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Starvation and anabolic steroids
Starvation and Anabolic Steroids ByVLADIMIRKFUZEK, PAVELSTJZPANEK AND LUWIK SAIXUX The effect of the administration of methandrostenolone (Danabol, Ciba) on the course of three-day complete starvation was investigated. The preparation was given to nine male volunteers, while a control group of the same size was subjected to starvation without the steroid. A negative nitrogen balance was found in both groups, but in the group which received 20 mg. methandrostenolone daily, nitrogen excretion was 30 per cent lower. Differences in body weight changes were also approximately the same. In the
group given the anabolic steroid during starvation, a significantly greater increase in the blood cholesterol, lipid and uric acid level was observed and ketonuria was also more pronounced. Smaller doses of
the steroid (5 mg. Danabol daily) had similar, but quantitatively somewhat lower, effects. Study of the nitrogen balance during complete starvation can be used to test and compare the efficacy of the protein-conserving action of different anabolic preparations. (Metabolism 18: No. 7, July, 585-592, 1969)
POMPLETE STARVATION leads to a large negative protein balance. _ _ b The aim of our study was to determine whether, and to what extent, anabolic steroids would influence protein catabolism and hence the nitrogen balance of starving subjects. The Clinical Department of the Institute of Physical Medicine and Balneology in Marianske L&n& provided us with an opportunity of carrying out a rigorously controlled trial. Eighteen male volunteers, most of whom were undergoing weight reduction, were subjected to three days’ complete starvation. The majority were obese, but none of them suffered from a metabolic disease or any other disorder which might have affected the results of the test. They had likewise not been receiving drugs. The test subjects were divided into two groups, one of which was subjected to three days’ starvation without drugs and the other, under otherwise exactly the same conditions, was given Danabole (Ciba; methandrostenolone, 17-aplha-methyl-17-beta-hydroxyandrostane-1,4-dione-3-one.) The test subjects were arranged in nine pairs of approximately the same weight and degree of obesity and one of each pair was included by random choice in each of the groups, as previous experience had shown that the response to a reducing diet and to starvation was markedly correlated to the From the Research Institute of Physical Medicine and Balneology, MaridnskS L&G, Marienbad, Czechoslovakia. Received for publication April 1, 1969. VLADIMIR KRIZEK, M.D., CSc.: Head Clinical Department, Research Institute of Physical Medicine and Balneology, Marianske Lazne, Marienbad, Czechoslovakia. PAVEL STEPANEK, M.D.: Research Member, Research Institute of Physical Medicine and Balneology, Mariarwke Lazne, Marienbad, Czechosluvakia. LULWIKSADILEX, PH.D.: Research Chemist, Research Institute of Physical Medicine and Balneology, Marianske Lazne, Marienbad, Czechoslovakia. METABOLISM, VOL. 18, No. 7 (JULY), 1969
585
586
KRIZEK, Table
l.--Clinical
STEPANEK
AND SADILEK
Data _
Dayabo’
41.3; 104.9; 171.3;
Age
Weight Height Overweight “Broca”
+47.1;
goup
Cojtro~Ggr;,up
23:; yrs. 79.8-142.0 Kg. 165-180 Cm. +1o&S+93.5
%
45.6; 105.3; 175.3; $39.2;
2&O yrs. 80.0-145.6 Kg. 169184 Cm. +11.0-+75.0
%
weight and degree of obesity of the subjects observed. The main findings in all the test subjects are given in Table 1. MATERIALS
AND METHODS
Three days’ complete starvation was observed. The interval between the last meal before starting the test on the previous evening and the first meal in the morning after terminating it was longer, of course, and amounted to 85 hours. Fluid intake was not restricted, but a record was kept of the amount. Weak tea sweetened with saccharin or plain water was allowed. The subjects collected all their urine in 24-hour portions, starting 24 hours before commencing starvation on the day after completing the test, i.e., a total of five days. Blood for laboratory tests was taken twice: on the morning of the first day of starvation and three days later, before the first meal was eaten. Every morning, after urinating, the subjects were weighed on the same scales to within + 50 Gm. To complete the picture it should be added that before and after the starvation the test subjects underwent reducing regimen with a daily diet of 1700 Cal. containing 95-100 Gm. protein.
Analytical Methods Nitrogen was determined in the urine by the micro-Kjeldahl method,” creatinine in urine and serum by the method of Bonsnes and Taussky,r uric acid in urine and serum by King’s method,4 total serum cholesterol by the Searcy-Berquist method,12 and total serum lipids by the method of De La Huerga.2 Creatinine and uric acid clearance was determined by the formula U.V/P, in which U is the urine concentration of the substance, V the urine flow per minute and P the serum concentration of the substance. Since blood was collected only twice, at the onset and end of the experiment, the P values for the individual days are extrapolated. The difference between the mean clearance values for three days’ starvation and the values on the day immediately preceding starvation was evaluated. Ketonuria was evaluated by semiquantitative assay of the urine, using the nitroprusside test with five-point scale. This test primarily characterizes aceto-acetic acid excretion and is less specific for acetone; it is not used for the evaluation of hydroxybutyric acid.
Statistical Evaluation The arithmetical means of the experimental data were determined, together with the standard deviations (S.D.). Differences between the two groups of test subjects were evaluated by the Student’s t test.
Toleration of Starvation Both groups tolerated starvation relatively well. Most subjects complained of hunger chiefly on the first day. We did not observe greater hunger, or subjectively worse or better toleration of starvation in those who received the anabolic steroid.
Anabolic
Steroid
Half (nine) of the test subjects received Danabol (Ciba) in amounts of 20 mg. on each of the three days of the starvation test and in amounts of 10 mg./day on the two days
STARVATION
AND ANABOLIC
587
STEROIDS
Table 2.-Effect
of 3-day Starvation
Co,tro,,Tp n-
Weight loss (Kg.) after S-days starvation Weight loss ( % ) of initial weight Total urinary nitrogen loss (Gm./3 days) Creatinine excretion (Cm./3 days) Uric acid excretion (Gm./3 days) Urine flow/3 days Fluid intake/3 days Changes in: Serum uric acid mg.% Total cholesterol mg.% Total lipids mg.% Creatinine clearance ml./min. Uric acid clearance ml./min.
Danabol Group (27ng;cY)
S.D.
n
S.D.
Difference
t
Significance
4.09
1.07
2.99
0.84
-1.10
2.36
P
3.98
0.94
2.82
0.33
-1.16
3.56
P
33.21
7.74
23.03
5.00
-10.18
3.33
P
5.92
0.99
5.29
0.88
-0.63
1.56
1.55
0.46
4.12 4.02
1.90 1.73
1.09 1.99 4.22
0.39 0.86 1.15
-0.46 -2.13 +0.20
2.30 3.08 0.28
P
0.64 18.62 78.16 9.01 3.79
+0.58 +28.66 +SS.O -2.08 -0.76
1.74 3.34 2.53 0.25 0.52
P=O.O5 P
+1.78 +4.67 +31.11 -4.65 -2.91
0.79 17.8 65.54 22.9 2.13
t-2.36 +33.33 t117.11 -6.73 -3.67
preceding the test. The total dose for five days was thus 80 mg. In a short-term test with relatively large dose we observed no side effects from this preparation. In another test, described below, we tested the effect of daily doses of 5 mg. Danabol, which we administered to five male subjects for five days, two days before starvation and during three days starvation, making a total dose of 25 mg. for five days. RESULTS
In Table 2 we compare metabolic processes in aggregate for a three-day period of either complete starvation or complete starvation with administration of the anabolic steroid. In over 100 subjects of different constitutions we verified whether such comparisons are possible, especially on choosing subjects with approximately the same body weight, the same degree of overweight and the same amount of lean body mass. In the actual experimental subjects, we also determined the value on one or two days preceding starvation. Those in the group given the steroid received a daily dose of 10 mg. for two days prior to starvation, however. We therefore intentionally did not compare the base-line values, but only the values for three days of actual starvation. Statistically significant differences: In the group given the steroid, the absolute body weight decrement was 27 per cent smaller, the relative decrement, in relation to the initial weight, was 29 per cent smaller than in the control group and nitrogen losses were 30 per cent smaller. The urine flow, with approximately the same fluid intake, was halved. In this group there was also a significantly greater increase in the blood uric acid, cholesterol and lipid levels. Differences in the amount of creatinine excreted in the urine were not statistically significant. A drop, however, was recorded, and the same applies to endogenous creatinine and uric acid clearance. There was a decrease
5%
KRIZEK,
STEPANEK
AND
SADILEK
in both groups. In the control group (simple starvation), pronounced ketonuria was not observed until the third day, and then in only one-third of the subjects, while in the group which received the anabolic steroid it was already demonstrated on the first day of starvation in eight subjects out of nine. On the third day of the test, all subjects given the steroid had severe ketosis, which in two thirds still persisted 24 hours after terminating the test. In the control group ketonuria persisted in only one subject out of nine. Specification
of Weight
Decrement
During
Starvation
From the values decisive in starvation, i.e., changes in body weight, the amount of given substances excreted in the urine and the estimated energy output, we attempted to determine the proportion of various body constituents in the weight decrement. Since the subjects were starved, we did not take excretion in the stools and losses through the skin into account. The calculation presumes a zero calorie intake, the same glycogen losses in all the subjects and a calorie output estimated from tabular values of basal metabolism for approximately the same physical activity for all the subjects. The weight changes measured and nitrogen losses in the urine make it possible, by using the calorie balance, to calculate the proportion of fat, lean body mass and water in the total weight decrement. Calorie balance in the interval studied equals calorie intake from diet plus calorie intake from catabolized tissues (lean body mass plus glycogen and fat) minus calorie output. In our study, calorie intake from the diet is taken zero. The calories from catabolized lean body mass (LBM) are expressed as the number of grams of nitrogen in the urine X 6.25 X 4.1.11 Glycogen losses in both groups were estimated at 250 Gm., i.e., 1000 Cal. per subject. Calorie output is taken from tabular basal metabolism values? plus 33 per cent to allow the limited and controllable amount of physical activity in a clinical metabolic ward. In the calorie balance equation, only catabolized fat energy is unknown. It was calculated as follows: the catabolized fat calories divided by the factor 9.3 (1 Gm. fat = 9.3 available Cal.) gives the following Cal. catab. fat 1.33 X basal. fat losses: fat losses (3 days’ starvation) =--~-~3~-~ = -_____ metab. X 3 -
Cal. catab. LBM
-
Cal. catab. glycogen.
LBM losses = grams 9.3 of urine nitrogen X 32.1° Water losses = total body water changes calculated from body weight and LBM changes, glycogen and fat: A HZ0 = A body weight - A LBM - A glycogen - A fat.8sQ Differences in the structure of weight decrements in the group subjected only to starvation and in the group given the anabolic steroid are illustrated in Fig. 1. The group without the steroid shows a bigger decrease in the water content as well as in the lean body mass. In the control group, the caloric equivalent for a loss of 1 Kg. body weight during three days’ starvation was 2080 Cal., while in the group given Danabol it was 2800 Cal. The caloric equivalent was expressed as the ratio of the calorie balance to the body weight change during the given period.
STARVATION
AND ANABOLIC
589
STEROIDS
*a 4-
0 AMABOL 3-
Fig. L-Composition of weight decrement during three-day starvation. (Left column) simple starvation without medicament; (Right column) starvation with 20 mg. Danabol/day.
2-
l-
L
Table 3.Effect
of Anabolic Steroid Dose on Metabolic Response During 3-day
Starvation
Witfp&e@ Weight loss (Kg.) after 3-day starvation Weight loss ( % ) of initial weight Total urinary nitrogen loss (Gm./3 days) Creatinine excretion (Gm./3 days) Uric acid excretion (Gm./3 days) Urine (L/3 days) Changes in: Serum uric acid (mg.% ) Total cholesterol ( mg. % ) Total lipids (mg.% )
lnfbmce
Dar&ml (5 m&/day; n = 5)
Danabol (20 mg./day; n = 9)
4.09 Kg.
3.23 Kg.
2.99 Kg.
3.98
3.00
2.82
33.21
24.55
23.04
5.92
5.11
5.29
1.55 4.12
1.07 2.31
1.09 1.99
+1.77 -+4.67 t31.11
of Anubolic Steroid Dose During
+z.e2 f23.6 +lll.S
+2.36 +33.33 +117.11
Starvation
The given dose (20 mg. Danabol daily) is relatively high. We investigated the influence of smaller dose by carrying out a similar three days’ starvation test with five male subjects. The experimental conditions and urine and blood tests were the same as above. On the two days preceding starvation and during the three days of the test, these subjects were given daily doses of 5 mg. Danabol, a total dose of 25 mg. The results are given in Table 3. These findings are also compared with the results in the group subjected only to starvation and those in the group which received 20 mg. Danabol daily. Since the supplementary test was carried out with only five subjects, the differences in the values were too small to reach the level of statistical significance and the results of the statistical evaluation are therefore omitted. For most of the criteria used, however, a trend can be seen from the group given 20 mg. Danabol, via the group which received the smaller dose of the preparation, to the control group subjected only to starvation.
590
KRIZEK,
STEPANEK
AND
SADILEK
DISCUSSION
Our test conclusively shows that the administration of methandrostenolone during several days’ starvation influenced the protein balance. Protein intake was completely stopped, but its catabolism continued and the nitrogen balance was consequently negative. The anabolic steroid reduced protein catabolism and the negativity of the nitrogen balance, however. This occurred not only after the administration of relatively large doses (20 mg. methandrostenolone daily) but also after relatively small doses (5 mg. daily). When comparing the effects of anabolic steroids, Metcalf used the term “excretion ratio,” which expresses the ratio of nitrogen excretion during a given period in a group given such a drug to its excretion in a group not given the drug.5 He also used the term “retention ratio,” which is 1 minus the excretion ratio. For optimal activity of anabolic steroids he gave the retention ratio 23 A 4 per cent, which he found to be independent of age, sex, weight, surface area, basal nitrogen excretion and general metabolic status. In our tests, the retention ratio for a daily dose of 20 mg. Danabol was 30.5 per cent and for a daily dose of 5 mg. 26.2 per cent (Table 3). Methandrostenolone has a very good protein-conserving effect and its influence on protein metabolism is also manifested if the organism’s protein supply is reduced or stopped. In practice, it means that this preparation can be used in situations similar to the one in the present test for mitigating body protein losses, e.g., in certain post-operative states, sudden abdominal episodes necessitating interruption of the food intake, emergency situations and natural catastrophes. It also raises the question of the use of these substances during starvation as part of a radical weight-reducing course.13.*-3Relatively small doses of Danabol also have a significant anabolic effect on protein metabolism and by using such doses we avoid any possible side effects, even if we administer the preparation over a longer period than in the present test. The finding that anabolic steroids have a protein-conserving effect under starvation conditions could also be of significance for testing various preparations of this type with the same effect. In a healthy subject it is always only a question of time before the nitrogen balance becomes stabilized at a level higher than the physiological minimum. The only optimal observations, therefore, are those based on a minimum nitrogen balance. It is obvious that, under clinical conditions, it is not easy to obtain this ideal basis for evaluation of the anabolic effect.6 Under these circumstances, therefore, we find it satisfactory to test and compare anabolic steroids in subjects undergoing short-term starvation. When setting up the control groups it should be borne in mind that, given otherwise the same condition, negativity of the nitrogen balance is usually greater in males than in females. The following points in our findings also merit attention. Interpretation of the somewhat obscure changes in water balance is impeded by the fact that we did not simultaneously study the excretion of certain basic electrolytes. With the same spontaneous fluid intake, the urine flow in starved subjects given the anabolic steroid was about half the volume in the group to which the steroid
STARVATION
AND ANABOLIC
591
STEROIDS
Danabol
Fig. B.--Mean weight curve before, during and after starvation in male subjects given no drug (Left) and given Danabol (Right). (Below) mean daily .:b
::/7t
weight
changes.
was not administered. With reference to our estimate of the change in the composition of the body weight decrement during starvation, it is clear that a negative water balance developed in both groups, but that in the group given the steroid negativity was about 790 Gm. lower. The difference in the amount of lean body mass catabolized thus corresponded to only 250 Gm. water. During the first days of starvation, a markedly negative water and electrolyte balance develops. Danabol seems to promote water retention, manifested in a lower urine ilow and a less negative water balance. It is worth mentioning, however, that after completing three days’ starvation and resuming a daily diet of 1700 Cal. containing 100 Gm. protein and a normal amount of salt, a weight increment was observed in both groups. In the group not given the steroid, this increment lasted four days and in the other group, which received Danabol, it lasted only three days, but was significantly higher. This again raises the question of the extent to which the waning anabolic effect of the steroid on protein, or its possible effect on electrolytes, participates in this increment. Figure 2 illustrates the mean weight curves for both groups of test subjects, including their course for a whole week after discontinuing starvation. The mean daily weight changes are given in the lower part of the figure. We also drew attention to a number of other changes produced in lipid balance by the anabolic steroid: a significant increase in the serum cholesterol and total serum lipid level and pronounced ketonuria. These phenomena could have been the outcome of the greater catabolism of adipose tissue which occurs during starvation with the administration of anabolic steroids. Apart from a few exceptions, however, very little attention was paid to these questions in the part.3 REFERENCES 1. Bonsnes, R. W., and Taussky, H. H.: On the calorimetric determination of creatinine by the Jaffe reaction. J. Biol. Chem. 158:581, 1945. 2. De La Huerga, J., Yesinick, C., and Estimation of total serum Popper, H.:
lipids by a turbidimetric method. Amer. J. Clin. Pathol. 23:1163, 1953. 3. Fiegel, G., Bargheer, R., and Kulwa, D.: Beeinflussung des Lipidhaushaltes durch anabole Substanzen. Arzneimittel-Forsch. 16:202, 1966.
592 4. King, E. J.: Micro-analysis in Medical Biochemistry. London, Churchill, 1954. 5. Metcalf, W., and Greene, H. G.: A quantitative expression for nitrogen retention with anabolic steroids. Metabolism 12: 899, 1963. 6. Noelle H.: Stickstoffbilanzuntersuchungen zur Uberpriifung anaboler Steroide. Zs.ges.exp.Medizin 145: 138, 1968. Bes:~moumgr;~~ SanTiiz Grundumsatz8. Passmore, R., Strong, J. A., and Ritchie, F. J.: The chemical composition of the tissue lost by obese patients on a reducing regimen. Brit. J. Nutr. 12: 113, 1958. 9. -, -, and -: Water and electrolyte exchanges of obese patients on a reducing regimen. Brit. J. Nutr. 13:17, 1959. 10. Reifenstein, E. C., Jr., Albright, F., and Wells, S. L.: The accumulation, inter-
KRIZEK, STEPANEK
AND SADILEK
pretation, and presentation of data pertaining to metabolic balances, notably those of calcium, phosphorus and nitrogen. J. Clin. Endocr. 5:367, 1945. 11. Russell, G. F. M.: The effect of diets of different composition on weight loss, water and sodium balance in obese patients. Clin. Sci. 22:269, 1962. 12. Searcy, R. L., and Berquist, L. M.: A new color reaction for the quantitation of serum cholesterol. Clin. Chim. Acta 5:192, 1960. 13. Slabochovi, Z., Rath, R., and Placer, Z.: Effect of methandrostenolon and dimethandrostenolo in the dietotherapy of obesity. Nutr. Dieta 7:271, 1965. 14. Stepanek, P., Kfiiek, V., and Sadilek, L.: Die Anwendung von Trijodthyronin und Dianabol bei Abmagerungskuren. Med. Welt 17:2393, 1966.
group given the anabolic steroid during starvation, a significantly greater increase in the blood cholesterol, lipid and uric acid level was observed and ketonuria was also more pronounced. Smaller doses of
the steroid (5 mg. Danabol daily) had similar, but quantitatively somewhat lower, effects. Study of the nitrogen balance during complete starvation can be used to test and compare the efficacy of the protein-conserving action of different anabolic preparations. (Metabolism 18: No. 7, July, 585-592, 1969)
POMPLETE STARVATION leads to a large negative protein balance. _ _ b The aim of our study was to determine whether, and to what extent, anabolic steroids would influence protein catabolism and hence the nitrogen balance of starving subjects. The Clinical Department of the Institute of Physical Medicine and Balneology in Marianske L&n& provided us with an opportunity of carrying out a rigorously controlled trial. Eighteen male volunteers, most of whom were undergoing weight reduction, were subjected to three days’ complete starvation. The majority were obese, but none of them suffered from a metabolic disease or any other disorder which might have affected the results of the test. They had likewise not been receiving drugs. The test subjects were divided into two groups, one of which was subjected to three days’ starvation without drugs and the other, under otherwise exactly the same conditions, was given Danabole (Ciba; methandrostenolone, 17-aplha-methyl-17-beta-hydroxyandrostane-1,4-dione-3-one.) The test subjects were arranged in nine pairs of approximately the same weight and degree of obesity and one of each pair was included by random choice in each of the groups, as previous experience had shown that the response to a reducing diet and to starvation was markedly correlated to the From the Research Institute of Physical Medicine and Balneology, MaridnskS L&G, Marienbad, Czechoslovakia. Received for publication April 1, 1969. VLADIMIR KRIZEK, M.D., CSc.: Head Clinical Department, Research Institute of Physical Medicine and Balneology, Marianske Lazne, Marienbad, Czechoslovakia. PAVEL STEPANEK, M.D.: Research Member, Research Institute of Physical Medicine and Balneology, Mariarwke Lazne, Marienbad, Czechosluvakia. LULWIKSADILEX, PH.D.: Research Chemist, Research Institute of Physical Medicine and Balneology, Marianske Lazne, Marienbad, Czechoslovakia. METABOLISM, VOL. 18, No. 7 (JULY), 1969
585
586
KRIZEK, Table
l.--Clinical
STEPANEK
AND SADILEK
Data _
Dayabo’
41.3; 104.9; 171.3;
Age
Weight Height Overweight “Broca”
+47.1;
goup
Cojtro~Ggr;,up
23:; yrs. 79.8-142.0 Kg. 165-180 Cm. +1o&S+93.5
%
45.6; 105.3; 175.3; $39.2;
2&O yrs. 80.0-145.6 Kg. 169184 Cm. +11.0-+75.0
%
weight and degree of obesity of the subjects observed. The main findings in all the test subjects are given in Table 1. MATERIALS
AND METHODS
Three days’ complete starvation was observed. The interval between the last meal before starting the test on the previous evening and the first meal in the morning after terminating it was longer, of course, and amounted to 85 hours. Fluid intake was not restricted, but a record was kept of the amount. Weak tea sweetened with saccharin or plain water was allowed. The subjects collected all their urine in 24-hour portions, starting 24 hours before commencing starvation on the day after completing the test, i.e., a total of five days. Blood for laboratory tests was taken twice: on the morning of the first day of starvation and three days later, before the first meal was eaten. Every morning, after urinating, the subjects were weighed on the same scales to within + 50 Gm. To complete the picture it should be added that before and after the starvation the test subjects underwent reducing regimen with a daily diet of 1700 Cal. containing 95-100 Gm. protein.
Analytical Methods Nitrogen was determined in the urine by the micro-Kjeldahl method,” creatinine in urine and serum by the method of Bonsnes and Taussky,r uric acid in urine and serum by King’s method,4 total serum cholesterol by the Searcy-Berquist method,12 and total serum lipids by the method of De La Huerga.2 Creatinine and uric acid clearance was determined by the formula U.V/P, in which U is the urine concentration of the substance, V the urine flow per minute and P the serum concentration of the substance. Since blood was collected only twice, at the onset and end of the experiment, the P values for the individual days are extrapolated. The difference between the mean clearance values for three days’ starvation and the values on the day immediately preceding starvation was evaluated. Ketonuria was evaluated by semiquantitative assay of the urine, using the nitroprusside test with five-point scale. This test primarily characterizes aceto-acetic acid excretion and is less specific for acetone; it is not used for the evaluation of hydroxybutyric acid.
Statistical Evaluation The arithmetical means of the experimental data were determined, together with the standard deviations (S.D.). Differences between the two groups of test subjects were evaluated by the Student’s t test.
Toleration of Starvation Both groups tolerated starvation relatively well. Most subjects complained of hunger chiefly on the first day. We did not observe greater hunger, or subjectively worse or better toleration of starvation in those who received the anabolic steroid.
Anabolic
Steroid
Half (nine) of the test subjects received Danabol (Ciba) in amounts of 20 mg. on each of the three days of the starvation test and in amounts of 10 mg./day on the two days
STARVATION
AND ANABOLIC
587
STEROIDS
Table 2.-Effect
of 3-day Starvation
Co,tro,,Tp n-
Weight loss (Kg.) after S-days starvation Weight loss ( % ) of initial weight Total urinary nitrogen loss (Gm./3 days) Creatinine excretion (Cm./3 days) Uric acid excretion (Gm./3 days) Urine flow/3 days Fluid intake/3 days Changes in: Serum uric acid mg.% Total cholesterol mg.% Total lipids mg.% Creatinine clearance ml./min. Uric acid clearance ml./min.
Danabol Group (27ng;cY)
S.D.
n
S.D.
Difference
t
Significance
4.09
1.07
2.99
0.84
-1.10
2.36
P
3.98
0.94
2.82
0.33
-1.16
3.56
P
33.21
7.74
23.03
5.00
-10.18
3.33
P
5.92
0.99
5.29
0.88
-0.63
1.56
1.55
0.46
4.12 4.02
1.90 1.73
1.09 1.99 4.22
0.39 0.86 1.15
-0.46 -2.13 +0.20
2.30 3.08 0.28
P
0.64 18.62 78.16 9.01 3.79
+0.58 +28.66 +SS.O -2.08 -0.76
1.74 3.34 2.53 0.25 0.52
P=O.O5 P
+1.78 +4.67 +31.11 -4.65 -2.91
0.79 17.8 65.54 22.9 2.13
t-2.36 +33.33 t117.11 -6.73 -3.67
preceding the test. The total dose for five days was thus 80 mg. In a short-term test with relatively large dose we observed no side effects from this preparation. In another test, described below, we tested the effect of daily doses of 5 mg. Danabol, which we administered to five male subjects for five days, two days before starvation and during three days starvation, making a total dose of 25 mg. for five days. RESULTS
In Table 2 we compare metabolic processes in aggregate for a three-day period of either complete starvation or complete starvation with administration of the anabolic steroid. In over 100 subjects of different constitutions we verified whether such comparisons are possible, especially on choosing subjects with approximately the same body weight, the same degree of overweight and the same amount of lean body mass. In the actual experimental subjects, we also determined the value on one or two days preceding starvation. Those in the group given the steroid received a daily dose of 10 mg. for two days prior to starvation, however. We therefore intentionally did not compare the base-line values, but only the values for three days of actual starvation. Statistically significant differences: In the group given the steroid, the absolute body weight decrement was 27 per cent smaller, the relative decrement, in relation to the initial weight, was 29 per cent smaller than in the control group and nitrogen losses were 30 per cent smaller. The urine flow, with approximately the same fluid intake, was halved. In this group there was also a significantly greater increase in the blood uric acid, cholesterol and lipid levels. Differences in the amount of creatinine excreted in the urine were not statistically significant. A drop, however, was recorded, and the same applies to endogenous creatinine and uric acid clearance. There was a decrease
5%
KRIZEK,
STEPANEK
AND
SADILEK
in both groups. In the control group (simple starvation), pronounced ketonuria was not observed until the third day, and then in only one-third of the subjects, while in the group which received the anabolic steroid it was already demonstrated on the first day of starvation in eight subjects out of nine. On the third day of the test, all subjects given the steroid had severe ketosis, which in two thirds still persisted 24 hours after terminating the test. In the control group ketonuria persisted in only one subject out of nine. Specification
of Weight
Decrement
During
Starvation
From the values decisive in starvation, i.e., changes in body weight, the amount of given substances excreted in the urine and the estimated energy output, we attempted to determine the proportion of various body constituents in the weight decrement. Since the subjects were starved, we did not take excretion in the stools and losses through the skin into account. The calculation presumes a zero calorie intake, the same glycogen losses in all the subjects and a calorie output estimated from tabular values of basal metabolism for approximately the same physical activity for all the subjects. The weight changes measured and nitrogen losses in the urine make it possible, by using the calorie balance, to calculate the proportion of fat, lean body mass and water in the total weight decrement. Calorie balance in the interval studied equals calorie intake from diet plus calorie intake from catabolized tissues (lean body mass plus glycogen and fat) minus calorie output. In our study, calorie intake from the diet is taken zero. The calories from catabolized lean body mass (LBM) are expressed as the number of grams of nitrogen in the urine X 6.25 X 4.1.11 Glycogen losses in both groups were estimated at 250 Gm., i.e., 1000 Cal. per subject. Calorie output is taken from tabular basal metabolism values? plus 33 per cent to allow the limited and controllable amount of physical activity in a clinical metabolic ward. In the calorie balance equation, only catabolized fat energy is unknown. It was calculated as follows: the catabolized fat calories divided by the factor 9.3 (1 Gm. fat = 9.3 available Cal.) gives the following Cal. catab. fat 1.33 X basal. fat losses: fat losses (3 days’ starvation) =--~-~3~-~ = -_____ metab. X 3 -
Cal. catab. LBM
-
Cal. catab. glycogen.
LBM losses = grams 9.3 of urine nitrogen X 32.1° Water losses = total body water changes calculated from body weight and LBM changes, glycogen and fat: A HZ0 = A body weight - A LBM - A glycogen - A fat.8sQ Differences in the structure of weight decrements in the group subjected only to starvation and in the group given the anabolic steroid are illustrated in Fig. 1. The group without the steroid shows a bigger decrease in the water content as well as in the lean body mass. In the control group, the caloric equivalent for a loss of 1 Kg. body weight during three days’ starvation was 2080 Cal., while in the group given Danabol it was 2800 Cal. The caloric equivalent was expressed as the ratio of the calorie balance to the body weight change during the given period.
STARVATION
AND ANABOLIC
589
STEROIDS
*a 4-
0 AMABOL 3-
Fig. L-Composition of weight decrement during three-day starvation. (Left column) simple starvation without medicament; (Right column) starvation with 20 mg. Danabol/day.
2-
l-
L
Table 3.Effect
of Anabolic Steroid Dose on Metabolic Response During 3-day
Starvation
Witfp&e@ Weight loss (Kg.) after 3-day starvation Weight loss ( % ) of initial weight Total urinary nitrogen loss (Gm./3 days) Creatinine excretion (Gm./3 days) Uric acid excretion (Gm./3 days) Urine (L/3 days) Changes in: Serum uric acid (mg.% ) Total cholesterol ( mg. % ) Total lipids (mg.% )
lnfbmce
Dar&ml (5 m&/day; n = 5)
Danabol (20 mg./day; n = 9)
4.09 Kg.
3.23 Kg.
2.99 Kg.
3.98
3.00
2.82
33.21
24.55
23.04
5.92
5.11
5.29
1.55 4.12
1.07 2.31
1.09 1.99
+1.77 -+4.67 t31.11
of Anubolic Steroid Dose During
+z.e2 f23.6 +lll.S
+2.36 +33.33 +117.11
Starvation
The given dose (20 mg. Danabol daily) is relatively high. We investigated the influence of smaller dose by carrying out a similar three days’ starvation test with five male subjects. The experimental conditions and urine and blood tests were the same as above. On the two days preceding starvation and during the three days of the test, these subjects were given daily doses of 5 mg. Danabol, a total dose of 25 mg. The results are given in Table 3. These findings are also compared with the results in the group subjected only to starvation and those in the group which received 20 mg. Danabol daily. Since the supplementary test was carried out with only five subjects, the differences in the values were too small to reach the level of statistical significance and the results of the statistical evaluation are therefore omitted. For most of the criteria used, however, a trend can be seen from the group given 20 mg. Danabol, via the group which received the smaller dose of the preparation, to the control group subjected only to starvation.
590
KRIZEK,
STEPANEK
AND
SADILEK
DISCUSSION
Our test conclusively shows that the administration of methandrostenolone during several days’ starvation influenced the protein balance. Protein intake was completely stopped, but its catabolism continued and the nitrogen balance was consequently negative. The anabolic steroid reduced protein catabolism and the negativity of the nitrogen balance, however. This occurred not only after the administration of relatively large doses (20 mg. methandrostenolone daily) but also after relatively small doses (5 mg. daily). When comparing the effects of anabolic steroids, Metcalf used the term “excretion ratio,” which expresses the ratio of nitrogen excretion during a given period in a group given such a drug to its excretion in a group not given the drug.5 He also used the term “retention ratio,” which is 1 minus the excretion ratio. For optimal activity of anabolic steroids he gave the retention ratio 23 A 4 per cent, which he found to be independent of age, sex, weight, surface area, basal nitrogen excretion and general metabolic status. In our tests, the retention ratio for a daily dose of 20 mg. Danabol was 30.5 per cent and for a daily dose of 5 mg. 26.2 per cent (Table 3). Methandrostenolone has a very good protein-conserving effect and its influence on protein metabolism is also manifested if the organism’s protein supply is reduced or stopped. In practice, it means that this preparation can be used in situations similar to the one in the present test for mitigating body protein losses, e.g., in certain post-operative states, sudden abdominal episodes necessitating interruption of the food intake, emergency situations and natural catastrophes. It also raises the question of the use of these substances during starvation as part of a radical weight-reducing course.13.*-3Relatively small doses of Danabol also have a significant anabolic effect on protein metabolism and by using such doses we avoid any possible side effects, even if we administer the preparation over a longer period than in the present test. The finding that anabolic steroids have a protein-conserving effect under starvation conditions could also be of significance for testing various preparations of this type with the same effect. In a healthy subject it is always only a question of time before the nitrogen balance becomes stabilized at a level higher than the physiological minimum. The only optimal observations, therefore, are those based on a minimum nitrogen balance. It is obvious that, under clinical conditions, it is not easy to obtain this ideal basis for evaluation of the anabolic effect.6 Under these circumstances, therefore, we find it satisfactory to test and compare anabolic steroids in subjects undergoing short-term starvation. When setting up the control groups it should be borne in mind that, given otherwise the same condition, negativity of the nitrogen balance is usually greater in males than in females. The following points in our findings also merit attention. Interpretation of the somewhat obscure changes in water balance is impeded by the fact that we did not simultaneously study the excretion of certain basic electrolytes. With the same spontaneous fluid intake, the urine flow in starved subjects given the anabolic steroid was about half the volume in the group to which the steroid
STARVATION
AND ANABOLIC
591
STEROIDS
Danabol
Fig. B.--Mean weight curve before, during and after starvation in male subjects given no drug (Left) and given Danabol (Right). (Below) mean daily .:b
::/7t
weight
changes.
was not administered. With reference to our estimate of the change in the composition of the body weight decrement during starvation, it is clear that a negative water balance developed in both groups, but that in the group given the steroid negativity was about 790 Gm. lower. The difference in the amount of lean body mass catabolized thus corresponded to only 250 Gm. water. During the first days of starvation, a markedly negative water and electrolyte balance develops. Danabol seems to promote water retention, manifested in a lower urine ilow and a less negative water balance. It is worth mentioning, however, that after completing three days’ starvation and resuming a daily diet of 1700 Cal. containing 100 Gm. protein and a normal amount of salt, a weight increment was observed in both groups. In the group not given the steroid, this increment lasted four days and in the other group, which received Danabol, it lasted only three days, but was significantly higher. This again raises the question of the extent to which the waning anabolic effect of the steroid on protein, or its possible effect on electrolytes, participates in this increment. Figure 2 illustrates the mean weight curves for both groups of test subjects, including their course for a whole week after discontinuing starvation. The mean daily weight changes are given in the lower part of the figure. We also drew attention to a number of other changes produced in lipid balance by the anabolic steroid: a significant increase in the serum cholesterol and total serum lipid level and pronounced ketonuria. These phenomena could have been the outcome of the greater catabolism of adipose tissue which occurs during starvation with the administration of anabolic steroids. Apart from a few exceptions, however, very little attention was paid to these questions in the part.3 REFERENCES 1. Bonsnes, R. W., and Taussky, H. H.: On the calorimetric determination of creatinine by the Jaffe reaction. J. Biol. Chem. 158:581, 1945. 2. De La Huerga, J., Yesinick, C., and Estimation of total serum Popper, H.:
lipids by a turbidimetric method. Amer. J. Clin. Pathol. 23:1163, 1953. 3. Fiegel, G., Bargheer, R., and Kulwa, D.: Beeinflussung des Lipidhaushaltes durch anabole Substanzen. Arzneimittel-Forsch. 16:202, 1966.
592 4. King, E. J.: Micro-analysis in Medical Biochemistry. London, Churchill, 1954. 5. Metcalf, W., and Greene, H. G.: A quantitative expression for nitrogen retention with anabolic steroids. Metabolism 12: 899, 1963. 6. Noelle H.: Stickstoffbilanzuntersuchungen zur Uberpriifung anaboler Steroide. Zs.ges.exp.Medizin 145: 138, 1968. Bes:~moumgr;~~ SanTiiz Grundumsatz8. Passmore, R., Strong, J. A., and Ritchie, F. J.: The chemical composition of the tissue lost by obese patients on a reducing regimen. Brit. J. Nutr. 12: 113, 1958. 9. -, -, and -: Water and electrolyte exchanges of obese patients on a reducing regimen. Brit. J. Nutr. 13:17, 1959. 10. Reifenstein, E. C., Jr., Albright, F., and Wells, S. L.: The accumulation, inter-
KRIZEK, STEPANEK
AND SADILEK
pretation, and presentation of data pertaining to metabolic balances, notably those of calcium, phosphorus and nitrogen. J. Clin. Endocr. 5:367, 1945. 11. Russell, G. F. M.: The effect of diets of different composition on weight loss, water and sodium balance in obese patients. Clin. Sci. 22:269, 1962. 12. Searcy, R. L., and Berquist, L. M.: A new color reaction for the quantitation of serum cholesterol. Clin. Chim. Acta 5:192, 1960. 13. Slabochovi, Z., Rath, R., and Placer, Z.: Effect of methandrostenolon and dimethandrostenolo in the dietotherapy of obesity. Nutr. Dieta 7:271, 1965. 14. Stepanek, P., Kfiiek, V., and Sadilek, L.: Die Anwendung von Trijodthyronin und Dianabol bei Abmagerungskuren. Med. Welt 17:2393, 1966.