logging in or signing up quantitative analysis by UV spectroscopy samaju Download Post to : URL : Related Presentations : Share Add to Flag Embed Email Send to Blogs and Networks Add to Channel Uploaded from authorPOINT lite Insert YouTube videos in PowerPont slides with aS Desktop Copy embed code: (To copy code, click on the text box) Embed: URL: Thumbnail: WordPress Embed Customize Embed The presentation is successfully added In Your Favorites. Views: 1372 Category: Education License: All Rights Reserved Like it (4) Dislike it (0) Added: September 07, 2010 This Presentation is Public Favorites: 0 Presentation Description No description available. Comments Posting comment... By: anuradharc (13 month(s) ago) can u mail me the presentation to anuradharaichowdhury19@gmail.com Saving..... Post Reply Close Saving..... Edit Comment Close By: KhanBangash (14 month(s) ago) Good Job Saving..... Post Reply Close Saving..... Edit Comment Close By: samaju (18 month(s) ago) if nu need the presentation mail me at ajussamuel@gmail.com Saving..... Post Reply Close Saving..... Edit Comment Close By: samaju (18 month(s) ago) if nu need the presentation mail me at ajussamuel@gmail.com Saving..... Post Reply Close Saving..... Edit Comment Close By: samaju (18 month(s) ago) if nu need the presentation mail me at ajussamuel@gmail.com Saving..... Post Reply Close Saving..... Edit Comment Close loading.... See all Premium member Presentation Transcript QUANTITATIVE ANALYSIS BY UV - SPECTROSCOPY : QUANTITATIVE ANALYSIS BY UV - SPECTROSCOPY Aju S. Sam 1st year M.Pharm Nehru College of Pharmacy Introduction : Introduction LAMBERT’S LAW: “When a beam of light is allowed to pass through a transparent medium the rate of decrease in intensity with the thickness of the medium is directly proportional to intensity of the light.” -dI/dt ∞ I It = I0 e-kt (1) Slide 3: BEER’S LAW “The intensity of a beam of monochromatic light decreases exponentially with increase in concentration of absorbing substances arithmetically.” It = I0e–k’c (2) On simplification and combining (1) and (2) we get Slide 4: log (I0/It) = act This is called BEER-LAMBERT’S LAW. Relationship between Absorbance(A), Transmittance(T) , & Molar extinction coefficient(ε). A = εct = log (I0/It)= log (1/T) = log -T METHODS USED FOR ASSAY : METHODS USED FOR ASSAY USE OF CALIBRATION GRAPH: Step 1: select the λmax . Step 2: prepare series of known conc. Solutions. Step 3: set λmax in spectrophotometer. Step 4: measure absorbance. Step 5: plot calibration curve. absorbance conc. 2. USE OF STANDARD ABSORPTIVE VALUE : 2. USE OF STANDARD ABSORPTIVE VALUE In this method standard ε value ie ε 1%1cm is used. A = ε1%1cmct e.g. ε1%1cm for methyl testosterone in B.P is 540 at 241 nm. 3.SINGLE OR DOUBLE POINT STANDARDISATION : 3.SINGLE OR DOUBLE POINT STANDARDISATION Involves measurement of sample and reference. Conc. of reference close to that of sample. C test= A test × C std A std Double point involves use of 2 reference. one greater conc. than sample, other lower conc. than sample. C test=(Atest- Astd1)(Cstd1- Cstd2)+Cstd1(Astd1- Astd2) Astd1-Astd2 4.ASSAY USING ABSORBANCE CORRECTED FOR INTERFERENCE : 4.ASSAY USING ABSORBANCE CORRECTED FOR INTERFERENCE The concentration of absorbing component = total absorbance – absorbance of interfering substance. 5.Assay after solvent extraction of sample : 5.Assay after solvent extraction of sample Involves separation of absorbing interferents by solvent extraction, choice of pH . e.g. B.P Assay of Caffeine in Aspirin & Caffeine tablet 6.SIMULTANEOUS EQUATION METHOD : 6.SIMULTANEOUS EQUATION METHOD Total absorbance of a solution is equal to the sum of absorbance of individual components present. absorbance λ1 wave length λ2 mixture x y Slide 11: A1= ax1bcx+ay1bcy (at 1) A2= ax2bcx+ay2bcy (at 2) C x = A2ay1-A1ay2 ax2ay1-ax1ay2 Cy= A1ax2-A2ax1 ax2ay1-ax1ay2 e.g. B.P assay of quinine related alkaloids in Cinchona bark. 7.ABSORBANCE RATIO METHOD : 7.ABSORBANCE RATIO METHOD For a substance which obeys BEER’S LAW at all wavelengths, the ratio of absorbance at any two wavelengths is a constant value independent of concentration or path length. In USP the ratio is referred as Q value. e.g. cyanocobalamin exhibits three λmax at 278nm, 361nm, 550nm A361/A550 = 3.30 ± 0.15 A361/A278 = 1.79 ± 0.09 DIFFERENCE SPECTROSCOPY : DIFFERENCE SPECTROSCOPY Two samples used, one in reference beam and one in sample beam. Conc. of absorbing substance identical. But some solution parameter like pH is different. Difference absorbance ∆A = A alk - A acid DIFFERENTIAL SPECTROSCOPY : DIFFERENTIAL SPECTROSCOPY Slide 15: 0 10 20 30 40 50 60 70 80 90 100 (BLANK) RECEIVER DARK 0 10 20 30 40 50 60 70 80 90 100 HIGH ABSORBANCE 0 10 20 30 40 50 60 70 80 90 100 (BLANK) 0 10 20 30 40 50 60 70 80 90 100 TRACE ANALYSIS 0 10 20 30 40 50 60 70 80 90 100 C1 C1>C2 C2 0 10 20 30 40 50 60 70 80 90 100 MAXIMUM PRECISION DERIVATIVE SPECTROPHOTOMETRY : DERIVATIVE SPECTROPHOTOMETRY Conversion of normal spectrum into its first or higher derivative spectrum. First derivative spectrum- (dA/dλ) v/s λ A λ dA/dλ λ + - λmax Cross over point (λmax) Slide 17: Second derivative spectrum A λ d2A/dλ2 λ + - X+Y X Y REFERENCES : REFERENCES A.H.BECKETT, J.B.STENLAKE.PRACTICAL PHARMACEUTICAL CHEMISTRY, 4TH EDITION, PART TWO; 2005: 275-300. WILLARD,MERRITT,DEAN,SETTLE. INSTRUMENTAL METHODS OF ANALYSIS, 7TH EDITION;1986:159-178. GURDEEP.R.CHATWAL,SHAM.K.ANAND. INSTRUMENTAL METHODS OF CHEMICAL ANALYSIS, 5TH EDITION;2007:140-178. Slide 19: THANKS You do not have the permission to view this presentation. In order to view it, please contact the author of the presentation.
quantitative analysis by UV spectroscopy samaju Download Post to : URL : Related Presentations : Share Add to Flag Embed Email Send to Blogs and Networks Add to Channel Uploaded from authorPOINT lite Insert YouTube videos in PowerPont slides with aS Desktop Copy embed code: (To copy code, click on the text box) Embed: URL: Thumbnail: WordPress Embed Customize Embed The presentation is successfully added In Your Favorites. Views: 1372 Category: Education License: All Rights Reserved Like it (4) Dislike it (0) Added: September 07, 2010 This Presentation is Public Favorites: 0 Presentation Description No description available. Comments Posting comment... By: anuradharc (13 month(s) ago) can u mail me the presentation to anuradharaichowdhury19@gmail.com Saving..... Post Reply Close Saving..... Edit Comment Close By: KhanBangash (14 month(s) ago) Good Job Saving..... Post Reply Close Saving..... Edit Comment Close By: samaju (18 month(s) ago) if nu need the presentation mail me at ajussamuel@gmail.com Saving..... Post Reply Close Saving..... Edit Comment Close By: samaju (18 month(s) ago) if nu need the presentation mail me at ajussamuel@gmail.com Saving..... Post Reply Close Saving..... Edit Comment Close By: samaju (18 month(s) ago) if nu need the presentation mail me at ajussamuel@gmail.com Saving..... Post Reply Close Saving..... Edit Comment Close loading.... See all Premium member Presentation Transcript QUANTITATIVE ANALYSIS BY UV - SPECTROSCOPY : QUANTITATIVE ANALYSIS BY UV - SPECTROSCOPY Aju S. Sam 1st year M.Pharm Nehru College of Pharmacy Introduction : Introduction LAMBERT’S LAW: “When a beam of light is allowed to pass through a transparent medium the rate of decrease in intensity with the thickness of the medium is directly proportional to intensity of the light.” -dI/dt ∞ I It = I0 e-kt (1) Slide 3: BEER’S LAW “The intensity of a beam of monochromatic light decreases exponentially with increase in concentration of absorbing substances arithmetically.” It = I0e–k’c (2) On simplification and combining (1) and (2) we get Slide 4: log (I0/It) = act This is called BEER-LAMBERT’S LAW. Relationship between Absorbance(A), Transmittance(T) , & Molar extinction coefficient(ε). A = εct = log (I0/It)= log (1/T) = log -T METHODS USED FOR ASSAY : METHODS USED FOR ASSAY USE OF CALIBRATION GRAPH: Step 1: select the λmax . Step 2: prepare series of known conc. Solutions. Step 3: set λmax in spectrophotometer. Step 4: measure absorbance. Step 5: plot calibration curve. absorbance conc. 2. USE OF STANDARD ABSORPTIVE VALUE : 2. USE OF STANDARD ABSORPTIVE VALUE In this method standard ε value ie ε 1%1cm is used. A = ε1%1cmct e.g. ε1%1cm for methyl testosterone in B.P is 540 at 241 nm. 3.SINGLE OR DOUBLE POINT STANDARDISATION : 3.SINGLE OR DOUBLE POINT STANDARDISATION Involves measurement of sample and reference. Conc. of reference close to that of sample. C test= A test × C std A std Double point involves use of 2 reference. one greater conc. than sample, other lower conc. than sample. C test=(Atest- Astd1)(Cstd1- Cstd2)+Cstd1(Astd1- Astd2) Astd1-Astd2 4.ASSAY USING ABSORBANCE CORRECTED FOR INTERFERENCE : 4.ASSAY USING ABSORBANCE CORRECTED FOR INTERFERENCE The concentration of absorbing component = total absorbance – absorbance of interfering substance. 5.Assay after solvent extraction of sample : 5.Assay after solvent extraction of sample Involves separation of absorbing interferents by solvent extraction, choice of pH . e.g. B.P Assay of Caffeine in Aspirin & Caffeine tablet 6.SIMULTANEOUS EQUATION METHOD : 6.SIMULTANEOUS EQUATION METHOD Total absorbance of a solution is equal to the sum of absorbance of individual components present. absorbance λ1 wave length λ2 mixture x y Slide 11: A1= ax1bcx+ay1bcy (at 1) A2= ax2bcx+ay2bcy (at 2) C x = A2ay1-A1ay2 ax2ay1-ax1ay2 Cy= A1ax2-A2ax1 ax2ay1-ax1ay2 e.g. B.P assay of quinine related alkaloids in Cinchona bark. 7.ABSORBANCE RATIO METHOD : 7.ABSORBANCE RATIO METHOD For a substance which obeys BEER’S LAW at all wavelengths, the ratio of absorbance at any two wavelengths is a constant value independent of concentration or path length. In USP the ratio is referred as Q value. e.g. cyanocobalamin exhibits three λmax at 278nm, 361nm, 550nm A361/A550 = 3.30 ± 0.15 A361/A278 = 1.79 ± 0.09 DIFFERENCE SPECTROSCOPY : DIFFERENCE SPECTROSCOPY Two samples used, one in reference beam and one in sample beam. Conc. of absorbing substance identical. But some solution parameter like pH is different. Difference absorbance ∆A = A alk - A acid DIFFERENTIAL SPECTROSCOPY : DIFFERENTIAL SPECTROSCOPY Slide 15: 0 10 20 30 40 50 60 70 80 90 100 (BLANK) RECEIVER DARK 0 10 20 30 40 50 60 70 80 90 100 HIGH ABSORBANCE 0 10 20 30 40 50 60 70 80 90 100 (BLANK) 0 10 20 30 40 50 60 70 80 90 100 TRACE ANALYSIS 0 10 20 30 40 50 60 70 80 90 100 C1 C1>C2 C2 0 10 20 30 40 50 60 70 80 90 100 MAXIMUM PRECISION DERIVATIVE SPECTROPHOTOMETRY : DERIVATIVE SPECTROPHOTOMETRY Conversion of normal spectrum into its first or higher derivative spectrum. First derivative spectrum- (dA/dλ) v/s λ A λ dA/dλ λ + - λmax Cross over point (λmax) Slide 17: Second derivative spectrum A λ d2A/dλ2 λ + - X+Y X Y REFERENCES : REFERENCES A.H.BECKETT, J.B.STENLAKE.PRACTICAL PHARMACEUTICAL CHEMISTRY, 4TH EDITION, PART TWO; 2005: 275-300. WILLARD,MERRITT,DEAN,SETTLE. INSTRUMENTAL METHODS OF ANALYSIS, 7TH EDITION;1986:159-178. GURDEEP.R.CHATWAL,SHAM.K.ANAND. INSTRUMENTAL METHODS OF CHEMICAL ANALYSIS, 5TH EDITION;2007:140-178. Slide 19: THANKS