OXYTOCIN Oxytocinum

Oxytocin EUROPEAN PHARMACOPOEIA 5.0
Injection: test solution and reference solution (a).
Calculate the percentage content of C 22
H 25
ClN 2
O 9
taking
1 mg of oxytetracycline as equivalent to 1.079 mg of
oxytetracycline hydrochloride.
STORAGE
In an airtight container, protected from light. If the
substance is sterile, store in a sterile, airtight, tamper-proof
container.
LABELLING
The label states, where applicable, that the substance is free
from bacterial endotoxins.
IMPURITIES
A. R1 = NH 2
,R2=N(CH 3
) 2
,R3=R4=H,R5=OH:
(4R,4aR,5S,5aR,6S,12aS)-4-(dimethylamino)-3,5,6,10,12,
12a-hexahydroxy-6-methyl-1,11-dioxo-1,4,4a,5,5a,6,11,12aoctahydrotetracene-2-carboxamide
(4-epioxytetracycline),
B.R1=NH 2
,R2=R4=R5=H,R3=N(CH 3
) 2
: tetracycline,
C. R1 = CH 3
,R2=R4=H,R3=N(CH 3
) 2
,R5=OH:
(4S,4aR,5S,5aR,6S,12aS)-2-acetyl-4-(dimethylamino)-
3,5,6,10,12,12a-hexahydroxy-6-methyl-4a,5a,
6,12a-tetrahydrotetracene-1,11(4H,5H)-dione
(2-acetyl-2-decarbamoyloxytetracycline),
D. R = OH, R′ =H:(3S,4S,5S)-4-[(1R)-4,5-dihydroxy-9-
methyl-3-oxo-1,3-dihydronaphtho[2,3-c]furan-1-yl]-
3-(dimethylamino)-2,5-dihydroxy-6-oxocyclohex-1-
enecarboxamide (α-apo-oxytetracycline),
E. R = H, R′ =OH:(3S,4S,5R)-4-[(1R)-4,5-dihydroxy-9-
methyl-3-oxo-1,3-dihydronaphtho[2,3-c]furan-1-yl]-
3-(dimethylamino)-2,5-dihydroxy-6-oxocyclohex-1-
enecarboxamide (β-apo-oxytetracycline),
F. (4S,4aR,5R,12aS)-4-(dimethylamino)-3,5,10,11,
12a-pentahydroxy-6-methyl-1,12-dioxo-1,4,4a,
5,12,12a-hexahydrotetracene-2-carboxamide
(anhydro-oxytetracycline).
OXYTOCIN
Oxytocinum
01/2005:0780
C 43
H 66
N 12
O 12
S 2
M r
1007
DEFINITION
Oxytocin is a cyclic nonapeptide having the structure of the
hormone produced by the posterior lobe of the pituitary
gland that stimulates contraction of the uterus and milk
ejection in receptive mammals. It is obtained by chemical
synthesis and is available in the freeze-dried form as an
acetate. It contains not less than 93.0 per cent and not
more than the equivalent of 102.0 per cent of the peptide
C 43
H 66
N 12
O 12
S 2
, calculated with reference to the anhydrous,
acetic acid-free substance.
By convention, for the purpose of labelling oxytocin
preparations, 1 mg of oxytocin peptide (C 43
H 66
N 12
O 12
S 2
)is
equivalent to 600 IU of biological activity.
CHARACTERS
A white or almost white powder, hygroscopic, very soluble in
water and in dilute solutions of acetic acid and of ethanol.
IDENTIFICATION
Examine the chromatograms obtained in the assay. The
retention time of the principal peak in the chromatogram
obtained with the test solution is approximately the same
as that of the principal peak in the chromatogram obtained
with the reference solution.
TESTS
pH (2.2.3). Dissolve 0.200 g in carbon dioxide-free water R
and dilute to 10.0 ml with the same solvent. The pH of the
solution is 3.0 to 6.0.
Amino acids. Examine by means of an amino-acid analyser.
Standardise the apparatus with a mixture containing
equimolar amounts of ammonia, glycine and the L-form of
the following amino acids:
lysine threonine alanine leucine
histidine serine valine tyrosine
arginine glutamic acid methionine phenylalanine
aspartic acid proline isoleucine
together with half the equimolar amount of L-cystine. For the
validation of the method, an appropriate internal standard,
such as DL-norleucine R, isused.
Test solution. Place 1.0 mg of the substance to be examined
in a rigorously cleaned hard-glass tube 100 mm long and
6 mm in internal diameter. Add a suitable amount of a
50 per cent V/V solution of hydrochloric acid R. Immerse
thetubeinafreezingmixtureat−5 °C, reduce the pressure
to below 133 Pa and seal. Heat at 110 °C to 115 °C for 16 h.
Cool, open the tube, transfer the contents to a 10 ml flask
with the aid of five quantities, each of 0.2 ml, of water R and
evaporate to dryness over potassium hydroxide R under
reduced pressure. Take up the residue in water R and
evaporate to dryness over potassium hydroxide R under
reduced pressure; repeat these operations once. Take up
the residue in a buffer solution suitable for the amino-acid
analyser used and dilute to a suitable volume with the same
buffer solution. Apply a suitable volume to the amino-acid
analyser.
2174 See the information section on general monographs (cover pages)

EUROPEAN PHARMACOPOEIA 5.0
Oxytocin bulk solution
Express the content of each amino acid in moles. Calculate
the relative proportions of the amino acids, taking one-sixth
of the sum of the number of moles of aspartic acid, glutamic
acid, proline, glycine, isoleucine and leucine as equal to one.
The values fall within the following limits: aspartic acid 0.95
to 1.05; glutamic acid 0.95 to 1.05; proline 0.95 to 1.05;
glycine 0.95 to 1.05; leucine 0.90 to 1.10; isoleucine 0.90 to
1.10; tyrosine 0.7 to 1.05; half-cystine 1.4 to 2.1; not more
than traces of other amino acids are present.
Related peptides. Examinebyliquidchromatography
(2.2.29) as described under Assay.
Inject 50 µl of the test solution. In the chromatogram
obtainedtheareaofanypeak,apartfromtheprincipal
peak, is not greater than 1.5 per cent of the total area of the
peaks; the sum of the areas of all the peaks, apart from the
principal peak, is not greater than 5 per cent of the total
area of the peaks. Disregard any peak due to the solvent
and any peak with an area less than 0.1 per cent of that of
the principal peak.
Acetic acid (2.5.34): 6.0 per cent to 10.0 per cent.
Test solution. Dissolve15.0mgofthesubstancetobe
examined in a mixture of 5 volumes of mobile phase B and
95 volumes of mobile phase A and dilute to 10.0 ml with the
same mixture of solvents.
Water (2.5.12). Not more than 5.0 per cent, determined on
at least 50 mg by the semi-micro determination of water.
Bacterial endotoxins (2.6.14): less than 300 IU/mg, if
intended for use in the manufacture of parenteral dosage
forms without a further appropriate procedure for removal
of bacterial endotoxins.
ASSAY
Examine by liquid chromatography (2.2.29).
Test solution. Prepare a 0.25 mg/ml solution of the
substance to be examined in a 15.6 g/l solution of sodium
dihydrogen phosphate R.
Reference solution. Dissolve the contents of a vial of
oxytocin CRS in a 15.6 g/l solution of sodium dihydrogen
phosphate R to obtain a concentration of 0.25 mg/ml.
Resolution solution. Dissolve the contents of a vial of
oxytocin/desmopressin validation mixture CRS in 500 µl
of a 15.6 g/l solution of sodium dihydrogen phosphate R.
The chromatographic procedure may be carried out using:
— a stainless steel column 0.125 m long and 4.6 mm in
internal diameter packed with octadecylsilyl silica gel for
chromatography R (5 µm),
— asmobilephaseataflowrateof1ml/min:
Mobile phase A. A 15.6 g/l solution of sodium
dihydrogen phosphate R,
Mobile phase B. Mix 1 volume of acetonitrile for
chromatography R with 1 volume of water R,
Time
(min)
Mobile phase A
(per cent V/V)
Mobile phase B
(per cent V/V)
Comment
0 - 30 70 → 40 30 → 60 linear gradient
30 - 30.1 40 → 70 60 → 30 switch to initial eluent
composition
30.1 - 45 70 30 re-equilibration
— as detector a spectrophotometer set at 220 nm.
Equilibrate the column with amixtureof30volumesof
mobile phase B and 70 volumes of mobile phase A.
Inject 25 µl of the resolution solution. When the
chromatograms are recorded in the prescribed conditions,
the retention times are: oxytocin about 7.5 min and
desmopressin about 10 min. The test is not valid unless the
resolution between the peaks corresponding to desmopressin
and oxytocin is at least 5.0.
Inject 25 µl of the test solution and 25 µl of the reference
solution.
Calculate the content of oxytocin (C 43
H 66
N 12
O 12
S 2
)fromthe
peak areas in the chromatograms obtained with the test
solutionandthereferencesolutionandthedeclaredcontent
of C 43
H 66
N 12
O 12
S 2
in oxytocin CRS.
STORAGE
Store in an airtight container, protected from light, at a
temperature of 2 °C to 8 °C. If the substance is sterile, store
in a sterile, airtight, tamper-proof container.
LABELLING
The label states:
— the oxytocin peptide content (C 43
H 66
N 12
O 12
S 2
),
— where applicable, that the substance is free from bacterial
endotoxins.
01/2005:0779
OXYTOCIN BULK SOLUTION
Oxytocini solutio
DEFINITION
Oxytocin bulk solution is a solution of oxytocin, a cyclic
nonapeptide having the structure of the hormone produced
by the posterior lobe of the pituitary gland that stimulates
contraction of the uterus and milk ejection in receptive
mammals. It is obtained by chemical synthesis. It is available
as a bulk solution with a stated concentration of not less
than 0.25 mg of oxytocin per millilitre, in a solvent that
may contain an appropriate antimicrobial preservative. It
contains not less than 95.0 per cent and not more than
105.0percentoftheamountofthepeptideC 43
H 66
N 12
O 12
S 2
stated per millilitre.
By convention, for the purpose of labelling oxytocin
preparations, 1 mg of oxytocin peptide (C 43
H 66
N 12
O 12
S 2
)is
equivalent to 600 IU of biological activity.
CHARACTERS
A clear, colourless liquid.
IDENTIFICATION
Examine the chromatograms obtained under Assay. The
retention time of the principal peak in the chromatogram
obtained with the test solution is similar to that of the
principal peak in the chromatogram obtained with the
reference solution.
TESTS
pH (2.2.3).ThepHofthepreparationtobeexaminedis3.0
to 5.0.
Amino acids. Examine by means of an amino-acid analyser.
Standardise the apparatus with a mixture containing
equimolar amounts of ammonia, glycine and the L-form of
the following amino acids:
lysine threonine alanine leucine
histidine serine valine tyrosine
arginine glutamic acid methionine phenylalanine
aspartic acid proline isoleucine
GeneralNotices(1)applytoallmonographsandothertexts 2175