Assay Of Boric Acid 4204o

1. ASSAY OF BORIC ACID AIM: To perform the assay of Boric Acid. APPARATUS: Volumetric flask, conical flask, Measuring cylinder and Burette. CHEMICALS: Glycerin, NaoH solution, Boric acid and Phenolphthalein. PRINCIPLE: Boric acid is a local infective. It is very weak acid and hence it can be titrated with glycerin, glyceryl-boric acid is formed which behaves as a strong acid and can be titrated against sodium hydroxide using phenolphthalein indicator. PREPARATION OF REAGENTS: PREPARATION OF 1M NaoH: 4gms of NaoH was accurately weighed and taken in a volumetric flask and dissolved in water and make up to 100 with distilled water. STANDARDIZATION OF 1M NaoH: 5gms of potassium hydrogen phthalate is weighed accurately and taken in conical flask and dissolved in 20 ml of distilled water, phenolphthalein indicator is added and titrated against with 1M NaoH. ASSAY: 1gm of boric acid is taken in conical flask and dissolved in distilled water to it add glycerin and phenolphthalein indicator titrated against with1M NaoH until a faint pink colour was produced.

REPORT:

2. ASSAY OF ASPIRIN

AIM: To perform the assay of Aspirin. APPARATUS: Volumetric flask, conical flask, Measuring cylinder and Burette. CHEMICALS: NaoH solution, Aspirin and Phenolphthalein, sodium carbonate, Methyl red solution. PRINCIPLE: Assay of aspirin is acid base titration using phenolphthalein indicator. The amount of aspirin is determined by neutralizing sample solution with excess standard NaoH solution to acetoxy group. Cooling and back titrated with standard acid using phenol red as indicator. PREPARATION OF REAGENTS: PREPARATION of 1M NaoH: 4gms of NaoH was accurately weighed and taken in a volumetric flask and dissolved in water and make up to 100 with distilled water. STANDARDIZATION of 1M NaoH: 5gms of potassium hydrogen phthalate is weighed accurately and taken in conical flask and dissolved in 20 ml of distilled water, phenolphthalein indicator is added and titrated against with 1M NaoH. PREPARATION OF 0.5M HCL:-4.25ml of HCL was diluted with water and the volume was made up to 1000ml. STANDARDIZATION OF 0.5M HCL:-0.75gms of anhydrous sodium carbonate is weighed taken in conical flask and dissolved with 100 ml of water titrated with 0.5M HCL using methyl red solution as indicator. ASSAY: 1.5gm of aspirin is weighed taken in conical flask and dissolved in ethanol to it add 50 ml of NaoH boiled for 10mins. The solution was cooled and titrated against 0.5M HCL using phenol red as indicator.

REPORT:

3. STANDARDISATION OF DISODIUM EDETATE AIM: To perform the standardization of disodium edetate. APPARATUS: Conical flask, burette, pipette, volumetric flask. CHEMICALS: Disodium edetate, zinc, bromine, water, NaOH, ammonium buffer. PRINCIPLE: Chemically disodium edetate is salt of disodium ethylene diamine tetra acetate. It is pharmaceutical acid. It is used as chelating agent in metal poisoning. As it is not a primary standard, it is standardized by using a primary standard. Zinc chloride solution which is prepared from weighed amount of granulated zinc. PROCEDURE: Weigh accurately 0.125gms of granulated zinc dissolved in minimum volume of HCl and bromine water (0.2ml) to ensure oxidation of iron impurity to iron which forms a much less stable edetate complex than iron. Gently boil to remove excess bromine cool and add NaOH (2M) until it become neutral, it was diluted to 250ml and ammonia buffer PH (10) was added until the precipitate was just dissolved and then 5ml in excess was added. Then a mixture of mordant black-II and NaCl was added which acts as an indicator.It was then titrated with 0.05M disodium edetate solution until green colour was appeared. Each ml of 0.05M EDTA ≡ 0.003269gms of zinc.

REPORT: The molarity of given disodium edetate was found to be

4. ASSAY OF CALCIUM GLUCONATE

AIM: To perform the assay of calcium gluconate and repeat its percentage purity. APPARATUS: Conical flask, pipette, measuring cylinder. CHEMICALS: Disodium edetate, calcium gluconate, warm water, magnesium sulphate, strong ammonia, mordant black-II. PRINCIPLE: Calcium gluconate is used as calcium replenisher. Chemically it is calcium Dgluconate mono hydrate. Estimation of calcium gluconate is done by disodium edetate. PROCEDURE: Standardization of disodium edetate: 0.125gms of granulated zinc was accurately weighed and dissolved in few ml of HCl.To this 0.2ml of bromine water was added.solution was gently boiled to remove excess of bromine.Solution was cooled and NaOH was added until solution becomes neutral.dilute to 250ml Ammonia buffer of PH 10 was added until the precipitate was just dissolved, 5ml excess was added.Then a mixture of mordant black-II and NaCl was added as indicator and was titrated with 0.05M disodium edetate until the green colour appears. ASSAY: Drug equivalent to 0.5gms was taken and dissolved in 50ml of warm water. The solution was cooled and 5ml of 0.05M MgSO 4 and 10ml of NH3 was added. The resultant mixture was then titrated with 0.5M EDTA using mordant black-II as indicator. Each ml of 0.05M EDTA ≡ 0.02242gms of calcium gluconate.

REPORT: Percentage purity of calcium gluconate was found to be

5. ASSAY OF FEROUS SULPHATE

AIM: To perform the assay of given ferrous sulphate. APPARATUS: Conical flask, pipette, measuring cylinder. CHEMICALS: Ferrous sulphate, 0.1N KMno4, Oxalic acid, Sulphuric acid. PRINCIPLE: Ferrous sulphate is an example of haematinic agent and also reducing agent. It is an redox titration.KMno4 is an powerful oxidizing agent in the presence of dilute H2SO4.During the titration ferrous sulphate is oxidized to ferric sulphate. As soon as the oxidation of ferrous sulphate is completed addition of a drop of KMno4 produces pink color which indicates end point. PREPARATION OF REAGENTS: PREPARATION OF 0.1NKMno4: 0.31gms of KMno4 was accurately weighed and taken in a volumetric flask and dissolved in water and make up to 100 with distilled water. PREPARATION OF DILUTE SULPHURIC ACID:- 5.7ml of 10%w/w of sulphuric acid was diluted to 100 ml. PREPARATION OF 0.1N OXALIC ACID:- 0.63gms of oxalic acid was diluted with water and the volume was made up to 100ml. STANDARDIZATION OF 0.5M HCL:-10ml of 0.1N Oxalic acid was taken and to this add 10 ml of sulphuric acid .The contents were warmed to 70ºC and titrated with KMno4.The titration was continued until a faint pink color was produced. ASSAY: 1gm of ferrous sulphate was weighed accurately and dissolved in 20 ml of dilute sulfuric acid .Then it was titrated with 0.1N KMno4 until permanent color was produced. 1ml of 0.1N KMno4 ≡ 0.02789gms of Ferrous sulphate. REPORT: The percentage purity of given ascorbic acid tablet was found to be

6. ASSAY OF METRONIDAZOLE

AIM: To perform the assay of Metronidazole by non-aqueous titration. APPARATUS: Conical flask, burette, pipette, measuring cylinder. CHEMICALS: Perchloric acid, Metronidazole, acetone, acetic anhydride, glacial acetic acid. PRINCIPLE: The substances which are weakly basic and weakly acidic, give sharp end points in non-aqueous titrations. Non-aqueous titration is the titration of substances dissolved in non-aqueous solvents. It is the most common titrimetric procedure used in assays. These titrations provide insoluble organic compounds are soluble. PROCEDURE: Preparation and standardization of 0.1N perchloric acid:8.5ml of perchloric acid was dissolved in about 700ml of glacial acetic acid in 100ml flask.30ml of acetic anhydride was added slowly and the volume was made up to 1000ml with glacial acetic acid.The solution was kept aside for 24hrs by which any small amount of water present in either perchloric acid or glacial acetic acid would react with acetic anhydride and solution becomes non-aqueous. The glacial acetic acid was not dissolved in perchloric acid. Then acetic anhydride was added which results in the formation of perchloric acetic anhydride which is highly explosive in nature. (CH3CO)2O + H2O → 2CH3COOH Standardization of 0.1N perchloric acid: 0.5gms of potassium hydrogen phthalate was taken in a conical flask and dissolved in 100ml of acetic acid using crystal violet as indicator. End point is green colour. 1ml of 0.1N perchloric acid ≡ 0.20422gms of potassium hydrogen phthalate. ASSAY:    

20 tablets were weighed, powdered and a quantity equivalent of 0.2gms was taken. Solution is extracted with 6 quantities each of 10ml of hot acetone and cool. 50ml of acetic anhydride was added. 0.1ml of anhydrous glacial acetic acid was added and 1% solution of brilliant green which acts as indicator, and titrated against 0.1N perchloric acid. Each ml of HClO4 ≡ 0.01712gms of metronidazole.

REPORT: The percentage purity of metronidazole tablets was found to be

7. ASSAY OF METFORMIN

AIM: To perform the assay of metformin hydrochloride tablets. APPARATUS: spatula, measuring cylinder, mortar and pestle. CHEMICALS: Metformin hydrochloride tablets, water. PRINCIPLE: The substances which are weakly acidic and weakly basic give sharp endpoints in non-aqueous titrations. Non-aqueous titration is the titration of substances dissolved in non-aqueous solvent. It is most common titrimetric procedure used in pharmacopeial assays. These titrations provide solvents in which organic compounds are dissolved. PROCEDURE: 

20 tablets were weighed and powdered and 0.1gm was taken and dissolved in 70ml of



water and diluted to 100ml and filter. From above solution, 10ml was taken and diluted to 100ml with water, from 10ml was taken and further diluted to 100ml and absorbance was measured at 232nm.

REPORT: The percentage purity of metformin was found to be

8. ESTIMATION OF AMPICILLIN BY NINHYDRIN REAGENT

AIM: To estimate the given sample by using ninhydrin reagent. CHEMICALS: Ampicillin, ninhydrin, 0.1N HCl. PRINCIPLE: Ampicillin is a β-lactum antibiotic used against gram +ve bacteria. The primary amine present in ampicillin reacts with ninhydrin. This reaction in presence of 0.1N HCl using ninhydrin reagent, it forms L-amino acid which on reaction with ninhydrin gives colour complex. Absorbance was seen at 490nm. PROCEDURE: For calibration curve: 100mg of drug was dissolved in 100ml of 0.1N HCl to give 1mg/ml. 

From this 1, 2, 3, 4, 5ml was taken, to each 2ml of ninhydrin, 1.5ml of 0.1N HCl, 2ml



of methanol was added. Then it was heated until colour developed. It was then cooled and volume was made up to 100 ml with methanol and absorbance was measured at 490nm.

ASSAY: 

20 tablets were weighed and quantity equivalent to 100mg was dissolved in 100ml of



0.1N HCl. From this 3 ml was taken, 1.5N HCl, ninhydrin, 2ml methanol were added and heated



for 15min until the colour was developed. It was then cooled and diluted to 10ml and absorbance was measured at 492nm.

REPORT: The percentage purity of Ampicillin was found to be

9. ASSAY OF FUROSEMIDE

AIM: To perform the assay of furosemide by diazotization reaction APPARATUS: Measuring cylinder, pipette and volumetric flask. CHEMICALS: Furosemide pure drug and tablets, sodium nitrite, methanol, aniline, HCl, Water. PRINCIPLE: Furosemide is a diuretic drug. It is a sulphonamide derivative. This drug has a carbonyl group with sulphonyl moiety in Meta position (carbon no.5), a halide substitution is present at carbon-4 and substituted amino group is present at carbon-2. Furosemide is a highly potent diuretic related to the thiazides but more potent than them. Furosemide has high efficiency and rapid onset of action and short duration of action. It acts only on proximal and distal tubules and allow on the ascending limb of loop of Henley It is indicated in the treatment of edema associated with congestive heart failure, cirrhosis of liver, renal disease and hypertension The hydrolysed furosemide is diazotized with nitrite in acid medium and coupled with aniline to give a water soluble coloured dye having an absorption maximum at 480nm. PROCEDURE: Nitrite solution: A 1000 ppm nitrite solution was prepared by dissolving 0.15gms of NaNO2in some distilled water and finally diluted to 100ml. Aniline solution: A 10% v/v aniline solution was prepared by mixing 10ml of aniline in 50ml methanol and diluted to 100ml with water. PREPARATION OF STANDARD: 

0.05gms of furosemide was weighed and was dissolved in 50ml of methanol



From the above solution, 1ml was diluted with 200ml of methanol which gives

  

10ppm. From the above 2, 4, 6, 8and 10ml were pipetted and make up to 10ml with methanol To the aliquots of drug 2 ml of 3.5M HCl was added and 10ml of NaNO2was added. The solutions were kept for 5min and allow to diazotization reaction to go to



completion. The resulting solutions are coupled with aniline by adding 7ml of 10% aniline



followed by heating on water bath. The orange coloured azo dye was diluted to 100ml with distilled water and absorbance was measured at 480nm

ASSAY: 

10 tablets were taken , weight of drug equivalent to 0.05gms was taken and dissolved

 

in 50ml CH3OH 1ml of above solution was taken and diluted to 10ml with methanol From the resulting solution any volume ranging from 2-10ml was pipetted out and



diluted with ethanol to 10 ml To these solutions, 2ml of 3.5M HCl was added followed by addition of 10ml NaNO 2

 

and keep aside for 5min. Then 7ml of 10% aniline was added and was boiled in water bath for 6min, The resulting orange colour azo dye was diluted to 100ml with distilled water and absorbance was noted at 480nm.

REPORT: The percentage purity of furosemide was found to be

10. ESTIMATION OF CEPHADROXIL BY MBTH REAGENT AIM: To estimate the amount of cephadroxil by MBTH reagent. APPARATUS: Pipette, mortar and pestle, measuring cylinder, volumetric flask.

CHEMICALS: MBTH reagent, distilled water, cephadroxil pure drug and ceric ammonium nitrate. PRINCIPLE: MBTH→3 methyl, 2-benzo thiazolinone hydrazine was originally introduced for aldehydes, and later extended to a variety of organic compounds like phenol, aryl amines and different N and S heterocyclic compounds. Cephadroxil is an antibiotic which belongs to the category of cephalosporins which act by disrupting the cell wall. Cephodroxil consists of phenol group which interacts with MBTH and gives coloured solution which is observed under U.V. With phenols under reaction conditions MBTH loses two electrons and one proton to form the electrophilic intermediate, which has been identified as the active coupling species that undergoes electrophilic substitution with phenols and other groups to form coloured products. PROCEDURE: For calibration curve: 10 mg of pure drug was weighed and dissolved in 10ml of water. From the above solution 0.1, 0.2, 0.3, 0.4 and 0.5 ml are taken and to these solutions 2ml of MBTH solution was taken and to these solutions 2ml of MBTH solution was added and 2ml of Ce(IV) solution was added and volume was made up to 1oml with distilled water which produces concentration of 10, 20, 30, 40 and 50 µg/ml. The absorbance was noted at 410nm.

REPORT: The percentage purity of cephadroxil was found to be

11. ESTIMATION OF DICLOFENAC BY MBTH REAGENT AIM: To estimate the amount of diclofenac present in given tablet using MBTH. CHEMICALS: Diclofenac, MBTH, ceric ammonium nitrate, 1M H2SO4.

APPARATUS: Beaker, funnel, pipette, conical flask, volumetric flask. PRINCIPLE: Diclofenac 2(2, 6-dichlorophenyl) amino benzene acetic acid mono salt. It is an NSAID advised for use in painful and inflammation, rheumatic and non-rheumatic condition. The proposed mechanism involves the use of MBTH in presence ofcerric ammonium as the chromogenic agent for diclofenac. It obeys beers law at 0.2-8 µg/ml and absorbance was noted at 580nm. PROCEDURE:   

100 mg of drug was dissolved in 100 ml water to get 1000µg/ml. From this 2.5ml was taken and diluted to 25ml to get 100µg/ml (stock-1) From stock, pipette out 0.4, 0.5, 0.6, 0.7, 0.8 and 1ml in 10ml volumetric flask and



add 1ml MBTH and 2ml cerric ammonium nitrate and made up to 10ml with water. Blue colour develops due to complex formed. The concentrations are 4, 5, 6, 7, 8



µg/ml. The absorbance was measured at 580nm and curve was plotted.

ASSAY:  

20 tablets were weighed and average weight was noted. Powder equivalent to 100mg was weighed and dissolved in 100ml of water to get



1000µg/ml From this 0.5ml was taken and add 1ml MBTH, 2ml cerric ammonium nitrate. Make up to 10ml with water and absorbance was noted.

REPORT: The percentage purity of Diclofenac was found to be

12. ESTIMATION OF ATORVASTATIN BY MBTH REAGENT AIM: To estimate the amount of atorvastatin present in given tablet using MBTH. CHEMICALS: Atorvastatin (AVS) pure drug and tablets, MBTH, methanol, Ce(SO 4)2, sulphuric acid (0.368M).

APPARATUS: Beaker, funnel, pipette, conical flask, volumetric flask. PRINCIPLE: Statins are commonly used to treat several forms of hypercholesterolemia. They have potent cholesterol-lowering effects and they could reduce morbidity and mortality associated with coronary heart disease. Atorvastatin calcium is chemically {[R-(R, R*)]-2-(4fluro

phenyl)-β,δ-dihydroxy-5(1-methylethyl)-3-phenyl-4-[phenyl

amino

carbonyl]-1h-

pyrrole-1-heptanoic acid, calcium salt92:10trihydrate} is the most commonly occurring drug in commercially available pharmaceutical formulations.AVS reacts with MBTH in the presence of Ce (SO4)2in an acidic medium to form pink coloured oxidative coupling product that can be measured at 566nm. Under the reaction conditions on oxidation, MBTH loses two electrons and one H+ forming an electrophilic intermediate, which is active coupling species. The coupling of oxidized form of the drug with electrophilic intermediate of MBTH results in the formation of intensely coloured product.

PROCEDURE: Preparation of stock solutions: stock standard solution of 0.5 mg/ml was prepared by dissolving the appropriate weight of AVS in 100ml volumetric flask, 5ml of methanol was added the volume was then diluted to the mark with distilled water. 0.01M MBTH solution was prepared with doubled distilled water and 1% Ce (SO 4)2 solution was prepared with sulphuric acid of 0.369M, medium. Freshly prepared solutions were always used.

Procedure for calibration curve: Aliquots of standard AVS solution (0.1-1.0ml, 0.5mg/ml) were transferred into a series of 25ml calibrated volumetric flask. Then 1ml of MBTH solution was added and kept a side for 3 min. after that, 1 ml of Ce(SO 4)2 solution was added.

The volume was made up to mark with distilled water mix well and note the absorbance at 566nm as function of time between 0-20min against reagent blank treated similarly.

ASSAY: 20 tablets containing AVS were weighed and pulverized. An amount of the powder equivalent to 25mg of the cited drug was dissolved in a 25ml of CH 3OHand mixed for about 5min and filtered. The methanol was evaporated to about 5ml. the remaining portion of solution was diluted to 50ml with double distilled water to achieve a concentration of 0.5mg/ml.Then to this above solution 1ml of MBTH was added and kept aside for 3min. after that 1ml of Ce(SO4)2was added and the volume was made up to mark (25ml) with double distilled water. Mix well and absorbance was noted at 566nm.

REPORT: The percentage purity of atorvastatin was found to be

13. ESTIMATION OF CEPHALEXIN BY FC-REAGENT AIM: To estimate the amount of cephalexin by FC reagent. CHEMICALS: Cephalexin pure drug, FC reagent 10% NaOH, distilled water, tablets, NaNO2 APPARATUS: UV-visible spectrophotometer analytical mode, T-80, volumetric flask measuring cylinder, pipettes etc.

PRINCIPLE: Cephalexin is a first generation cephalosporin antibiotic. It is a bactericidal agent. It can be determined spectrophotometrically by using FC reagent in the presence of 10% NaOH, giving greenish blue colour. The absorbance was measured at 753nm. PROCEDURE: For calibration curve: 100mg of pure drug was weighed and dissolved in 100ml water. From this solution 10ml was taken and made to 100ml with water to give 100µg/ml. From this 1, 1.5, 2, 2.5, 3ml were pipetted out and 2ml of FC-reagent and 2.5ml NaOH was added volume was made up to 10ml with distilled water. The resultant concentrations were 10, 15, 20, 25 and 30 µg/ml respectively. Absorbance was noted at 753nm and curve was plotted. ASSAY: 

20 tablets were weighed and the drug equivalent to 100mg was taken and dissolved in



100ml of distilled water. From the above stock solution, 2ml was taken and the volume was made up to 100ml, from this 1ml was taken and to this solution 0.1 ml of FC reagent and 1ml of NaNO 2 was added and volume was made up to 10ml with distilled water and absorbance was noted at 753nm.

REPORT: The percentage purity of cephalexin was found to be

14. ESTIMATION OF CIPROFLOXACIN BY FCREAGENT AIM: To estimate the percentage purity of ciprofloxacin by UV-visible spectroscopy. CHEMICALS: 0.2% FeCl3, ciprofloxacin pure drug and tablet, FC-reagent, distilled water. APPARATUS: Volumetric flask, beaker. Pipette, UV spectrophotometer PRINCIPLE: Ciprofloxacin is a synthetic chemotherapeutic antibiotic of fluoroquinalone drug class. It obeys beer lamberts law in a range of 20-120 µg/ml.Folin ciocalteau reagent reacts with ciprofloxacin to form chromogen that can be detected spectrophotometrically. The colour development is due to the transfer of electrons at basic pH to reduce the phosphomolybdic acid (or) phosphotungstic acid complexes to form chromogen in which metals have lower valency. PROCEDURE For calibration curve: 0.5 mg/ml of stock solution was prepared by taking 50mg of drug in 100ml of water. Pipette out 2, 3, 4, 5, 6ml and diluted to 25ml and add 1ml FeCl 3 and 1ml FC reagent and then diluted to 25ml. Which gives concentration of 40, 60, 80, 100 and 120 µg/ml. The absorbance was measured and calibration curve was plotted. ASSAY: Weigh and powder tablets and take the weight equivalent to 50mg and diluted to 100ml with distilled water.From the above solution pipette out 2ml and add 1ml FeCl 3 and 1ml folin ciocalteau reagent and make up to the mark with distilled water to 25ml. The absorbance of the resulting solution was measured. REPORT: The percentage purity of ciprofloxacin was found to be

15. ESTIMATION OF RANITIDINE BY USING PDAB REAGENT

AIM: To estimate the amount of Ranitidine HCl pure drug and tablets, distilled water. APPARATUS: UV- visible spectrophotometer, volumetric flask, pipette, measuring cylinder etc. PRINCIPLE: Ranitidine HCl is chemically N, N dimethyl-5-[2-(1-methyl amine-2nitrovinyl)-ethylthiomethyl] furfuryl amine hydrochloride. It is a H 2-receptor antagonist and is widely used in short term treatment of duodenal ulcer and in the management of hyper secretory conditions. This method is based on the condensation of RNH with PDAB to form a red coloured product. The absorbance of coloured product measured is a quantitative measure of concentration of RNH. The absorbance was measured at 503nm. Procedure for preparation of standard solutions: PDAB solution of 0.025N was prepared in 2N HCl. Ranitidine HCl stock solution containing 1000µg/ml RNH was prepared by dissolving 100mg of pure drug in water and diluting to the mark in a 100ml calibrated flask. Procedure for calibration curve: Aliquots containing 50-350µg/ml RNH were transferred into a series of 10ml standard flask by means of a micro burette. To each flask 3ml of 0.025N PDAB was added and kept a side for 20min for colour development. It was then diluted up to the mark with distilled water. The absorbance was measured against reagent blank at 503nm. The graph of PDAB-RNH condensation product and corresponding reagent blank was plotted by taking concentration Vs absorbance. ASSAY: Three tablets were taken and ground into a fine powder. Powder equivalent to 168mg of RNH was weighed into a 100ml calibrated flask, 60 ml of water was added and shaken well and filtered. First 10ml portion of the filtrate was discarded. A suitable aliquot was taken and add 3ml of 0.025N P-Dimethyl amino benzaldehyde was added and kept aside for 20 min for colour development. It was then diluted up to the mark with distilled water. The absorbance was measured against the reagent blank at 503nm.

REPORT: The percentage purity of Ranitidine was found to be

.