines, which helps in improving memory. Whole

plant of Shankhapushpi is used in medical treatment. Various chemicalcompositions such as glucose, sucrose, glycosides, alkaloids and various acidsetc. are found in the plant. Shankhapushpi is also used as one of the mostimportant ingredient in treatment of disorders/syndromes such as hypertension,hypotension, anxiety neurosis, stresses etc.


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3.2 Ashwgandha

Family Name: Solanaceae

Botanical Name: Withania Somnifera

Common Name: Withania, Winter Cherry, Indian Winter Cherry, Indian Ginseng,Ashwagandha

Part Used: Roots, Leaves

Habitat: Cultivated throughout drier parts of india.Product offered: Roots

Chemical constituents: The major biochemical constituents of ashwagandaroot are steroidal alkaloids and steroidal lactonesin a class of constituents calledwithanolides.1 At present, 12 alkaloids, 35 withanolides, and severalsitoindosidesfrom this plant have been isolated and studied. A sitoindoside is a withanolidecontaining aglucose molecule at carbon 27. Much of ashwagandaspharmacological activity has been attributed to twomain withanolides, withaferin

A and withanolide D.


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withanolide D

Description:Ashwagandha is a small, woody shrub in the Solanaceae family that

grows about two feet in height.It can be found growing in Africa, theMediterranean, and India. As a result of this wide growing rangethere areconsiderable morphological and chemotypical variations in terms of localspecies. However, theprimary alkaloids of both the wild and the cultivatedspecies appear to be the same. The roots are the mainportion of the plant usedtherapeutically.The bright red fruit is harvested in the late fall and seeds are driedfor planting in the followingspring. The berries have been shown to have anemetic effect.

Uses:It is used as tonic, abortifacient, astringent, deobstruent, nervine,

aphrodisiac and sedative. It is official in Indian Pharmacoepeia. It is popularlyknown as Indian Ginseng. It gives vitality and vigour and helps in building greaterendurance. It has been used in diseases such as rheumatism, leprosy andarthritis. Used to treat general debility, arthritis, depression, chronic fatigue,insomnia, anxiety, depressed immunity, infertility and memory loss. It increasesthe iron content of blood.


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3.3Brahmi:

Family Name: Scrophuariaceae

Botanical Name: Bacopa monniera

Common Name: Bacopa, Babies tear, Bacopa monnieri, Hespestis monniera,Nirbrahmi, Indian Pennywort, Jalanevari and water hyssop are the other namesthat are used forBrahmi. In English it is commonly called as the 'Herb of Grace'.

Parts Used: Entire plant and juice

Habitat: Cultivated throughout drier parts of india.Chemical constituents: The pharmacological effects of Bacopa monniera areattributed to the presence of a number of biologically active compounds,including alkaloids, saponins and sterols. The compounds responsible for thememory enhancing effects of Bacopa monniera are triterpenoid saponins called"bacosides".


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The chemical structures of some of these compounds are as follows:

Description:Brahmi is the small creeping herb with the numerous branches. It grows

to a height of 2 -3 feet and its branches are 10 -35 cm long. It has oval shapedleaves that are 1-2 cm long and 3- 8 mm broad. Leaves are formed in pairs alongthe stems. Small- tubular, five petaled flowers are white- purple in colour. Its


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stem is soft, succulent, and hairy with the glands. Roots emerge out of thenodules and directly go to the soil. The fruit is oval and sharp at apex.

Uses: Brahmi is used in traditional Indian medicines for centuries. for thetreatment of bronchitis, chronic cough, asthma, hoarseness, arthritis,rheumatism, backache, fluid retention, blood cleanser, chronic skin conditions,constipation, hair loss, fevers, digestive problems, depression, mental andphysical fatigue and many more. It is used to treat all sorts of skin problems likeeczema, psoriasis, abscess and ulceration. It stimulates the growth of skin, hairand nails. Brahmi posses anticancer activity. It is taken to get relief from stressand anxiety. According to the Ayurveda Brahmi has an antioxidant property. Ithas been reported to reduce oxidation of fats in the blood stream, which is therisk factor for cardiovascular diseases. Brahmi is considered as the main

rejuvenating herb for the nerve and brain cells.Brahmi is rich in Vitamin C andcan be used in the salads, soups and sandwiches. Brahmi oil restores andpreserves the memory. In India it is given to the infants to boost memory powerintelligence


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MATERIALS AND METHODS:

MATERIALS:The authenticated crude material ofAshwgandha (Withania somnifera), Bramhi

(Bacopa monniera), and Shankpusphi (Canscora decussata) were procured fromAyurvedic Arkashala, Satara. All these are reported as nervine tonic andimmunomodulatory agents. These plants contain antioxidant principles,immunomodulatory agents used in many herbal products.

METHODS:Ashwgandha (Withania somnifera), Bramhi (Bacopa monniera),and

Shankpusphi (Canscora decussata).All these are reported as nervine tonicandimmunomodulatory agents.

These materials were powdered. Herbal oral liquid were obtained

by aqueous extraction of these crude materials with purified water at roomtemperature with occasional shaking for seven days, and then theaqueous extract was filtered. The filtrates were treated with lemon flavor,sodium saccharin and synthetic preservative. Formulated products wereoptimized using applicable physicochemical parameters.

Following parameters were studied for standardization of herbal oralliquids Applicable paramerters are:

General appearance

Colour

Odour

Taste

PH

Viscosity

Surface tension

Particle size

Specific gravity

Clarity or foreign particle determination

Critical parameters:

Antioxidant activity

Microbial determination

TLC profiles

HPTLC finger print

Heavy toxic metal ion determination

Pesticide residue


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Determination of pH:The pH values of herbal oral liquids were measueed by potentiometer.

For determination of pH glass electrode is incorporated in an ordinarypotentiometeric circuit and the off balance currents are amplified electronically so

that a milliammeter can be used to detect the balance point. The electricamplifier in this manner act as a null point indicator.Potentiometer is adjusted until the deflection is zero by using distil water. Formeasurement of unknown pH the potentiometer is first calibrated with a knownbuffer solution pH (4 and 90) and then measured the unknown pH.

Determination of Viscosity:Ostwald viscometer was used to determine the viscosity of all samples of herbaloral liquid.Clean the viscometer with warm chromic acid solution or organic

solvent. Mount the viscometer in a vertical position on suitable stand. Filled themeter the water and count the time required in second to flow the water betweenthe two marks. then filled with meter with the health drink and count the tine insecond required to flow between the two marks.Viscosity of the unknown healthdrink determine by using formula.

Viscosity = (2 x t2 / 1 x t1 ) 2

Where,2=Density of health drinks.t2 =Time taken for the health drinks1=Density of water.t1=Time taken for water2 =Viscosity of water


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Determination of surface tension:Herbal oral liquids were evaluated for surface tension, by stalagnometer.

Clean the stalagnometer with warm chromic acid solution or organic solvent.

Mount the stalagnomete in a vertical position on suitable stand. Filled the meterwith water and count number of drops in second between the two marks. thenfilled the meter with the health drink and count the number of drops in secondbetween the two marks.Surface tension of the herbal oral liquids were determine by using formula.

Surface tension = (2 x n1 / 1 x n2) 2

Where,2=Density of health drinks.

n2=Number of drops for the health drinks1=Density of water.n1=Number of drops for water2=surface tension of water

Determination of R. I.:Abbes refractometer was used to determine R. I. of Herbal oral liquids.

The drink is placed between the two prisms of the Abbes refractrometer. Adjustthe light and looked in to the eye piece. Adjust the cross and fine adjustmentknobs. Until the dividing line between light and dark halves of the visual fieldcoincides with the centre of the cross hairs. Then the refractive index is read.

Determination of specific gravity:Pcynometer is used to determine the specific gravity at 250C. It was

determined dividing the weight of sample (expressed in gms) by the weight ofwater (in gm).Clean the density bottle with chromic acid or organic solvent then distil water.Take weight of the empty density bottle with stopper. Filled the density bottle withwater close it by stopper and take the weight,Then replace the water with Herbaloral liquid take the weight of the density bottle with stopper. Density wasdetermined by using firmula

Density of Herbal oral liquid = weight of Herbal oral liquid / weight of water.


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5) Determination of clarity:Oral liquids were tested for the presence or absence of any particles by

clarity test apparatus.

Microbiological study:Different samples of herbal oral liquids were tested for their presence or

absence of Total viable aerobic count, Yeast and mold count, E Coli, salmonellaand other pathogens.

Total viable aerobic count by plate count Method:

For Bacteria:Petridishes of 9-10 cm in diameter were used. To one dish added a

mixture of 1ml of the pre treated material and about 15 ml of liquefied caseinsoybean digest agar at a temperature not exciding 450C. Alternatively, spread thepretreated material on the surface of the solidified medium in a Petridish. Ifnecessary, diluted the material as described above to obtain an expected colonycount of not more than 300. Prepare least two dish using the same dilution andincubate them at 30-350C for 5 days unless a more reliable count is obtained in a

shorter period of time. Counted the number of colonies formed and calculatedthe results using the plate with the largest number of colonies, up to a maximumof 300.

For fungi:Petridishes of 9-10 cm in diameter were used. To one dish added a

mixture of 1ml of the pre treated material and about 15 ml of liquefied sabour andglucose agar with antibiotics at a temperature not exceeding 450C Alternatively,


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spread the pretreated material on the surface of the solidified medium in aPetridish. If necessary diluted the pre treated material as described above toobtain an expected colony count of not more than 100. Prepared at least twodishes using the same dilution and incubated them at 20-250C for 5 days, unlessa more reliable count is obtained in a shorter period of time. 60 united the

number of colonies formed and calculate the results using the dish with not morethan 100 colonies.

Antioxidant activity:Antioxidant activity of herbal oral liquid was studied using different In vitro

chemical models as

DPPH assay:To 1ml extract of different concentrations, 1ml solution of 0.1mm of DPPH

(1, 1-diphenyl -2-picryl hydrazyl) was added. An equal amount of methods andDPPH solution sounded as control. After 20 min of incubation in the dark,absorbance was measured at 517 nm. The experiment was performed inreplicate and the percentage scavenging was calculated.

Iron Chelating activity:1ml of each extract was treated with an equivalent amount of reaction

mixture which contains 1ml , .05 5 0- phenan throline in treated compound wasincubated at ambient temperature for 10 min and the absorbance of same wasmeasured at 510 nm. The experiment was performed in triplicate.

Total antioxidant capacity:1 ml of extract of different concentrations was treated with 1 ml of reagent

solutions (0.6 M sulphuric acid, 28 mm sodium phosphate and 4 mm ammoniummolybalate) in eppendory tube. The tubes were capped and incubated in thermalblock at 950C for 90 min. After cooling to room temperature the absorbance was


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measured at 695 nm against blank. The ascorbic acid was used as standard.The experiment was performed in triplicate.

Scavenging of superoxide radial by riboflavin NBT system:The assay was based on the capacity of the sample to inhibit blue

formation by scavenging the superoxide radicals generated in the riboflavin NBTsystem. The recusttionmixture contains 50 mm phosphate buffer, pH 7.6, 20 gmriboflavin, 12 mm NBT. Reaction was started by illuminating the test samples ofthe extract. The absorbance was measured at 590 nm. Ascorbic acid was usedas positive control. The experiment was performed in triplicate.

Hydroxyl radical scavenging activity:Extracts of different concentrations were taken in different test tubes and

evaporated on water bath. To these, 1ml of fe EDTA, 0.5 ml of EDTA and 1 mlDM 50 were added and the reaction was initiated by adding 0.5 ml as wubic acidto each to the test tubes. Test tubes were capped tightly and heated on water

both at 80

0

C-90

0

C for 15 mm. then the reaction was terminated by addition of ice-cold TCA (17.5 % w/v) to the test tubes and kept aside for 5 min. Theformaldehyde formed was determined by adding 3 ml Nash reagent (75 gmammonium acute 3 gm glacial acetic acid, 2 ml acetyl acetone was mixed andraised to 1 liter with distilled water). This reaction mixture was kept aside for 15min for color development Intensity of yellow color formed was measuredspectrophotometrically at 412 nm against reagent blank. The experiment wasperformed in triplicate.

Scavenging of nitric oxide radical:Nitric oxide was generated from sodium nitroprusside and measured by

griss recuton as described previously 19, 20. Sodium nitroprusside(5 mm) inStandard phosphate buffer saline solution (0.025m, pH 7.4) was incubated with

different concentrations of (200 mg/ml, 300 mg, 500 mg/ml) of the hydroalcoholicbestrout dissolved in phosphate buffer saline (9.925 m, pH 7.4) and the tubeswere incubated at 250C for 5 hrs. Control experiments without test compoundsbut with equivalent amount of buffer were conducted in identifial maner. After 5hrs, 0.5 ml of solution was removed and diluted with 0.5 ml of grossiss reagent(1% nap thyl ethylene diamine dihydrochloride. The absorbance of chromophorefounded during diazotization of and its subsequent coupling with napthyl ethylenedemaine was read at 546 nm. The experiment was performed in triplicate.

Rapid screening for antioxidant compounds by using TLC:To make a semi quantitative visualization possible, Herbal oral liquid was

applied on a TLC plate and developed in solvent system consisting of ethylacetate formic acid, glacial acetic acid water (14.28 :1.42:1.42:2.85 v/v/v/v). Theplate was then dipped in a 0.2% solution of DPPH in ethanol. The yellow coloredspots on stationary phase are an indirect measure of antioxidant activity. Theexperiment was performed in triplicate.


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7) Qualitative Analysis:

For Arsenic:

1. Sample, when react with silver nitrate- No formation of yellow precipitateof silver arsenites soluble in ammonia solution and in nitric acid.

2. Sample, when react with solution of copper sulphate- - No formation ofgreen precipitate (scheelis green) which on boiling gives a red precipitateof cuprous oxides.

3. Sample, when react with silver nitrate- No formation of brownish redprecipitate of silver arsenate soluble in acids and in ammonia butinsoluble in acetic acid.

4. When nitric acid and excesses ammonium molybdate solution is added tothe sample- No yellow precipitate is obtained on boiling (Distraction fromarsenites which give no precipitate and from phosphates which giveprecipitate in cold or upon gentle warming).

5. Sample, when react with magnesia a mixture (a solution containing mgCl 2,NH4Cl and a little NH3)- No white precipitate which forms red silverarsenate upon treating with silver nitrate solution containing a few drop ofacetic acid (Distinction from phosphate).


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For Lead:

1. Sample, when treated with hydrochloric acid- No yields a white precipitatewhich is soluble in boiling water and is re-deposited as crystals when thesolution is cooled.

2. Sample, in the absence of very strong acid, when treated with hydrogen.Sulphide- No a black precipitate which is insoluble in dilute hydrochloricacid and in ammonium sulphide solution but is soluble in hot dilute nitricacid.

3. Sample, when treated with dilute sulphuric acid- No white precipitatewhich is almost insoluble in water, more nearly insoluble in dilute acetatesolution.

4. Sample, when treated with potassium iodide solution- No yellowprecipitate which dissolves when the mixture is boiled and is reprecipitateas glistening plates when the solution is loaded.

5. Sample, when treated with potassium chromate solution- No a yellow

precipitate which is readily soluble in sodium hydroxide solution and in hotnitric acid, sparingly soluble in dilute nitric acid, and insoluble in aceticacid.

6. Sample, when treated with potassium glyceride solution made alkalinewith dilute ammonia solution, and shaken with diphenyl thiocarbazonesolution- No a brick red colored lower layer.

For Mercury:-

1. Sample when treated with hydrogen sulphide- No formation of a blackprecipitate which is insoluble in ammonia sulphide solution and in boilingdilute nitric acid.

2. Sample, when treated with stannous chloride solution- No whiteprecipitate which rapidly becomes grey with an excess of the reagent

3. Sample, when treated with potassium iodide solution- No a scarletprecipitate which is soluble in excess of the reagent and in a considerablecub of the solution of the mercuric salt.

4. Sample, when treated with sodium hydroxide solutions- No a yellowprecipitate.

For chlorides:

1. Sample when treated with sulphuric acid and manganese dioxide Noblue color with potassium iodide and starch solution.

2. Sample when treated with silver nitrate solution No white cruddyprecipitate.


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For phosphate:

1. Sample when treated with silver nitrate solution No yellow precipitate2. Sample when treated with Magnesia mixture No white crystalline

precipitate of magnesium ammonium phosphate.

3. Sample when treated with large excess of ammonium molybate reagent No yellow crystalline precipitate of ammonium phosphomolybdate.4. Placed a drop of the acidic solution of sample upon evaluative filter paper

No blue stain is formed.

8) TLC Profile:-TLC plates were developed by using mobile phase as toluene: ethyl

acetate: Formic acid: ethanol (6: 4: 0.3: 0.4)Rf value was calculated by the ratio of the distance traveled by the spot to

the distance traveled by the solvent.For TLC silica gel G is used as a coating material. The TLC plate were preparedby appluiying silkica gel G on glass plate by poring method i.e. a measuredamoynt of slurry of silica gel is poured on glass plate which is kept on a levelsurface. The plate is then tipped back and forth to spread the salary uniformlyover the surface. Dried the plate at room temperature for thirty minutes thenactivate the plate at 110 oC for thirty minutes in an oven.

The sample of health drinks were applied by using capillary tubes then again drythe plate at room temperature. The plate is then placed in a developmentchamber. The bottom of the chamber is covered up to nearly one mm by thesolvent system. After solvent has transversed one half to two thirds the length ofthe plate. Plates were removed and dried at room temoerature. The position ofthe spots were determined by using iodine chamber. The Rf value of the spotswere determined by using formula.


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9) HPTLC Profile:Much of the crude materials used in herbal oral liquid contain Gallic acid

and quercetin as chemical markers. Therefore Gallic acid and quercetin were

used as standards for HPTLC determination.A simple, sensitive HPTLC method developed for the analysis of gallicacid, quercetin which present in herbal oral liquid formulation.

Apparatus used was Camag Linomat IV Automatic sampler applicator,Camag TLC Scanner III, ATS IV Soft waver.

Stationary Phase Pre coated silica Gel 60 F 254.Mobile phase

Sample Methanolic extract of oral liquid 30, 60, 90,120,150 and 180micro gm/ml was prepared by diluting with Methanol.

Standard. The same concentrations of Gallic acid and quercetin wereprepared.

Method 2Microliter of standard solution of mixture (Gallic acid and quercetin) was appliedon prey coated TLC. Silica gel G60 F254 plates using a Camag Linomat IV.

Automatic sample applicator. The plate was developed with a twin troughchamber to a distance of 10 cm. After removal from the chamber the plate wasdried in air for 15 min, was scanned and quantified at 540 nm using a Camag.TLC Scanner III. Data of peak area of each bond was recorded. Standard curvefor Gallic acid in the range of micro gram /ml and quercetin in the range ofmicrogram per ml was generated by plotting the peak area against concentrationof standards.

Developed oral liquid sample was extracted with methanol the extract was

filtered through Whattman filter paper no 41 and again the residue was washedwith 10ml of methanol. The extract and washing were transferred to a 100 mlvolumetric flash and the volume was made up to 100 ml with methanol. Fivemicroliter from the sample solution was spotted in triplicate on a prey coatedsilica gel g60 F254 TLC plates. The plate was developed as the same procedureand standard and the peak areas were recorded. Amount of Gallic acid,quercetin and stevioside in the foundation was estimated using the calibrationcurve for standards.

10) Sweetness potency:-Sweetness potency of oral liquids containing sweetener were evaluated

by take evaluation method.

Result and discussion:


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The result obtained indicating that the values of herbal oral liquid studied by allphysicochemical parameters are complies with the standard given in herbal and

Ayurvedic Formulary of India. It was less viscous liquid having pleasant odor,yellowish brown color with slightly sweet taste. Results are shown in Table no.1.This product showed significant antioxidant potential in comparison with std.

ascorbic acid. Qualitative analysis by different chemical tests gives an evidenceof absence of heavy toxic metal ions and pesticide residue. Total microbial count,fungal count, yeast and mould count were within limit as per WHO guidelines. E.coli, Salmonella, S. aureus, and other pathogens were found to be absent.

Table no. 1

Optimization of herbal oral liquid

Sr.

NoParameters Preparation1 Preparation2 Preparation3

1 Appearance Viscous Viscous Viscous

2 Color Brownish Brownish Brownish

3 Odor Agreeable Agreeable Agreeable

4 Taste Slightly sweet Slightly sweet Slightly sweet

5 Particle Size 125-180micron 125-180micron 125-180micron

6 Viscosity 1.15cp 1.28cp 1.32cp

7 ClarityFree from foreign

particle

Free from foreign

particle

Free from

foreign particle

8 pH 6.5 6.5 6.5

9 Sedimentation 2.5% /100ml 2.5% /100ml 2.5% /100ml

10 Total viable count / gm

Total aerobic count 4400 cfu / ml 4400 cfu / ml 4400 cfu / ml

Yeast and mould count < 10 cfu/ ml < 10 cfu/ ml < 10 cfu/ ml

E. coli _ _ _

Salmonella _ _ _

S. aureus _ _ _

11

Qualitative Analysis of

heavy metals

As _ _ _

Cd _ _ _

Hg _ _ _

Pb _ _ _

12 Pesticide Residues _ _ _


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Design and Optimization of Herbal Oral Liquid