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1.1 General introduction
Analytical chemistry may be defined as the science and art of determining the composition of materials in terms of the elements or compounds contained in them. Infact,analytical chemistry is the science of chemical identification and determination of the composition (atomic,molecular,phase) of substances and materials and and their chemical structure. The main object of analytical chemistry is to develop scientifically substantiated methods that allow the qualitative and quantitative evaluation of materials with certain accuracy. Analytical chemistry derives it’s principles from various branches of Science like chemistry,physics microbiology,nuclear science, electronics. This method provides information about the relative amount of one more of these components.
1.2 Selection of the most appropriate analytical method should taken in to account the following factors.
The purpose of the analysis, the required time scale and any cost constraints. The level of analyte (s) expected and the detection limit required. The nature of the sample, the amount available and the preparation procedure The accuracy required for a quantitative method. The availability of reference materials , standards chemicals instrumentation and special facilities. Possible interferences with the detection or quantitative measurement of the analyte and the possible need for sample clean up to avoid matrix interference. and solvents necessary sample
1.3 Different methods of analysis
By using this process the components of interest is separated and analysed by using the following techniques .
(a) Spectral methods :The spectral techniques are used to measure electromagnetic radiation which is either absorbed or emitted by the sample e.g.- U.V. visible spectroscopy ,I R spectroscopy, N M R, ESR spectroscopy, flame photometery , fluorimetery.
(b) Electro analytical method:Electro analytical methods involved the measurement of current as a property of concentration of the component in eg.:- Potentiometry , conductometry, amprometry. voltage or resistance solution mixture.
(c) Chromatographic methods :Chromatography is a technique in which chemicals in solutions travel down columns or over surface by means of liquids or gases and are separated from each other due to there molecular characteristics. eg:- Paper chromatography Thin layer chromatography High performance thin layer chromatography High performance liquid chromatography Gas chromatography
The technique of ultraviolet-visible spectroscopy is one of the most frequently employed techniques in pharmaceutical analysis. Molecular absorption in ultraviolet & visible region of spectrum is depend on the electronic structure of molecules. Absorption of energy as quantized, resulting in the elevation of electrons from orbital in the ground state to higher orbital in the exited state. The wavelength range of u.v. radiation starts at the blue end of the visible light and ends at 2000 Ǻ. The ultraviolet region is subdivided in to two spectral regions. (1) (2) The region between 2000Ǻ-4000 Ǻ is known as near as U.V. region. The region below 2000 Ǻ is called far or vaccume U.V. region.
Any molecule has either and n or combination of these electrons. These bonding and nonbonding (n) electrons absorb the characteristics radiation and undergo transition from ground state to exited state. By the characteristic absorption peaks the nature of electrons present and hence the molecular structure can be elucidated Ultraviolet absorption spectra arise from transition of electron or electrons with in a molecule or an ion from a lower to higher electronic energy level and the ultraviolet emission spectra arise from the reverse type of transition. When a molecule absorbs ultraviolet radiation of frequency v sec -1, the electron in that molecule undergoes transition from a lower to a higher energy level or molecular orbital, energy difference is given byE=h v erg The actual amount of energy required depends on the difference in energy between the ground state E0 and excited state E1 of the electrons. E1 -E0 =hv We known that total energy of a molecule is equal to the sum of electronic vibrational and rotational energy. The magnitude of these energies decrease in following order :- E etec , E vib and E rot.
The most important characteristic of spectrophotometry are there wide applicability, high sensitivity, moderate to high sensitivity, good accuracy and convenience. The assay of an absorbing substance may be quickly carried out by preparing a solvent in a transparent solvents and measuring its absorbance at a suitable wavelength. This is governed by Beer-Lambert’s law which statesA Where, A is the absorbance. a is the absorbitivity. b is the path length. c is the concentration. =abc
(a) Source of light: - The best source of light is the one which is more stable, more
intense and which gives range of spectrum from 180-360 nm (up to 400 nm). The different sources available are: 1. 2. 3. 4. 5. Tungston lamp.. Hydrogen discharge lamp Deuterium lamp. Xenon discharge lamp. Mercury arc.
The details are as follow:-
(i) Tungston lamp:- The various radiation sources are as follows:
The two most common radiation sources are tungsten lamps and hydrogen discharge lamps. The tungsten lamp is similar in' its functioning to an electric light bulb. It is tungsten. Filament heated electrically to white heat. It has two shortcomings. The intensity of radiation at short wavelengths (<350 nm) is small.
Furthermore, to maintain a constant intensity, the electrical current to the lamp must be carefully controlled. However, the lamps are generally stabled, robust, and easy to use. Typically, the emission intensity varies with wavelength.
(ii) Hydrogen discharge lamps:In these lamps, hydrogen gas is stored under relatively high pressure. When an electric discharge is passed through the lamp, excited hydrogen molecules will be produced which emit UV radiations. The high pressure in the hydrogen lamps causes the hydrogen to emit a continuum rather than a simple hydrogen spectrum. Hydrogen lamps cover the range 3500-1200 A. These lamps are stable, robust and widely used. The hydrogen discharge lamp consists of hydrogen gas under relatively high pressure through which there is an electrical discharge. The hydrogen molecules are excited electrically and emit UV radiation. The high pressure brings about many collisions between the hydrogen molecules, resulting in pressure broadening. This causes the hydrogen to emit a continuum (broad band) rather than a simple hydrogen line spectrum. The lamps are , stable, robust, and widely used. If deuterium (D2) is used instead of hydrogen, the emission intensity is increased by as much as a factor of 3 at the short-wavelength end of the UV range. Deuterium lamps are more expensive than hydrogen lamps but are used when higher intensity is required.
(iii) Deuterium lamps:If deuterium is used in place of hydrogen, the intensity of radiation emitted is 3 to 5 times the intensity of a hydrogen lamp of comparable design and wattage. Deuterium lamp is more expensive than hydrogen lamp. But, it is used when high intensity is required.
(iv) Xenon discharge lamps:In these lamps, xenon gas is stored under pressure in the range of 10-30 atmospheres. The xenon lamp possesses two tungsten electrodes separated by about 8 mm. When applying a low voltage forms an intense arc, the ultraviolet light is produced. The intensity of ultraviolet radiation produced by xenon discharge lamp is much greater than that of hydrogen lamp. 6
(v) Mercury arc:In this, the mercury vapour is under high pressure, and the excitation of mercury atoms is done by electric discharge. The mercury are, a standard source for much ultraviolet work, is generally not suitable for continuous spectral studies because of the presence of sharp lines or bands.
(b) Monochromators:Filters and prism monochromators are not used because of low resolution. On the other hand grating provides a band pass of 0.4 to 2nm. Hence they are more widely used, especially in expensive spectrophotometers .the mirrors, gratings etc are made up of quartz, since glass absorbs UV radiations from 200-300 nm. mirrors are front surfaced to prevent absorption of radiation . e.g.: - Prism.Gratings
(c) Sample cells:The design of sample cell used are similar to that used in colorimetry except that it is made up of quartz. Quartz cells only must be used UV spectroscopy since glass cells will absorb UV radiation. The path length of the cell are 10nm or 1cm.
(d) Solvents:Solvent plays an important role in UV spectra. Hence the solvent for a sample is selected in such a way that the solvent neither absorbs in the region of measurement not affects the absorption of the sample.
(e) Detectors:Although any one of the detectors used in calorimetric can be used, photomultipliter tubes are mainly used, since the cost of such UV spectrophotometers are high and more accurate measurements are to be made. E.g.: - (1) Barrier layer cell or photo voltaic cell. (2) Photo tubes (or) photo emissive cell. 7
Photo multiplier tubes (PMT).
- ULTRAVIOLET SPECROPHOTOMETER
(1) Qualitative Analysis:-
(a) Detection of impurities: To limit the presence of impurities, we can use Prism UV spectrophotometer measurements additional peaks can be due to Photomultiplier Photomultip M1 impurities in the sample and can be compared with that of standard raw Reference material. Also by absorbance measurement as specific wavelength the impurities can be detected. (b) Structure elucidation Rotating compound of organic Sector (c) Structure analysis of organic compound Lamp Beam MonochroSample (Source 2) Quantitative Analysis:- splitter chamber mator (a) Using (b)Single standard or direct comparison method. (c)Calibration curve method . (3) Determination of molecular weight (4) Determination of dissociation constant of acids and bases (5) Chemical kinetics (6) Identification of unknown compound. (7) Detection of poly nuclear hydrocarbons. (8)Determination of conjugation of geometrical isomerism. (9) Identification of the compound in different isomerism.
values can we find out the quantity of drug molecule?
(10) Detection of functional groups. (11) Dissociation in conjugate and non-conjugate
Drug name:DICLOFENAC SODIUM FENAC PLUS, DICLOWIN PLUS Structure :-
Systematic-(IUPAC) name:--2-(2-(2,6-dichlorophenylamino)phenyl)acetic acid Molecular MASS :- 296.148 g/mol Empirical formula: - C14H11Cl2NO
Oral: Adults: 50 mg twice daily
Therapeutic category:- NSAID’S Reported use:- PAIN KILLER
International-Brand-Name :- DOLO, DOLOPAR, CALPOL,
Molecular MASS :- 151.169 g/mol Empirical formula: - C8H9NO2 Dose:Oral: Adults: 500 mg twice daily
Therapeutic category:- NSAID’S Reported use:- ANTIPYRETIC Description of drug:These are the drug used in the treatment of fever pain respectievely.these have novel mode of action that differentiates it from non-steroidal anti-inflamm Clinical trials have shown tha Pracetamol & diclofenac sodium is highly effective in relieving the symptoms of fever and pain. antiflammatory drugs (NSAIDs) and other conventional forms of drug therapy.
Paracetmol:- It is soluble in hydrochloric acid . Diclofenac sodium:Storage:Store in tightly closed container, and reach out from the children,store the tablet away from light , at room temperature or in the refrigerator.
It is soluble in sodium hydroxide
FENAC PLUS - Blister of 10 Tablet.
OBJECTIVE AND PLAN OF WORK
DICLOFENAC SODIUM FORMULATION.
1) Plan of work
1. 2. 3. 4. 5. 6. 7. Determination of the partition coefficient. λ max determination of Paracetamol. λ max determination of Diclofenac sodium. Preparation of calibration curve of Paracetamol. Preparation of calibration curve of Diclofenac sodium. Preparation of sample of paracetamol. Preparation of sample of Diclofenac sodium.
2) Material & Method
1. 2. Hydrochloric sodium 0.1 N . Sodium hydroxide 0.1 N .
Distilled water (H2O ) Paracetamol drug ( Pure form ) Diclofenac sodium.
3.1 λ max determination:-
The uv spectrum of paracetamol and diclofenac sodium in solvent like methanol,0.1N HCL , 0.1N NAOH . were recorded for a different concentration 10µ g /ml. The spectrum of paracetamol and diclofenac sodium was found to have good spectrum pattern and maximum absorbance was obtained . Hence 0.1 N NAOH was selected as the solvent . the maximum absorbance was found to 257 nm for the paracetamol and 252 for the diclofenac sodium .
Fig.3.1.1 Spectrum of Diclofenac in NaOH
Fig.3.1.2 Spectrum of Paracetamol in hydrochloric acid
3.2 Preparation of caibration curve of 1) Paracetamol:The pure compound of paracetamol 100mg dissolve in 100 ml 0.1N HCL .form the 10ml take in 100ml volume metric falsk from this 10 ml take in 100 ml volume metric falsk to make 10 mg/ ml .solution form this we have made different concentration solution line 2 mg /ml,4 mg /ml,6 mg /ml,8 mg /ml,10 mg /ml. The paracetamol was found to have maximum absorbance 257nm and hence it was selected for further studies.
Diclofenac sodium :-
Stock solution of diclofenac sodium of 1000mg/ml was prepared by taking 100mg of durg dissolved and diluted to 100 ml with 0.1N NaOH.the above stock solution was suitably diluted with NaOH to get concentration ,ranging from 2-10µ g/ml and absorbance were noted at 252nm and the overlain spectra were show in fig.3.2.2.
Table3.1 :- Data represent the absorbance of the different concentration of paracetamol at 257.0 nm.
S no. 1 2 3 4 5
Cocentration Цg /ml 1 5 10 15 20
absorbance 1.008 1.625 1.756 1.757 1.758
Table3..2 :- Data represent the absorbance of the different concentration of diclofenac sodium in 252.0 nm. S no. 1 2 3 4 5 Cocentration Цg /ml 2 4 6 8 10 0.789 1.365 1.677 1.712 1.731 absorbance
Fig.3.2.1 Dilution of paracetomol
Fig.3.2.2 Dilution of diclofenac sodium
parcetomal standard graph
1 0.9 0.8 0.7 0.6 Abs 0.5 0.4 0.3 0.2 0.1 0 1µg/ml 5µg//ml 10µg/ml conc 15µg/ml 20µg/ml Series1
Fig.3.2.3 Calibration graph of paracetamol in 257nm
Dichlofinac sodium standard graph
0 2µg/ml 4µg/ml 6µg/ml conc 8µg/ml 10µg/ml
Fig.3.2.4 Calibration graph of diclofenac sodium in 252nm.
3.3 prepartion estimation in sample solution of fenac plus :-
20 tablet of 250mg of paracetamol and diclofenac sodium and average weight. Was calculated
sample solution of paracetamol:Weigh accurately powder sample equivalent to about 100mg of paracetamol carry out
the dry portion with three 25 ml portion of 0.1N HCL filtering through by sinterdred glass funnel. Dilute the combined filterate to 100ml with acid dilute further appropriately with acid got final concentration 100mg /ml .
paracetamol sample graph
2 1.8 1.6 1.4 1.2 abs 1 0.8 0.6 0.4 0.2 0 2 µg/ml 4µg/ml 6µg/ml conc 8µg/ml 10µg/ml Series1
2) sample:The entire residue left after the extraction of paracetamol as described above is used for estimation of diclofenac sodium. Transfer the residue left in the conical flask and the top of sintered fannel.qualitatively to 100ml volumetric flask with the help of 0.1 N NAOH make up the volume and carry out the further dilution appropriately with NAOH toget final concentration of 100 µ g/ml.
diclofenac sodium sample
2 1.8 1.6 1.4 1.2 abs 1 0.8 0.6 0.4 0.2 0 2 µg/m l 4µg/m l 6µg/m l conc 8µg/m l 10µg/m l Series1
RESULT AND DISCUSSION:On the basis of experimental data.The paracetamol showed maximum absorbance at 252nm in HCl and diclofenac sodium showed maximum absorbance at 257nm in NaOH.The data of precision given on method is more accurate and an equal but little time different from the standard value or calibration curve.
• Douglas-A.-Skoog,-Donald M.West, and JamesF. Holler: Fundamentals of analytical chemistry :Page No564-567 5th edition.
• Gurdeep=R.chatwal,-Sham-K.ANAND:Instrumentalmethods_of_chemical analysis:Page No. 2.149-2.151, 2.160-2.161,2.176-2.178: 5th edition. • J.denny-R.C,-Barnesj.d,Thomas M:Textbook of quantitative chemical analysis:Page No. 278-279:6th edition. • Scientific & research Publication from , Indian Drug manufactures association :Indian Drugs :Vol.44 : No 1 :January 2007 :Page No 13 . • Scientific & research Publication from , Indian Drug manufactures association :Indian Drugs :Vol.44 : No 2 : February 2007 :Page No 145 . • Scientific & research Publication from , Indian Drug manufactures association :Indian Drugs :Vol.44 : No 3 : March 2007 :Page No 209-210 . • Sharma B. K. Instrumental Methods of chemical analysis :Page No.S-46,S-52 :1 st edition. INTERNET WEB SITE • www.sciencedirect.com • www.pubmed.com • www.google.com • www.wikipedia.com • www.encylopedia.com
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