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A note on the new U. S. P. XIV method for assay of progesterone injection
A Note on the New U. S. P. XIV Method for Assay of Progesterone Injection* By ERNEST J. UMBERGER
I
method described in U. S. P. XIV for the assay of progesterone injection, a sample equivalent t o about 20 mg. of progesterone in a vegetable oil solution is added to a separator containing 40 cc. of petroleum benzin. The progesterone is extracted from this mixture with five successive 20-cc. portions of 90% alcohol. Because of the mutual solubility of petroleum benzin and 90% alcohol, only about 5 cc. of the petroleum benzin layer remains after the last extraction. It has been found that by the simple expedient of equilibrating the solvents before use, about 35 cc. of the petroleum benzin layer remains after the five extractions with 90% alcohol. N THE
EXPERIMENTAL
*
TABLE I.-RESULTS OF ANALYSES OF ALconoL AND SESAME OIL SOLUTIONS OF PROGESTERONE USING EQUILIBRATED AND NONEQUILIBRATED SOLVENTS IN THE U. S. P. XIV METHODFOR PROGESTERONE INJECTION
Sample No.
The solvents were equilibrated by shaking equal volumes of petroleum benzin and 90% alcohol in a separatory funnel and separating. In the first experiment, 1cc. of sesame oil was extracted using equilibrated solvents, and a similar amount extracted using nonequilibrated solvents. The petroleum benzin layers were then transferred to weighed 50-cc. beakers and evaporated to dryness on the steam bath under a stream of air. After weighing, it was found that 89% of the oil was retained in the petroleum benzin layer when equilibrated solvents were used, but only 73% when nonequilibrated solvents were used. The second experiment was designed t o show whether or not there was any loss in efficiency by the use of equilibrated solvents in the extraction of progesterone. b n alcohol solution was prepared containing approximately 20 mg. of progesterone per cubic centimeter. Duplicate I-cc. samples of this solution were extracted by each of the two methods. As shown in Table I, sample nos. 1 4 , identical results were obtained. Finally, both methods were used for the assay of 0.8 cc. of a sesame oil solution containing 25 mg. of progesterone per cubic centimeter. These results Received November 4, 1950, from the Division of Pharmacology, Food and Drug Administration, Federal Security Agency, Washington, D. C.
are shown as sample nos. 5-8 in Table I. There appears to be a tendency toward high values when nonequilibrated solvents are used. This may be due to the incomplete removal of the oil residue from the 2.4-dinitrophenyl hydrazine derivatives, which is reflected in the melting points, also shown in Table I.
Method
Mg.
Found
M.p. of 2.4Dinitrophenyl Hydrazine
Alcohol Solution, 20 Mg./Cc. Used 1.0 Cc. Nonequilibrated 19.8 277-279' 2 Nonequilibrated 19.8 276-279 ' 3 Equilibrated 19.8 276-278' 4 Equilibrated 19.7 277-279'
1
5 6
7 8
Oil Solution, 25 Mg./Cc. Used Nonequilibrated 20.2 21.3 Nonequilibrated 19.8 Equilibrated 20.0 Equilibrated
0.8 Cc. 275-278' 273-277' 276-278' 276-278'
DISCUSSION
It is apparent that the use of equilibrated solvents instead of nonequilibrated solvents in the U. S. P. XIV assay method for progesterone injection, results in a better separation of progesterone from its oily vehicle, and consequently in a purer 2,4-dinitrophenyl hydrazine derivative. Danger of mechanical loss is minimized since about 35 cc. of the petroleum benzin layer remains after the alcohol extractions using equilibrated solvents as compared to about 5 cc. using nonequilibrated solvents. This may be particularly important on a warm day when there would be greater loss of solvent due t o evaporation. There is no decrease in the efficiency of the extraction of progesterone by the use of equilibrated solvents.
GERMAN JOURNALS MERGED It has been announced that the Sueddeutsche Apotheker-Zeitung has merged with the Deutsche Apotheker Zeitung, and will be published under the name Deutscher Apotheker k'erkzg. The editor of the new publication will he nr. Roland Schiniedel, former editor of the SuedZeutsche Apathekq-Zeiilztng. T h e address of the editorial riflicc is 'I'uebinger Str. 53, Stiittgart S,(;mnany. 'i()O
I
method described in U. S. P. XIV for the assay of progesterone injection, a sample equivalent t o about 20 mg. of progesterone in a vegetable oil solution is added to a separator containing 40 cc. of petroleum benzin. The progesterone is extracted from this mixture with five successive 20-cc. portions of 90% alcohol. Because of the mutual solubility of petroleum benzin and 90% alcohol, only about 5 cc. of the petroleum benzin layer remains after the last extraction. It has been found that by the simple expedient of equilibrating the solvents before use, about 35 cc. of the petroleum benzin layer remains after the five extractions with 90% alcohol. N THE
EXPERIMENTAL
*
TABLE I.-RESULTS OF ANALYSES OF ALconoL AND SESAME OIL SOLUTIONS OF PROGESTERONE USING EQUILIBRATED AND NONEQUILIBRATED SOLVENTS IN THE U. S. P. XIV METHODFOR PROGESTERONE INJECTION
Sample No.
The solvents were equilibrated by shaking equal volumes of petroleum benzin and 90% alcohol in a separatory funnel and separating. In the first experiment, 1cc. of sesame oil was extracted using equilibrated solvents, and a similar amount extracted using nonequilibrated solvents. The petroleum benzin layers were then transferred to weighed 50-cc. beakers and evaporated to dryness on the steam bath under a stream of air. After weighing, it was found that 89% of the oil was retained in the petroleum benzin layer when equilibrated solvents were used, but only 73% when nonequilibrated solvents were used. The second experiment was designed t o show whether or not there was any loss in efficiency by the use of equilibrated solvents in the extraction of progesterone. b n alcohol solution was prepared containing approximately 20 mg. of progesterone per cubic centimeter. Duplicate I-cc. samples of this solution were extracted by each of the two methods. As shown in Table I, sample nos. 1 4 , identical results were obtained. Finally, both methods were used for the assay of 0.8 cc. of a sesame oil solution containing 25 mg. of progesterone per cubic centimeter. These results Received November 4, 1950, from the Division of Pharmacology, Food and Drug Administration, Federal Security Agency, Washington, D. C.
are shown as sample nos. 5-8 in Table I. There appears to be a tendency toward high values when nonequilibrated solvents are used. This may be due to the incomplete removal of the oil residue from the 2.4-dinitrophenyl hydrazine derivatives, which is reflected in the melting points, also shown in Table I.
Method
Mg.
Found
M.p. of 2.4Dinitrophenyl Hydrazine
Alcohol Solution, 20 Mg./Cc. Used 1.0 Cc. Nonequilibrated 19.8 277-279' 2 Nonequilibrated 19.8 276-279 ' 3 Equilibrated 19.8 276-278' 4 Equilibrated 19.7 277-279'
1
5 6
7 8
Oil Solution, 25 Mg./Cc. Used Nonequilibrated 20.2 21.3 Nonequilibrated 19.8 Equilibrated 20.0 Equilibrated
0.8 Cc. 275-278' 273-277' 276-278' 276-278'
DISCUSSION
It is apparent that the use of equilibrated solvents instead of nonequilibrated solvents in the U. S. P. XIV assay method for progesterone injection, results in a better separation of progesterone from its oily vehicle, and consequently in a purer 2,4-dinitrophenyl hydrazine derivative. Danger of mechanical loss is minimized since about 35 cc. of the petroleum benzin layer remains after the alcohol extractions using equilibrated solvents as compared to about 5 cc. using nonequilibrated solvents. This may be particularly important on a warm day when there would be greater loss of solvent due t o evaporation. There is no decrease in the efficiency of the extraction of progesterone by the use of equilibrated solvents.
GERMAN JOURNALS MERGED It has been announced that the Sueddeutsche Apotheker-Zeitung has merged with the Deutsche Apotheker Zeitung, and will be published under the name Deutscher Apotheker k'erkzg. The editor of the new publication will he nr. Roland Schiniedel, former editor of the SuedZeutsche Apathekq-Zeiilztng. T h e address of the editorial riflicc is 'I'uebinger Str. 53, Stiittgart S,(;mnany. 'i()O