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SAMPLE
Matrix: blood
Sample p repa ra ti o n : 1 mL Serum + 3 mL MTBE, vortex for 1 min, centrifuge at 1500
rpm for 5 min. Remove the organic layer and evaporate it to dryness under a stream of
nitrogen at room temperature, reconstitute the residue in 100 jxL MeOH, inject an aliquot.
HPLCVARIABLES
G uar d co lum n: 18 mm long (Brownlee)
Column: 300 X 3.6 10 |xm jxBondapak C18
Mobile ph as e: MeOH: water 80:20
Flow rate: 1
Dete cto r: UV 254; RIA
C H R O M A T O G R A M
Retention time: 5.6
OTHER SUBSTANCES
Extracted: metabolites, norgestimate
KEYWORDS
serum
REFERENCE
Wong, F.A.; Juzw in, S.J.; Tischio, N.S.; Flor, S.C. Determ ination of norgestim ate in serum by a utom ated
high-performance liquid chromatography and subsequent radioimmunoassay. J.Liq.Chromatogr.
1995,28, 1851-1861
SAMPLE
Matrix: bulk
HPLCVARIABLES
Colu mn: 250 X 4 10 |xm LiChrosorb RP-18
Mobile ph as e: MeOH: water 70:30
Flow rate: 1
Injection volume: 25
D etecto r: UV 240
C H R O M A T O G R A M
Retention t ime: 9
OTHER SUBSTANCES
Simultaneous: impurities, norethindrone
Interfering: ethinyl estradiol
REFERENCE
Gorog, S.; Hereny i, B. An alysis of stero ids. XXXVIII. The u se of high-perform ance liquid chro m atog raph y
with diode-array UV detection for es tim ating imp urity profiles of steroid drug s. J.Chromatogr. 1987,
400
177-186
Norgestrel
Molecular formula: C
21
H
28
O
2
Molecular weight: 312.5
CAS Registry No :
797-63 -7, 797-64-8 -) form), 6533-00 -2
Previous page
http://81765_13b.pdf/http://81765_13b.pdf/
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SAMPLE
Matrix: culture medium
Sample preparat ion: Extract culture medium twice with 2 volumes of ether, combine the
extracts and evaporate them to dryness, reconstitute with MeOH, inject an aliquot.
HPLCVARIABLES
Column: 300 X 3.9 Techopak 10 C18 (HPLC Technology)
Mobile phase:
MeOH: water 70:30
F low rate: 1.5
Detector:
UV 240; Radioactivity
CHROMATOGRAM
Retent ion t ime: 5
OTHER SUBSTANCES
Extracted:
metabolites, norgestimate
KEYWORDS
tritium labeled
REFERENCE
Wild, M.J.; Rudland, RS.; Back, D.J. Metabolism of the oral contraceptive steroids ethynylestradiol and
norge stima te by norm al (Hum a 7) and m align ant (MCF-7 and ZR-75-1) hu m an b rea st cells in culture.
J.Steroid Biochem.Mol.BioL 1991, 39 5 3 5 - 5 4 3
SAMPLE
Matrix:
formulations
Sample preparat ion: Centrifuge oil formulation at 30° at 2000 rpm for 30 min, filter
(W hatman No. 1 paper), collect the last 4 mL of th e n itra te. D ilute a 10 |xL aliquot to 10
mL with M eCN : w ater 60 :40 containin g 0.3% Tween 80, remove a 2 mL aliquot a nd add
it to 1 mL 3.33 |xg/mL progesterone, vortex for 10 s, inject a 50 |xL aliquot.
HPLCVARIABLES
Column:
300 X 3.9 Novapak C18
Mobile phase: MeCN: water 60:40
Flow rate: 2
Inject ion volume: 50
Detector: UV
248
CHROMATOGRAM
Internal s tandard:
progesterone
KEYWORDS
oils
REFERENCE
Gao, Z.-H.; Shu kla, A. J.; Joh nso n, J.R.; Crowley, W.R. Controlled re leas e of a con tracep tive ste roid from
biodegradable and injectable gel formulations: In vitro evaluation. Ph arm.Res. 1995, 12 857—863
SAMPLE
Matrix:
formulations
Sample preparat ion: Oils. 1 mL Sam ple + 25 mL MeO H-.water 90: 10 , sha ke vigorously
for 5 min, centrifuge, inject a 10
|JLL
aliquot of the supernatant. Tablets. Grind a tablet
to a fine powder, add 25 mL MeOH, sonicate for 5-10 min, filter (0.45 |min), discard first
5 mL of filtrate, inject a 10 |xL aliquot of the remaining filtrate. Suspensions (aqueous).
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Make up 5 mL to 50 mL with MeOH, filter (0.45 jjim), discard first 5 mL of filtra te, inject
a 10 |xL aliquot of the remaining filtrate.
HPLCVARIABLES
Column: 250 X 4.6 5 |xm Zorbax ODS
Mobile phase:
MeOH: water 75:25
Flow rate: 1.5
Injection volume: 10
Detector: UV
240
CHROMATOGRAM
Retention time: 5.9
Limit of detection:
5 |xg/mL
OTHER SUBSTANCES
Simultaneous:
aspirin, benzyl alcohol, benzyl benzoate, boldenone, caffeine, caluster6ne,
cortisone, dehydroepiandrosterone (UV 210), fluoxym esterone, formebolone, mesterolone
(UV 210), methandriol (UV 210), methenolone acetate, methyltestosterone, mibolerone,
nandrolone acetate, nandrolone propionate, norethandrolone, norethindrone, norethin-
drone acetate, oxandrolone (UV 210), oxymetholone, stanozolol, testolactone, testosterone
acetate, testosterone propionate, trenbolone acetate
Interfering:
ethisterone, methandrostenolone, nandrolone, testosterone
KEYWORDS
oils;
tablets; suspensions
REFERENCE
Walters, M.J.; Ayers, R.J.; Brown, D.J. Analysis of illegally distributed anabolic steroid products by
liquid chromatography with identity confirmation by mass spectrometry or infrared spectrophotom-
etry J.Assoc.OffAnaLChem. 1990, 73 904-926
SAMPLE
Matrix:
microsomal incubations, mucosal fluid
Sample preparation:
Mucosal fluid . Extract 1 mL mucosal fluid twice with 5 mL diethyl
ether, evaporate extracts to dryness, resuspend residue in 100 |xL MeOH, inject an ali-
quot. Microsomal incubations. Extract 2.5 mL microsomal preparation with 5 mL diethyl
ether, proceed as before.
HPLCVARIABLES
Guard column:
on-line guard column
Column:
100 X 8 ixBondapak radial compression module
Mobile phase: MeOH: water 70:30
Flow rate: 1.5
Injection volume:
100
Detector: UV 240
CHROMATOGRAM
Retention time: 6
OTHER SUBSTANCES
Simultaneous:
3-ketonorgestimate, 17-deacetylnorgestimate, norgestimate
REFERENCE
Madden, S.; Back, D.J. Metabolism of norgestimate by human gastrointestinal mucosa and liver micro-
somes in vitro. J.Steroid Biochem.Mol.Biol. 1991, 38 4 9 7 - 5 0 3
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SAMPLE
Matrix: solutions
HPLC VARIABLES
Column: 50 X 4.6 5 |xm Supelcosil LC-18
Mobile phase: MeOH: THF: water 10:20:70
Flow rate: 2
Injection volume: 20
Detector: UV
220
CHROMATOGRAM
Retention time:
5.5
OTHER SUBSTANCES
Simultaneous:
ethinyl estradiol, norethindrone, norethindrone acetate, norethynodrel
acetate
REFERENCE
Supelco Catalog Supelco Inc. Bellefonte PA
1996, p. A130
SAMPLE
Matrix: solutions
Sample preparation:
Inject an aliquot of a 10 |xg/mL solution.
HPLC VARIABLES
Column:
150
X 4.6
Supelco
ODS
Mobile phase: MeCN: water 25:75 containing 14 mM cyclodextrin
Column temperature: 0
Flow rate: 1 3
Injection volume: 20
Detector: UV 240
CHROMATOGRAM
Retention time: 16 D +)), 18 D -))
KEYWORDS
chiral
REFERENCE
Lamparczyk, H.; Zarzycki, RK.; Nowakowska, J. Effect of temperature on separation of norgestrel en-
antiomers
by
high-performance liquid chromatography. J.Chromatogr.A
1994,
668 413-417
SAMPLE
Matrix: solutions
Sample preparation: Direct injection.
HPLCVARIABLES
Column:
250
X 4.6 10
m Partisil
C 18 ODS 3
Mobile phase: MeCN-.water 50:50
Flow rate: 2
Detector: UV 243
CHROMATOGRAM
Retention time: 6.0
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KEYWORDS
see also J.Pharm.Sci. 1989, 78, 477
REFERENCE
Catz, P.; Friend, D.R. In vitro evaluations of transdermal levonorgestrel.
D rug D es.Deliv.
1990 6 4 9 -
60
SAMPLE
Matrix: solutions
HPLCVARIABLES
Column: 250 X 4.6 10 jim Partisil ODS-3 C18
Mobile phase: MeCN-.water 50:50
Flow rate: 2
D etecto r: UV 243
C H R O M A T O G R A M
Retention time: 6.0
REFERENCE
Friend, D.R.; C atz, P.; Heller, J.; O kagaki, M. Tran sderm al delivery of levonorgestrel IV : Ev aluation of
membranes.
J.Pharm.Sci.
1989
78
477-480
SAMPLE
Matrix: solutions
HPLCVARIABLES
Guard column: Chromsep reverse phase guard column (Chrompack)
Column: 100 X 3 5 |xm Chromspher glass column
M obile ph as e: MeCN: water 35:65
Flow rate: 0.4
Detec tor: UV 247
C H R O M A T O G R A M
Retention time: 20
OTHER SUBSTANCES
Sim ul tane ou s: nandrolone, trenbolone (at UV 340)
REFERENCE
Haa snoot, W.; Schilt, R.; Ha m ers , A.R.; Huf, F.A.; Farjam , A.; Fre i, R.W.; B rin km an , U.A. Dete rm ina tion
of p-19-nortestosterone a nd its metab olite a-19-nortestosterone in biological samples at th e sub pa rts
per billion level by high-performance liquid chromatography with on-line immunoaffinity sample
pretreatment. J.Chromatogr 1989 489 157-171
SAMPLE
Matrix: solutions
HPLCVARIABLES
Colum n: 250 X 4.6 10 jxm Nucleosil C18
Mobile pha se : MeCN:THF: water 12.9:22.4:64.7
Flow rate: 1
D etecto r: UV 240
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C H R O M A T O G R A M
Retention t ime: 13
OTHER SUBSTANCES
Sim ul tane ous : estrone, ethinyl estradiol, mestranol, norethindrone, norethindrone acetate
REFERENCE
Gazdag, M.; Szepesi, G.; Szeleczki, E. Selection of high-performance liquid chromatographic methods in
pharmaceutical analysis. I. Optimization for selectivity in reversed-phase chromatography.
J.Chromatogr.
1988
454
8 3 - 9 4
SAMPLE
Matrix: solutions
HPLCVARIABLES
Co lumn: 250 X 4.6 5
\xm
LiChrosorb Si 60
Mobile phase : Hexane: dioxane: isopropanol 95 : 3:2
Flow rate: 1
D etec tor: UV 254
C H R O M A T O G R A M
Retention t ime: 13
OTHER SUBSTANCES
Sim ul tane ou s: estrone, ethinyl estradiol, mestranol, norethindrone, norethindrone acetate
KEYWORDS
normal phase
REFERENCE
Gazdag, M.; Szepesi, G.; Fabian-Varga, K. Selection of high-performance liquid chro ma tographic m eth-
ods in pharmaceutical analysis. II. Optimization for selectivity in normal-phase systems.
J.Chromatogr.
1988
454 9 5 - 1 0 7
SAMPLE
Matrix: solutions
HPLCVARIABLES
Colum n: 250 X 4.6 jmBondapak C18
M obile phase: Dioxane: water 50:50 (Caution Dioxane is a carcinogen )
Flow rate: 1.4
D etec tor: UV 254
OTHER SUBSTANCES
Simultaneous: norgestimate
REFERENCE
Killinger, J.; Ha hn, D.W.; Phillips, A.; Hetyei, N.S.; McGuire, J.L. T he affinity of norgestim ate for u ter ine
progestogen receptors and its direct action on the uterus.
Contraception
1985,
32
311-319
SAMPLE
Matrix: tissue
Sa m ple p re pa ra ti o n : Incubate endometrial tissue with buffer, remove tissue, extract me-
dium twice with 2 volumes of diethyl ether, evaporate to dryness, reconstitute in a small
volume of MeOH, inject an aliquot.
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HPLC VARIABLES
Column: 300 X 3.9 Technopak 10 C18
Mobile phase: MeOH: water 70:30
Flow rate: 1.5
Detector: UV
240
CHROMATOGRAM
Retention time:
OTHER SUBSTANCES
Simultaneous:
metabolites, norgestimate
KEYWORDS
endometrial tissue
REFERENCE
Wild,
M.J.;
Rudland,
RS.;
Back, D.J. M etabolism of the oral contraceptive steroids ethyny lestradiol,
norgestimate and 3-ketodeso gestrel by a hu m an endo me trial cancer cell line (HEC-IA) and endo-
metrial tissue in
vitro. J.Steroid Biochem .Mol.BioL
1993,
45
407-420
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