Application Note AN 102-C
Pharmaceutical Purity Testing by OPLC
A
Nandrolone is a steroid that is used in the total synthesis of a number of norsteroids. Since its impurity profile strongly influences the composition of the final product, it bstract is necessary to obtain a good understanding of the impurities that are present. OPLC can provide an efficient and rapid test for the purity of a drug that is capable of indicating the presence of a broad range of impurities. The number of compounds that can be determined is significantly greater than the one resulting from TLC, GC or HPLC separations which are commonly used in process laboratories. In addition, it is important to note that OPLC requires less mobile phase and takes less time than HPLC. OPLC …
… Compared to HPLC and to TLC
Bagocsi et al.1 have compared a variety of chromatographic methods for the testing of impurities of Nandrolone. These workers demonstrated that a simple separation of the compounds of interest was obtained using OPCL using silica gel plates with a cyclohexane: ethyl acetate:chloroform (2:1:1) as shown in Figure 1. The total analysis time for simultaneous separation of numerous samples was of 40 minutes.
TLC is a low resolution technique as shown in figure 3. A combination of GC and HPLC could determine all of the compounds on a quantitative basis, but this is more tedious, and requires considerably more mobile phase (ca 10 fold).
Figure 2: HPLC of Standard Mixture. Stationary Phase C18 column (150 x 3.1 mm), 3.5 µm particle. Mobile phase: water:acetonitrile:methanol gradient from 75:10:15 to 4:56:40 in 80 min.
Figure 1: Videodensitogram obtained from OPLC of Standard Mixture. Sample: 1) Nandrolone 2) 5α-4,5-Dihydronandrolone 3) Nandrolone∆)5(10)-isomer 4) 8,14-Didehydronandrolone 5) Nandrolinedieneolether 6) 6β-Hydroxynandrolone 7) Estradiol methylether. X axis is % Full length of plate
Figure 3: Videodensitogram obtained from TLC of Standard mixture. Stationary Phase: Silica Gel, Mobile phase: hexane:ethyl acetate (1:2)
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In a process lab, the assay can be performed rapidly, at low cost and with reasonable precision. As shown in table 1 (reverse side) and figure 2, other chromatographic techniques were able to determine some but not all of the compounds of interest, (peak 2 cannot be observed in HPLC).
HPLC Quantitative
GC Quantitative
TLC Semiquantitative
OPLC Semiquantitative
5α-4,5 Dihydronandrolone
-*
+
-**
+
Nandrolone ∆5(10)isomer
-*
+
+
-***
8,14 Didehydronandrolone
-*
+
+
-***
Dienolether
+
+
+
-***
6β-Hydroxynandrolone
+
+
+
-***
Estradiol methylether
+
+
+
+
Separation
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Impurity
Table1
+ = suitable, / - = unsuitable / * = cannot be separated, ** = cannot be detected
/
*** = degradation occurred
References 1. B. Bagocsi, D. Fabian, A. Lauko, M.Mezei, A. Maho, Z. Vegh and K. Ferenzi-Fodor,, Comparison of OPLC and other Chromatographic Methods (TLC, HPLC, and GC) for In-Process Purity Testing of Nandrolone, J. Planar Chromatography.15 (2002) 252-257.
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