Нестероидни противовъзпалителни средства

• Salicylates (Asrpirin);• Arylalcanoic acids (Indometacin, Diclofenac);• 2-Arylpropionic acids (Ibuprofen, Ketoprofen);• N-Arylanthranilic acids (fenamic acids);• Pyrazolidine derivatives (Metamizole);• Oxicams (Piroxicam);• Sulphonanilides (Nimesulide);• COX-2 Inhibitors (Celecoxib);• Други (Omega-3 fatty acids).

Most NSAIDs act as non-selective inhibitors of the enzyme cyclooxygenase, inhibiting both the cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) isoenzymes. Cyclooxygenase catalyses the formation of prostaglandins and thromboxane from arachidonic acid (itself derived from the cellular phospholipid bilayer by phospholipase A2). Prostaglandins act (among other things) as messenger molecules in the process of inflammation. This mechanism of action was elucidated by John Vane, who later received a Nobel Prize for this work.

NSAIDs

Indometacin contains not less than 98.5 per cent and not more than the equivalent of 100.5 per cent of [1-(4-chlorobenzoyl)- 5 - methoxy-2-methylindol-3-yl] acetic acid, calculated with reference to the dried substance.

A white or yellow, crystalline powder, practically insoluble in water, sparingly soluble in alcohol.

Indometacin

Indometacin

IDENTIFICATION

First identification: A, C.

Second identification: A, B, D, E.

A. Melting point (2.2.14): 158°C to 162°C.

B. Dissolve 25 mg in a mixture of 1 volume of 1M hydrochloric acid and 9 volumes of methanol R and dilute to 100.0 ml with the same mixture of solvents. Dilute 10.0 ml of the solution to 100.0 ml with a mixture of 1 volume of 1M hydrochloric acid and 9 volumes of methanol R. Examined between 300 nm and 350 nm (2.2.25), the solution shows an absorption maximum at 318 nm. The specific absorbance at the maximum is 170 to 190.

C. Examine by infrared absorption spectrophotometry (2.2.24), comparing with the spectrum obtained with indometacin CRS. Examine the substances in the solid state without recrystallisation.

+ R R C O H 2 N O H H C l ' .=

H 2 O H C lN + R R C O H ' = +

D. Dissolve 0.1 g in 10 ml of alcohol R, heating slightly if necessary. To 0.1 ml of the solution add 2 ml of a freshly prepared mixture of 1 volume of a 250 g/l solution of hydroxylamine hydrochloride R and 3 volumes of dilute sodium hydroxide solution R. Add 2 ml of dilute hydrochloric acid R and 1 ml of ferric chloride solution R2 and mix. A violet-pink colour develops.

NCH3

O Cl

H3COCOOH

Indomethacin

OH-

NCH3

H3COCOO-

O Cl

H

+

-O

Indometacin

E. To 0.5 ml of the solution in alcohol prepared in identification test D, add 0.5 ml of dimethylaminobenzaldehyde solution R2. A precipitate is formed that dissolves on shaking. Heat on a water- bath. A bluish-green colour is produced. Continue to heat for 5 min and cool in iced water for 2 min. A precipitate is formed and the colour changes to light greyish-green. Add 3 ml of alcohol R. The solution is clear and violet-pink in colour.

Indometacin

NCH3

H3COCOOCH3

R1

Indomethacin

NCH3

H3COCOOCH3

R2

methyl ester

R1 = C Cl

O

R2= N(CH3)2

R2 CHO/H+

R1+

1

1- H+

CH3OHN

CH3

H3COCOOCH3

H

R2

H3CO

4 6

Indometacin

1-H+

NR1

R2

H

NR1 CH3

COOCH3

OCH3

H3C

H3OOC

H3CO

7

Fe3+/H+

NR1

R2

NR1 CH3

COOCH3

OCH3

H3C

H3OOC

H3CO

+

5

Indometacin

ASSAY

Dissolve 0.300 g in 75 ml of acetone R, through which nitrogen R, free from carbon dioxide, has been passed for 15 min. Maintain a constant stream of nitrogen through the solution. Add 0.1 ml of phenolphthalein solution R. Titrate with 0.1 M sodium hydroxide. Carry out a blank titration.

1 ml of 0.1 M sodium hydroxide is equivalent to 35.78 mg of C19H16ClNO4.

STORAGE

Store in a well-closed container, protected from light.

IMPURITIES

A. 4-chlorobenzoic acid.

Indometacin

Sodium diclofenac

Sodium diclofenac contains not less than 99.0 per cent and not more than the equivalent of 101.0 per cent of sodium 2-[(2,6- dichlorophenyl)amino]phenyl]acetate, calculated with reference to the dried substance. A white or slightly yellowish, crystalline powder, slightly hygroscopic, sparingly soluble in water, freely soluble in methanol, soluble in alcohol, slightly soluble in acetone, practically insoluble in ether.

Примеси

Sodium diclofenac

ASSAY

Dissolve 0.250 g in 30 ml of glacial acetic acid R. Titrate with 0.1M perchloric acid, determining the end-point potentiometrically (2.2.20).

1 ml of 0.1 M perchloric acid is equivalent to 31.81 mg of C14H10Cl2NNaO2.

STORAGE

Store in a airtight container, protected from light.

Sodium diclofenac

Киселинно-основно титруване в ледена оцетна киселина:

Ледената оцетна киселина е един от най-широко използваните разтворители за определяне на слаби незаредени бази. Тя има силно изразен протон-донорен характер, поради което засилва базичния и отслабва киселинния характер на протолитите. Напр. незаредени бази стават с около 3 рКа единици по-силни. Ниската диелектрична константа на средата (4.9) води до силна йонизация и ниска дисоциация на протолитите. Като стандартен разтвор при титруването на бази се използва перхлорна киселина в оцетна киселина. При разтварянето протичат следните процеси:

HClO4 + CH3COOH = CH3COOH2+ . ClO4

-

B + CH3COOH = BH+. CH3COO-

BH+. CH3COO- + CH3COOH2+ . ClO4

- = BH+. ClO4- + 2 CH3COOH

Когато базите са във вид на хидрохлориди или хидробромиди се прибавя разтвор на живачен (ІІ) ацетат. В резултат на тази реакция се освобождават базите, а хлоридните и бромидните йони се свързват в малко разтворими съединения.

неводна среда

Ketoprofen

Ketoprofen contains not less than 99.0 per cent and not more than the equivalent of 100.5 per cent of (RS)-2-(3-benzoylphenyl) propionic acid, calculated with reference to the dried substance.

A white or almost white, crystalline powder, practically insoluble in water, freely soluble in acetone, in alcohol and in methylene chloride.

Примеси

Ketoprofen

The chromatographic procedure for Related substances may be carried out using:

— a stainless steel column 0.15 m long and 4.6 mm in internal diameter packed with a spherical octadecylsilyl silica gel for chromatography R (5 µm) with a specific surface of 350 m2·g-1 and a pore size of 0.01 µm (10 nm), — as mobile phase at a flow rate of 1 ml per minute a mixture of 2 volumes of phosphate buffer solution pH 3.5 R, freshly prepared, 43 volumes of acetonitrile R and 55 volumes of water R,

— as detector a spectrophotometer set at 233 nm,

— a loop injector.

Inject 20 µl of reference solution (d). The substances are eluted in the following order: ketoprofen and ketoprofen impurity A (3-acetylbenzophenone). Adjust the sensitivity of the detector so that the heights of the two principal peaks in the chromatogram obtained are not less than 50 per cent of the full scale of the recorder. The test is not valid unless the resolution between the peaks corresponding to ketoprofen and ketoprofen impurity A is at least 7.0.

Ketoprofen

Ibuprofen

Ibuprofen contains not less than 98.5 per cent and not more than the equivalent of 101.0 per cent of (RS)-2-(4- isobutylphenyl) propionic acid, calculated with reference to the dried substance.

A white, crystalline powder or colourless crystals, practically insoluble in water, freely soluble in acetone, in ether, in methanol and in methylene chloride. It dissolves in dilute solutions of alkali hydroxides and carbonates.

Ibuprofen

IDENTIFICATION

First identification: A, C.

Second identification: A, B, D.

A. Melting point (2.2.14): 75°C to 78°C.

B. Dissolve 50.0 mg in a 4 g/l solution of sodium hydroxide R and dilute to 100.0 ml with the same alkaline solution. Examined between 240 nm and 300 nm (2.2.25), using a spectrophotometer with a band width of 1.0 nm and a scan speed of not more than 50 nm per minute, the solution shows a shoulder at 258 nm and two absorption maxima, at 264 nm and 272 nm. The ratio of the absorbance measured at the maximum at 264 nm to that measured at the shoulder at 258 nm is 1.20 to 1.30. The ratio of the absorbance measured at the maximum at 272 nm to that measured at the shoulder at 258 nm is 1.00 to 1.10.

C. Examine by infrared absorption spectrophotometry (2.2.24), comparing with the spectrum obtained with ibuprofen CRS. Examine the substances prepared as discs.

D. Examine by thin-layer chromatography (2.2.27), using silica gel H R as the coating substance.

Ibuprofen

ОСНОВНИ СТРУКТУРИ

NNH

pirazole

1

2

3

NNH

1

2

3

3 pirazoline

NNH

1

2

3

H H

pirazolidine

NNH

1

2

3

H

O

1,2 dihydro 3 H pyrazol

NN1

2

3

H

O

HO

py3 one razolidine 3,5 dione

Пиразолинови и пиразолидинови производни

Metamizole sodium

Metamizole sodium contains not less than 99.0 and not more than the equivalent of 100.5 per cent of sodium [(1,5-dimethyl-3-oxo-2- phenyl-2,3-dihydro-1H -pyrazol-4-yl)-N- methylamino]methanesulphonate, calculated with reference to the dried substance.

A white or almost white, crystalline powder, very soluble in water, soluble in alcohol.

Metamizol sodium

H+

NN

CH3

H3C

N

CH3

OHO3S

+ H2O H2SO3

NN

CH3

H3C

N

CH3

OHO

+H2O HO3SCH2OH

NN

CH3

H3C

N

CH3

OHHCHO

NN

CH3

H3C

N

CH3

NaO3 S O

H2O

H2O

NN

CH3

H3C

N

CH3

O H+ HCHO + NaHSO3H

NN

CH3

H3C

N

CHOOH

NN

CH3

H3C

N OH2

O2/Cu2+

O2/Cu2+

NN

CH3

H3C

OHOO

A. R at C4 = NHCHO: 4-formylamino-1,5-dimethyl-2-phenyl-1,2- dihydro-3 H-pyrazol-3-one,

B. R at C4 = NH2: 4-amino-1,5-dimethyl-2-phenyl-1,2-dihydro-3H - pyrazol-3-one,

C. R at C4 = NHCH3: 4-methylamino-1,5-dimethyl-2-phenyl-1,2- dihydro-3 H-pyrazol-3-one,

D. R at C4 = N(CH3)2: 4-dimethylamino-1,5-dimethyl-2-phenyl-1,2- dihydro-3 H-pyrazol-3-one.

Metamizole sodiumImpurities

Metamizole sodium

Related substances Examine by liquid chromatography (2.2.29).

Prepare the solutions immediately before use. Test solution. Dissolve 50.0 mg of the substance to be examined in methanol R and dilute to 10.0 ml with the same solvent. Reference solution (a). Dissolve 10.0 mg of metamizole impurity A CRS in methanol R and dilute to 20.0 ml with the same solvent. Reference solution (b). Dilute 1.0 ml of reference solution (a) to 20.0 ml with methanol R. Reference solution (c). Dissolve 40 mg of metamizole sodium CRS in methanol R and dilute to 20.0 ml with the same solvent. Reference solution (d). Take 10 ml of reference solution (c) and boil under a reflux condenser for 10 min. Allow to cool to room temperature and dilute to 20.0 ml with methanol R. Reference solution (e). To 6 ml of reference solution (a) add 1 ml of reference solution (c).

The chromatographic procedure may be carried out using: — a stainless steel column 0.25 m long and 4.6 mm in internal diameter packed with base-deactivated octadecylsilyl silica gel for chromatography R (5 µm), — as mobile phase at a flow rate of 1.0 ml/min a mixture of 28 volumes of methanol R and 72 volumes of a buffer solution prepared by adjusting a mixture of 1000 volumes of a 6.0 g/l solution of sodium dihydrogen phosphate R and 1 volume of triethylamine R to pH 7.0 with strong sodium hydroxide solution R, — as detector a spectrophotometer set at 254 nm.

Metamizole sodium

When the chromatograms are recorded in the prescribed conditions, the substances elute in the following order: impurity A, metamizole, impurity B, impurity C and impurity D. Inject 10 µl of reference solution (b). Adjust the sensitivity of the system so that the height of the principal peak in the chromatogram obtained is at least 50 per cent of the full scale of the recorder.

Inject 10 µl of reference solution (d). The chromatogram shows two principal peaks due to metamizole and impurity C.

Inject 10 µl of reference solution (e). The test is not valid unless in the chromatogram obtained the resolution between the peaks corresponding to impurity A and metamizole is at least 2.5.

Inject 10 µl of the test solution and 10 µl of reference solution (b) and continue the chromatography for 3.5 times the retention time of metamizole. In the chromatogram obtained with the test solution: the area of any peak corresponding to impurity C is not greater than the area of the principal peak in the chromatogram obtained with reference solution (b) (0.5 per cent), the area of any peaks, apart from the principal peak and the peak due to impurity C is not greater than 0.4 times the area of the principal peak in the chromatogram obtained with reference solution (b) (0.2 per cent). The sum of the areas of all the peaks, apart from the principal peak, is not greater than the area of the principal peak in the chromatogram obtained with reference solution (b) (0.5 per cent). Disregard any peak with an area less than 0.05 times that of the principal peak in the chromatogram obtained with the reference solution (b).

Metamizole sodium

Dissolve 0.200 g in 10 ml of 0.01 M hydrochloric acid previously cooled in iced water and titrate immediately, dropwise, with 0.05 M iodine. Before each addition of 0.05 M iodine dissolve the precipitate by swirling. At the end of the titration add 2 ml of starch solution R and titrate until the blue colour of the solution persists for at least 2 min. The temperature of the solution during the titration must not exceed 10°C.

1 ml of 0.05Miodine is equivalent to 16.67 mg of C13H16N3NaO4S.

Phenylbutazone contains not less than 99.0 per cent and not more than the equivalent of 101.0 per cent of 4-butyl-1,2- diphenyl-pyrazolidine-3,5-dione, calculated with reference to the dried substance.

CHARACTERS A white or almost white, crystalline powder, practically insoluble in water, soluble in ether, sparingly soluble in alcohol. It dissolves in alkaline solutions.

Phenylbutazone

A. Melting point (2.2.14): 104°C to 107°C. B. Dissolve 30.0 mg in 25 ml of methanol R, add 50 ml of 1M sodium hydroxide and dilute to 100.0 ml with water R. Dilute 5.0 ml of the solution to 250.0 ml with water R. Examined between 240 nm and 350 nm (2.2.25), the solution shows an absorption maximum at 264 nm. The specific absorbance at the maximum is 650 to 700. Use as the compensation liquid a mixture of 0.5 ml of methanol R, 1.0 ml of 1M sodium hydroxide and 98.5 ml of water R. C. Examine by infrared absorption spectrophotometry (2.2.24), comparing with the spectrum obtained with phenylbutazone CRS. D. To 0.1 g add 1 ml of glacial acetic acid R and 2 ml of hydrochloric acid R and heat the mixture under a reflux condenser for 30 min. Cool, add 10 ml of water R and filter. To the filtrate add 3 ml of a 7 g/l solution of sodium nitrite R. A yellow colour is produced. To 1 ml of the solution add a solution of 10 mg of b- naphthol R in 5 ml of sodium carbonate solution R. A reddish- brown to reddish-violet precipitate is forme.

Phenylbutazone

H2O + H2O

COOH

NNC6H5

HC6H5

n C4H9

CO2 t

NNC6H5

HC6H5

n C4H9

O

O

H3O+

CH COOH

HOOC

n C4H9

+ HN

NH

C6H5

C6H5

CO2 t

n C4H9 CH2 COOH

Dissolve 0.500 g in 25 ml of acetone R and add 0.5 ml of bromothymol blue solution R1. Titrate with 0.1M sodium hydroxide until a blue colour is obtained which persists for 15 s. Carry out a blank titration.

1 ml of 0.1 M sodium hydroxide is equivalent to 30. 84 mg of C19H20N2O2.

Phenylbutazone

SH2N

O O

NN

CF3

H3C

Celecoxib

4-[5-(4-Methylphenyl)-3-(trifluoromethyl)-1H -pyrazol-1-yl]benzene sulfonamide

Pale yellow solid, mp 157 -159 degrees.

Piroxicam contains not less than 98.5 per cent and not more than the equivalent of 101.0 per cent of 4-hydroxy-2-methyl-N-(pyridin-2-yl)-2H-1,2-benzothiazine-3-carboxamide 1,1-dioxide, calculated with reference to the dried substance.

Piroxicam

• Жълтеникав кристален прах

• неразтворим във вода, лесно разтворим в ацетон, трудно разтворим в етанол

• има полиморфни форми

• т.т. около149°С

• pKa=6.5

Nimesulide

N-(4-Nitro-2-phenoxyphenyl)methanesulphonamide

ASSAY Dissolve 0.240 g in 30 ml of previously neutralised acetone R and add 20 ml of water R. Titrate with 0.1 M sodium hydroxide, determining the end-point potentiometrically (2.2.20). 1 ml of 0.1 M sodium hydroxide is equivalent to 30.83 mg of C13H12N2O5S.

A: R1 = SO2-CH3, R2 = H, R3 = R4 = NO2 N-(2,4-dinitro-6-phenoxyphenyl) methane-sulphonamide;B: R1 = SO2-CH3, R2 = R3 = R4 = H N-(2-phenoxyphenyl) methane-sulphonamideC: R1 = R2 = R3 = R4 = H2-phenoxyanilineD: R1 = R2 = R4 = H, R3 = NO24-nitro-2-phenoxyanilineE: R1 = R2 = SO2-CH3, R3 = R4 = H N,N-bis(methylsulphonyl)-2-phenoxyanilineF: R1 = R2 = SO2-CH3, R3 = NO2, R4 = H N,N-bis(methylsulphonyl)-4-nitro-2-phenoxyaniline

Nimesulide

Nimesulide

• C18 3.5 µm (150 x 4.6 mm i.d.) заедно с C18 guard column (10 x 2.1 mm i.d.)

• подвижна фаза – ацетонитрил : 50 mM натриево цитратен буфер, рН=3(53:47)

• скорост – 1.1 ml/min• UV-детектор при 240 nm

• Hypersil BDS-C18 column• подвижна фаза – 1g/l

динатриевхидрогенфосфат (pH=7.6) : метанол (56:44)

• UV-детектор при 302 nm

• 50mmx4.6mm i.d. monolithic column• подвижна фаза – ацетонитрил :

фосфатен буфер, pH=7; 10mM (34:66, v/v)

• скорост – 4.0 ml/min

• Supersphere Select B, 5µM, 125 x 3mm i.d.

• подвижна фаза – 1000g метанол, 650g ацетонитрил, 1650g вода, 10g динатриевхидрогенфосфат, 4g хептан сулфонова киселина. pH=5.5 (чрез фосфорна киселина 85%)

• скорост – 0.7 ml/min• UV-детектор при 297 nm

Omega-3 fatty acids are a family of polyunsaturated fatty acids which have in common a carbon-carbon double bond in the ω-3 position.

Omega-3 fatty acids

Alpha-linolenic acid

Eicosapentaenoic acid

Docosahexaenoic acid

Omega-3 fatty acids

Column 1:  —dimensions: l = 0.3 m, Ø = 7.8 mm; —stationary phase: styrene-divinylbenzene copolymer R (7 µm), with a pore size of 10 nm. Columns 2 and 3 placed closest to the injector: —dimensions: l = 0.3 m, Ø = 7.8 mm; —stationary phase: styrene-divinylbenzene copolymer R (7 µm), with a pore size of 50 nm. Mobile phase tetrahydrofuran R. Flow rate 0.8 ml/min. Detection Differential refractometer. Injection 40 µl.

Omega-3 fatty acids

Divinylbenzene

Styrene (Vinyl benzene) can be copolymerized with other monomers; for example, divinylbenzene for cross-linking the

polystyrene chains

(Ph. Eur. method 2.2.30) Size-exclusion chromatography is a chromatographic technique which separates molecules in solution according to their size. With organic mobile phases, the technique is known as gel-permeation chromatography and with aqueous mobile phases, the term gel-filtration chromatography has been used. The sample is introduced into a column, which is filled with a gel or a porous particle packing material, and is carried by the mobile phase through the column. The size separation takes place by repeated exchange of the solute molecules between the solvent of the mobile phase and the same solvent in the stagnant liquid phase (stationary phase) within the pores of the packing material. The pore-size range of the packing material determines the molecular-size range within which separation can occur.

Omega-3 fatty acids

Size-exclusion – изключваща по размер

gel-permeation – гел проникваща

gel-filtration – гел филтрираща