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Effects of caloric restriction on body composition and total body nitrogen as measured by neutron activation
Effects
of Caloric Restriction on Body Composition and Total Nitrogen as Measured by Neutron Activation Ashok N. Vaswani,
David Vartsky,
Kenneth
J. Ellis, Seiichi Yasumura,
and Stanton
Body
H. Cohn
The purpose of this study was to compare the effects of two isocaloric diets (800 Kcals) on the changes in body composition during weight reduction.
While the protein content of both diets was 70 g, the carbohydrate
that of diet 6 was 70 g. The various (TBK) by “K counting, neutron
total body water (TBW)
activation analysis (PGNAA)
chemistries intervals.
were performed
Seventeen
(group A-10
obese women
subjects).
different
by measuring
the skinfold thickness.
by radioimmunoassay
Routine serum
were done at 4-wk
for the outpatient
study.
for the two groups. The biochemical changes were consistent with the
We conclude that: (1) the technique
long term changes
potassium
and nitrogen)
as well as fat tissue occurred to diet B; however,
Total body potassium
total body nitrogen (TBN) by prompt gamma
At the end of the 12 wk study, the percent changes in the above parameters
to determine
expressed
content of diet A was 10 g and
as follows:
who were at least 30% above ideal body weight volunteered
effectively
less for diet A compared
were determined
by the tritated water technique,
and total body fat was estimated
were not significantly
degree of caloric restriction,
of body composition
every 2 wk and serum insulin and triiodothyronine
subjects, group B-7
of body composition
parameters
of prompt gamma neutron activation analysis can be used
in total body nitrogen
during weight
during weight reduction.
there was no significant difference
reduction,
(2) loss of lean tissue (water,
The loss of TBN in absolute quantities was between
as a percent change from the baseline values, and 13) TBK determination
the two diets when the data was
probably provides the best estimate of
total body fat.
I
T IS well established that hypocaloric weight reducing diets are useful in the management of obehowever, over the sity;’ 3 there is some controversy most effective dietary composition of such regimens in protein sparing. Nitrogen wasting or the demonstration of protein sparing during weight reduction has relied upon balance studies with their inherent errors in collection of body excretions and in the chemical analysis of nitrogen. The development of a direct measurement of total body nitrogen (TBN) by prompt gamma neutron activation analysis (PGNAA) provides a method which is comparable to the metabolic balance technique. The present study was designed to compare the effkacy of two isocaloric diets on the preservation of lean body mass during weight reduction. MATERIALS
AND
METHODS
Selection of Subjects Twenty-twoobese women were accepted for this out patient study. The subjects were at least 30% above ideal body weight (according to the Metropolitan Life Insurance tables, 1959) and showed no evidence of endocrine, hepatic, renal or cardiac diseases. Fasting blood glucose, lipids, liver function tests, electrolytes, uric acid, creatinine and BUN were determined at the time of the initial evaluation, and repeated every 2 wk. Serum insulin and triiodothyronine (T,) were determined by radioimmunoassay (Bioscience Laboratory) at 4-wk intervals starting with the first visit. Twenty-four hour urine collections were made at the time of the initial evaluation, and repeated every 2 wk during the I2 wk study: the urine was analyzed for urea, uric acid and electrolytes. Pregnancy tests were done every 2 wk and electrocardiograms were repeated at least every 4 wk. Seventeen out of 22 women completed the 12 wk protocol, and were readmitted to the Medical Research Center (MRC) for repeat studies. Of these 17 subjects there were 3 black women in group A.
Metabolism. Vol. 32, No. 2
(February),
1983
Special Studies Body composition studies were performed at the time of the first admission and repeated at I2 wk. Total body water (TBW) was determined from a blood sample taken 3 hr after the patient drank 50 &i of tritiated water mixed with orange juice. The coefficient of variation for the measurement of TBW with this technique is less than 1%.4Total body potassium (TBK) was determined by counting 40K in the 54 detector Brookhaven Whole Body Counter.5 This facility, with its on-line computer. corrects for body size and the internal location of the naturally occurring isotope of potassium (40K). The precision and accuracy of this technique is k34. in the Alderson Phantom. Total body nitrogen (TBN) was determined by prompt gamma neutron activation.” The subjects were scanned in both the supine and prone positions on a motor driven bed which passes over a collimated neutron beam measuring I3 x 60 cm at the level of the bed. A Z3sPu. Be source delivers neutrons for the ‘“N (n, y) IiN reaction, and the gamma emission from the body is detected by two 15.24 x 15.24 cm Nal (Tl) crystals coupled to photomultiplier tubes and to an analyzer. In addition to nitrogen. the gamma rays from the neutron capture provides a way to measure hydrogen. The use of total body hydrogen (TBH) as an internal standard reduces the errors due to the effect of body size and shape. The total body nitrogen is determined by the formula: TBN = K x N/H x TBH where K is a constant determined from irradiation of an Alderson
From the Brookhaven National Laboratory, Medical Research Center, Upton, New York, and the Nassau Hospital, Department of Medicine, Mineola. New York. Received for publication July 13, 1982. Supported in part by grant IROI AM.27533~01. Additional parts of the study are supported by DE-AC02-76CH00016 with the U.S. Department of Energy. Accordingly, the U.S. Government retains a nonexclusive royalty-free license to publish or reproduce the published form of this contribution. or allow others to do so. for the U.S. Government purposes. Address reprint requests to A. N. Vaswani. M.D., Nassau Hospital, Department of Medicine,Minneola. NY 11501. d 1983 bv Grune & Stratton. Inc.
185
VASWANI ET AL.
186
Phantom containing known amounts of nitrogen and hydrogen, and N/H is the corrected ratio for the differences in thickness between the patient and the Alderson Phantom. The precision of the method as determined from multiple measurements of Alderson Phantoms is +4%. The total scanning time is less than 20 min and the radiation dose delivered by this procedure is 26 mrem (0.0026 rad). Total body fat (TBF) was estimated by measuring skinfolds with the Lange calipers. Four conventional sites (biceps, triceps, suprailiac and subscapular) were selected and the mean value of three observations at each site was recorded. Body density and hence total body fat was calculated using standard equations.’ We have previously shown that the skinfold method is a poor indicator of the body fat content in normal subjects and cancer patients.* However, the data for TBF was included in the study to determine the utility of the technique in measuring TBF changes during weight reduction.
Dietary Protocol All subjects were fed an 800 Kcal (3200 Kj) diet starting on the first day of the initial hospitalization. Diet A consisted of meat, poultry, fish, eggs, cheese and salad greens and provided about 10 g of carbohydrate. Subjects consuming diet B were given an additional 30 g of glucose twice daily as a concentrated solution (solution B), while those on diet A were given tap water twice daily as solution A. Everyone in the study received daily supplements of potassium (25 mEq b.i.d.), 1 g of elemental calcium daily in divided doses and one multivitamin with minerals as well as 1 mg of folic acid daily. Milk, sugar substitutes, sugar-free gum and alcoholic beverages were not permitted for the duration of the study. All participants were ambulatory but were not permitted to initiate any new form of exercise program during the 12 wk. Following the 5-day stay at the MRC, the subjects were followed on an out-patient basis every 2 wk. At each outpatient visit, urine was examined for ketones, diet diaries were checked for adherence to the prescribed calories, and appropriate solutions (A or B) were dispensed by the pharmacist. Seventeen subjects completed the dietary protocol and were readmitted to the MRC for repeat studies as well as initiation of refeeding. The refeeding protocol will not be discussed in detail. In brief, all subjects consumed about 1000 Kcal daily, (4000 Kj) by the end of 5 days of refeeding.
Statistics The statistical significance of the means of the various parameters of body composition was evaluated by the Student’s t-test. Multivariate analysis was carried out for the various parameters of body composition using the Hotelling T square. Analysis of variance for repeated measurements was carried out for the biochemical changes. The data were analyzed using the Statistical Package for Social Sciences (SPSS) and BMDP Biomedical Computer Programs PSeries (1979). All values are reported as mean it S.D.
RESULTS
Mean age (+ S.E.) of the 10 subjects consuming diet A was not significantly different from that of the 7 subjects following diet B (30.3 -t 2.1 and 30.9 + 1.7 respectively). The baseline weight and other parameters of body composition were not significantly different between the two groups (Table 1). We have previously reported racial differences in body composition;’ however, no significant differences were observed in the present study. All subjects tolerated the diets well although postural hypotension was noted and fatigue was a common complaint. These symptoms were treated by increasing the salt intake and rest, respectively. The use of a carbohydrate-free bulk laxative alleviated the problem of constipation, while none of the subjects required medications to lower the uric acid level. The changes in body composition during 12 wk of dieting are shown in Table 1. Total body fat was calculated as the difference between the weight and the lean body mass derived from total body potassium determinations. The body weight, TBF, TBW, and TBK for each group decreased significantly when compared to the baseline; the change in TBN. however, was minimal and failed to reach statistical significance for group A. There was no significant difference between the two groups for any of the measured parameters, either at the baseline or at the end of the 12 wk, although the absolute values for all components of body composition were greater for group B subjects (Table 1). When group A was compared to group B (expressed as a percent change from the baseline) no significant differences were noted in the above parameters (Fig. 1). Moreover, no significant difference was found between the two groups of subjects using the Hotelling T square for five variables: weight, TBF, TBW, TBK, and TBN (f-value 0.6725, p-value 0.645). The serum insulin levels at the start of the study were comparable for both groups, 8.6 + 4.7 pu/ml and 8.1 + 4.1 vu/ml for diets A and B respectively. The decline in serum insulin following weight reduction
187
EFFECTS OF CALORIE RESTRICTION
Total body fat was calculated as the Fig. 1. between the weight and the lean body mass derived body potassium determination.
difference from total
not statistically significant either between the groups or in comparison to the baseline value for each group (6.0 * 3.0 pu/ml and 5.7 + 3.1 pu/ml for diets A and B respectively). The fasting blood glucose was not significantly different for the two groups at the baseline, 94 + 7 mg/dl and 97 f 6 mg/dl for groups A and B respectively, or at the end of 12 wk, 85 * 10 mg/dl and 92 + 8 mg/dl respectively. Baseline triiodothyronine (T3) for groups A and B was 138 + 16 ng/dl and 159 + 21 ng/dl respectively (p < 0.05) and 101 i43 ng/dl for diet A and 149 + 27 ng/dl for diet B (p < 0.25) at 12 wk. The initial serum uric acid was 5.4 k 0.8 mg/dl and 6.5 + 1.0 mg/dl respectively for groups A and B (p < 0.05). No significant difference was noted for the values at 12 wk, 5.8 f 0.8 mg/dl and 6.2 + 1.3 mg/dl for the two diets. However, the percent changes (+ S.D.) in serum uric acid and triiodothyronine were not significantly different between the two diets at the end of 12 wk. was
DISCUSSION
Although the adaptive changes during starvation or caloric restriction tend to conserve nitrogen, protein supplementation is critical in minimizing nitrogen depletion.” Moreover, it has been observed that ingestion of approximately I .0-l .5 g of protein/kg ideal body weight per day can modify nitrogen loss and even produce a positive nitrogen balance in spite of a reduced caloric intake.’ While protein intake is undoubtedly instrumental in minimizing nitrogen loss
during hypocaloric weight reduction, the utility of adding carbodydrate to reduce nitrogen depletion has been debated.‘,” Studies comparing protein diets with mixed diets have not adequately answered this controversy since most investigations have been short term or too restrictive in calories.“-” Moreover, very few studies have used an adequate amount of carbohydrate in the hypocaloric regimen to make a meaningful impact on nitrogen economy. Yang and Van Itallie, using conventional balance techniques, have shown no significant difference in nitrogen balance on either an 800 cal mixed or ketogenic diet over a 10 day period.” These investigators reached similar conclusions when their studies were extended to 64 days.lh The limitations and problems associated with conventional balance studies have been reviewed by Hegsted.” Prompt gamma neutron activation analysis offers several distinct advantages for the measurement of total body nitrogen. The short duration for the procedure is preferred by patients to the repetitive, cumbersome collection of excreta required for balance studies. Moreover, the conventional balance technique merely determines a change in nitrogen over a period of time, while the determination of the absolute levels for the measured nitrogen can accurately reflect the percent change. Secondly, with a precision of ~t4%, the direct, noninvasive measurement of the absolute quantity of TBN is convenient and comparable to any of the other available methods to determine total body nitrogen. Overall, prompt gamma neutron activation analysis is useful for the evaluation of long term sequential changes in TBN and can be used to study the patients outside the confines of a metabolic unit. Total body potassium measured by “OK counting has been successfully employed by several investigators for body composition studies.‘8,‘9 Pierson et al.‘” used this technique to determine the changes in adiposefree mass in five obese subjects maintained on approximately 800 Kcal (3200 Kj) daily for a period of 46 days. Of the 17.4 kg mean weight loss achieved by these subjects, 66% was adipose tissue and the remainder, (34%) adipose-free mass. The mean loss of lean tissue based on TBK determination in the present study was 3 1%. It is important to point out that the different components of lean tissue, i.e., water, potassium and nitrogen, may respond or adapt differently during weight reduction. In this regard, nitrogen balance, as measured by prompt gamma neutron activation analysis, was negative for both groups during the 12 wk of dieting. However, the nitrogen loss was minimal and failed to reach statistical significance for diet A. Whether this finding was due to individual variability in the loss of
VASWANI
188
nitrogen or the absence of dietary carbohydrate awaits further confirmation. It should be noted however, that the measurement of absolute values of any of the components of lean tissues was advantageous, since the alterations in these parameters could be expressed as a percent change thereby normalizing the individual values. In this regard, there was no significant difference in the percent change between the two diets for any of the measured parameters. Moreover, multivariate analysis for the loss of two important intracellular components, potassium and nitrogen, indicated no overall difference between the two diets (f-value 0.9314,p-value 0.419). It is also interesting to note that on the basis of skinfold thickness measurement, the loss of total body fat was estimated to be 6.75 f 1.98 kg and 7.51 f 2.90 kg for diets A and B respectively. When these values were compared to the TBF calculated from TBK determination, (Table 1) the skinfold technique underestimated the TBF loss by 20.5% and 12.5% for diets A and B respectively. However, since skinfold thickness actually measures adipose tissue rather than “fat,” and fat accounts for about 80%-85% of adipose tissue, the difference between the two techniques could be easily reconciled. The biochemical changes during the 12 wk of weight reduction reflected the adaptive changes to the two diets. Normoglycemia was maintained, while hyper-
ET AL.
uricemia occured, as expected, with normalization of uric acid by the end of 12 wk. The reduction in T, was greater for group A subjects, however, the changes in glucose, uric acid and triiodothyronine, expressed as a percent change from the baseline values, were not significantly different between the two diets. In summary, it can be concluded that, (1) the technique of prompt gamma neutron activation analysis was used effectively to determine long-term changes in total body nitrogen during weight reduction, (2) of all components of lean body mass, total body nitrogen showed the maximal resistance to change, however, the loss of lean tissue, expressed as a percent change, was comparable for the two diets, suggesting that caloric restriction rather than dietary composition was responsible for the observed changes in the measured parameters, and (3) estimation of body fat by TBK determination is probably the best available technique. ACKNOWLEDGMENT The authors wish to acknowledge the contribution to the study of the following persons: M. Stravino. CRT, LRT, and J Rothmann, who performed the total body potassium measurements; A.F. LoMonte, M.S., who performed the total body water measurements; S. Minton. B.S., A. Kronenberg, B.S. and R. Skelaney for their participation in the measurement of total body nitrogen; V. Brooks, R.N. and Nursing Staff: J. Matkovitch, Patient Coordinator; Kathleen Roberts and F. O’Brien for typing the manuscript.
REFERENCES 1. MacLean WC, Graham GG: The effect of level of protein intake in isoenergetic diets on energy utilization. Am J Clin Nutr 32:1381-1387,1979 2. Bistrian BR, Winterer J, Blackbur GL, et al: Effect of a protein-sparing diet and brief fast on nitrogen metabolism in mildly obese subjects. J Lab Clin Med 89:1030, 1977 3. Jourdan M, Margen S, Bradfield RB: Protein-sparing effect in obese women fed low calorie diets. Am J Clin Nutr 27:3-12, 1974 4. Vaughn BE, Boling EA: Rapid assay procedures for tritium labeled water in body fluids. J Lab CLin Med 57: 159- 166, 196 1 5. Cohn SH, Dombrowski CS, Pate HR. et al: A whole-body counter with an invariant response to radionuclide distribution and body size. Physics Med Biol 14:645-658, 1969 6. Vartsky D, Ellis KJ, Cohn SH: In vivo measurement of body nitrogen by analysis of prompt gammas from neutron capture. J Nucl Med 20:1158-l 165, 1979 7. Durnin JVGA, Rahaman MM: The assessment of the amount of fat in the human body from measurements of skin-fold thickness. Br J Nutr 21:681-689,1967 8. Cohn SH, Ellis, KJ, et al: Comparison of methods of estimating body fat in normal subjects and cancer patients. Am J Clin Nutr 34:2839-2846, 1981 9. Cohn SH, Abesamis C, Yasumura S, et al: Comparative skeletal mass and radial bone mineral content in black and white women. Metabolism Vol 26 No. 2, 1977 10. Marliss EB, Murray F, Nakhooda A: The metabolic response to hypocaloric protein diets in obese man. J Clin Invest 16:468-479, 1978 11. DeHaven J, Sherwin R, Hendler R, et al: Nitrogen and
sodium balance and sympathetic nervous system activity in obese subjects treated with a low caloric protein or mixed diet. N Eng J Med 302:4777482, 1980 12. Howard L, Dobs A. Chodos R, et al: A comparison of administering protein alone and protein plus glucose on nitrogen balance. Am J Clin Nutr 31:226-9, 1978 13. Zollner N, Keller C: A 300 kcal (1.2MJ) diet using conventional food. International Journal of Obesity 5:217-220, 1981 14. Contaldo F, Di Biase G, Fischetti A, et al: Evaluation of the safety of very-low-calorie diets in the treatment of severely obese patients in a metabolic ward. International Journal of Obesity 5:221-226, 1981 15. Yang M-U, Van Itallie TB: Composition of weight lost during short-term weight reduction. J Clin Investigation. 58:722-730. 1976 16. Yang M-U, Barbosa-Saldivar JL, Pi-Sunyer FX, et al: Metabolic effects of substituting carbohydrates for protein in a low calorie diet: a prolonged study in obese patients. Intl J Obesity 5:23 1. 198 I 17. Hegsted
DM: Balance Studies. J of Nutr
106:307,
18. Cohn SH, Vaswani AN, Zani I, et al: Changes chemical composition with age measured by total-body activation. Metabolism. 25: 1, 1976
1976 in body neutron
19. Jourdan MH, Bradfield RB: Body composition changes during weight loss estimated from energy, nitrogen, sodium, and potassium balances. Am J Clin Nutr 26:144-149, 1973 20. Pierson RN Jr., Wang J, Yang M-U, et al: The assessment of human body composition during weight reduction: Evaluation of a new model for clinical studies. J Nutr 106:1694-1701, 1976
of Caloric Restriction on Body Composition and Total Nitrogen as Measured by Neutron Activation Ashok N. Vaswani,
David Vartsky,
Kenneth
J. Ellis, Seiichi Yasumura,
and Stanton
Body
H. Cohn
The purpose of this study was to compare the effects of two isocaloric diets (800 Kcals) on the changes in body composition during weight reduction.
While the protein content of both diets was 70 g, the carbohydrate
that of diet 6 was 70 g. The various (TBK) by “K counting, neutron
total body water (TBW)
activation analysis (PGNAA)
chemistries intervals.
were performed
Seventeen
(group A-10
obese women
subjects).
different
by measuring
the skinfold thickness.
by radioimmunoassay
Routine serum
were done at 4-wk
for the outpatient
study.
for the two groups. The biochemical changes were consistent with the
We conclude that: (1) the technique
long term changes
potassium
and nitrogen)
as well as fat tissue occurred to diet B; however,
Total body potassium
total body nitrogen (TBN) by prompt gamma
At the end of the 12 wk study, the percent changes in the above parameters
to determine
expressed
content of diet A was 10 g and
as follows:
who were at least 30% above ideal body weight volunteered
effectively
less for diet A compared
were determined
by the tritated water technique,
and total body fat was estimated
were not significantly
degree of caloric restriction,
of body composition
every 2 wk and serum insulin and triiodothyronine
subjects, group B-7
of body composition
parameters
of prompt gamma neutron activation analysis can be used
in total body nitrogen
during weight
during weight reduction.
there was no significant difference
reduction,
(2) loss of lean tissue (water,
The loss of TBN in absolute quantities was between
as a percent change from the baseline values, and 13) TBK determination
the two diets when the data was
probably provides the best estimate of
total body fat.
I
T IS well established that hypocaloric weight reducing diets are useful in the management of obehowever, over the sity;’ 3 there is some controversy most effective dietary composition of such regimens in protein sparing. Nitrogen wasting or the demonstration of protein sparing during weight reduction has relied upon balance studies with their inherent errors in collection of body excretions and in the chemical analysis of nitrogen. The development of a direct measurement of total body nitrogen (TBN) by prompt gamma neutron activation analysis (PGNAA) provides a method which is comparable to the metabolic balance technique. The present study was designed to compare the effkacy of two isocaloric diets on the preservation of lean body mass during weight reduction. MATERIALS
AND
METHODS
Selection of Subjects Twenty-twoobese women were accepted for this out patient study. The subjects were at least 30% above ideal body weight (according to the Metropolitan Life Insurance tables, 1959) and showed no evidence of endocrine, hepatic, renal or cardiac diseases. Fasting blood glucose, lipids, liver function tests, electrolytes, uric acid, creatinine and BUN were determined at the time of the initial evaluation, and repeated every 2 wk. Serum insulin and triiodothyronine (T,) were determined by radioimmunoassay (Bioscience Laboratory) at 4-wk intervals starting with the first visit. Twenty-four hour urine collections were made at the time of the initial evaluation, and repeated every 2 wk during the I2 wk study: the urine was analyzed for urea, uric acid and electrolytes. Pregnancy tests were done every 2 wk and electrocardiograms were repeated at least every 4 wk. Seventeen out of 22 women completed the 12 wk protocol, and were readmitted to the Medical Research Center (MRC) for repeat studies. Of these 17 subjects there were 3 black women in group A.
Metabolism. Vol. 32, No. 2
(February),
1983
Special Studies Body composition studies were performed at the time of the first admission and repeated at I2 wk. Total body water (TBW) was determined from a blood sample taken 3 hr after the patient drank 50 &i of tritiated water mixed with orange juice. The coefficient of variation for the measurement of TBW with this technique is less than 1%.4Total body potassium (TBK) was determined by counting 40K in the 54 detector Brookhaven Whole Body Counter.5 This facility, with its on-line computer. corrects for body size and the internal location of the naturally occurring isotope of potassium (40K). The precision and accuracy of this technique is k34. in the Alderson Phantom. Total body nitrogen (TBN) was determined by prompt gamma neutron activation.” The subjects were scanned in both the supine and prone positions on a motor driven bed which passes over a collimated neutron beam measuring I3 x 60 cm at the level of the bed. A Z3sPu. Be source delivers neutrons for the ‘“N (n, y) IiN reaction, and the gamma emission from the body is detected by two 15.24 x 15.24 cm Nal (Tl) crystals coupled to photomultiplier tubes and to an analyzer. In addition to nitrogen. the gamma rays from the neutron capture provides a way to measure hydrogen. The use of total body hydrogen (TBH) as an internal standard reduces the errors due to the effect of body size and shape. The total body nitrogen is determined by the formula: TBN = K x N/H x TBH where K is a constant determined from irradiation of an Alderson
From the Brookhaven National Laboratory, Medical Research Center, Upton, New York, and the Nassau Hospital, Department of Medicine, Mineola. New York. Received for publication July 13, 1982. Supported in part by grant IROI AM.27533~01. Additional parts of the study are supported by DE-AC02-76CH00016 with the U.S. Department of Energy. Accordingly, the U.S. Government retains a nonexclusive royalty-free license to publish or reproduce the published form of this contribution. or allow others to do so. for the U.S. Government purposes. Address reprint requests to A. N. Vaswani. M.D., Nassau Hospital, Department of Medicine,Minneola. NY 11501. d 1983 bv Grune & Stratton. Inc.
185
VASWANI ET AL.
186
Phantom containing known amounts of nitrogen and hydrogen, and N/H is the corrected ratio for the differences in thickness between the patient and the Alderson Phantom. The precision of the method as determined from multiple measurements of Alderson Phantoms is +4%. The total scanning time is less than 20 min and the radiation dose delivered by this procedure is 26 mrem (0.0026 rad). Total body fat (TBF) was estimated by measuring skinfolds with the Lange calipers. Four conventional sites (biceps, triceps, suprailiac and subscapular) were selected and the mean value of three observations at each site was recorded. Body density and hence total body fat was calculated using standard equations.’ We have previously shown that the skinfold method is a poor indicator of the body fat content in normal subjects and cancer patients.* However, the data for TBF was included in the study to determine the utility of the technique in measuring TBF changes during weight reduction.
Dietary Protocol All subjects were fed an 800 Kcal (3200 Kj) diet starting on the first day of the initial hospitalization. Diet A consisted of meat, poultry, fish, eggs, cheese and salad greens and provided about 10 g of carbohydrate. Subjects consuming diet B were given an additional 30 g of glucose twice daily as a concentrated solution (solution B), while those on diet A were given tap water twice daily as solution A. Everyone in the study received daily supplements of potassium (25 mEq b.i.d.), 1 g of elemental calcium daily in divided doses and one multivitamin with minerals as well as 1 mg of folic acid daily. Milk, sugar substitutes, sugar-free gum and alcoholic beverages were not permitted for the duration of the study. All participants were ambulatory but were not permitted to initiate any new form of exercise program during the 12 wk. Following the 5-day stay at the MRC, the subjects were followed on an out-patient basis every 2 wk. At each outpatient visit, urine was examined for ketones, diet diaries were checked for adherence to the prescribed calories, and appropriate solutions (A or B) were dispensed by the pharmacist. Seventeen subjects completed the dietary protocol and were readmitted to the MRC for repeat studies as well as initiation of refeeding. The refeeding protocol will not be discussed in detail. In brief, all subjects consumed about 1000 Kcal daily, (4000 Kj) by the end of 5 days of refeeding.
Statistics The statistical significance of the means of the various parameters of body composition was evaluated by the Student’s t-test. Multivariate analysis was carried out for the various parameters of body composition using the Hotelling T square. Analysis of variance for repeated measurements was carried out for the biochemical changes. The data were analyzed using the Statistical Package for Social Sciences (SPSS) and BMDP Biomedical Computer Programs PSeries (1979). All values are reported as mean it S.D.
RESULTS
Mean age (+ S.E.) of the 10 subjects consuming diet A was not significantly different from that of the 7 subjects following diet B (30.3 -t 2.1 and 30.9 + 1.7 respectively). The baseline weight and other parameters of body composition were not significantly different between the two groups (Table 1). We have previously reported racial differences in body composition;’ however, no significant differences were observed in the present study. All subjects tolerated the diets well although postural hypotension was noted and fatigue was a common complaint. These symptoms were treated by increasing the salt intake and rest, respectively. The use of a carbohydrate-free bulk laxative alleviated the problem of constipation, while none of the subjects required medications to lower the uric acid level. The changes in body composition during 12 wk of dieting are shown in Table 1. Total body fat was calculated as the difference between the weight and the lean body mass derived from total body potassium determinations. The body weight, TBF, TBW, and TBK for each group decreased significantly when compared to the baseline; the change in TBN. however, was minimal and failed to reach statistical significance for group A. There was no significant difference between the two groups for any of the measured parameters, either at the baseline or at the end of the 12 wk, although the absolute values for all components of body composition were greater for group B subjects (Table 1). When group A was compared to group B (expressed as a percent change from the baseline) no significant differences were noted in the above parameters (Fig. 1). Moreover, no significant difference was found between the two groups of subjects using the Hotelling T square for five variables: weight, TBF, TBW, TBK, and TBN (f-value 0.6725, p-value 0.645). The serum insulin levels at the start of the study were comparable for both groups, 8.6 + 4.7 pu/ml and 8.1 + 4.1 vu/ml for diets A and B respectively. The decline in serum insulin following weight reduction
187
EFFECTS OF CALORIE RESTRICTION
Total body fat was calculated as the Fig. 1. between the weight and the lean body mass derived body potassium determination.
difference from total
not statistically significant either between the groups or in comparison to the baseline value for each group (6.0 * 3.0 pu/ml and 5.7 + 3.1 pu/ml for diets A and B respectively). The fasting blood glucose was not significantly different for the two groups at the baseline, 94 + 7 mg/dl and 97 f 6 mg/dl for groups A and B respectively, or at the end of 12 wk, 85 * 10 mg/dl and 92 + 8 mg/dl respectively. Baseline triiodothyronine (T3) for groups A and B was 138 + 16 ng/dl and 159 + 21 ng/dl respectively (p < 0.05) and 101 i43 ng/dl for diet A and 149 + 27 ng/dl for diet B (p < 0.25) at 12 wk. The initial serum uric acid was 5.4 k 0.8 mg/dl and 6.5 + 1.0 mg/dl respectively for groups A and B (p < 0.05). No significant difference was noted for the values at 12 wk, 5.8 f 0.8 mg/dl and 6.2 + 1.3 mg/dl for the two diets. However, the percent changes (+ S.D.) in serum uric acid and triiodothyronine were not significantly different between the two diets at the end of 12 wk. was
DISCUSSION
Although the adaptive changes during starvation or caloric restriction tend to conserve nitrogen, protein supplementation is critical in minimizing nitrogen depletion.” Moreover, it has been observed that ingestion of approximately I .0-l .5 g of protein/kg ideal body weight per day can modify nitrogen loss and even produce a positive nitrogen balance in spite of a reduced caloric intake.’ While protein intake is undoubtedly instrumental in minimizing nitrogen loss
during hypocaloric weight reduction, the utility of adding carbodydrate to reduce nitrogen depletion has been debated.‘,” Studies comparing protein diets with mixed diets have not adequately answered this controversy since most investigations have been short term or too restrictive in calories.“-” Moreover, very few studies have used an adequate amount of carbohydrate in the hypocaloric regimen to make a meaningful impact on nitrogen economy. Yang and Van Itallie, using conventional balance techniques, have shown no significant difference in nitrogen balance on either an 800 cal mixed or ketogenic diet over a 10 day period.” These investigators reached similar conclusions when their studies were extended to 64 days.lh The limitations and problems associated with conventional balance studies have been reviewed by Hegsted.” Prompt gamma neutron activation analysis offers several distinct advantages for the measurement of total body nitrogen. The short duration for the procedure is preferred by patients to the repetitive, cumbersome collection of excreta required for balance studies. Moreover, the conventional balance technique merely determines a change in nitrogen over a period of time, while the determination of the absolute levels for the measured nitrogen can accurately reflect the percent change. Secondly, with a precision of ~t4%, the direct, noninvasive measurement of the absolute quantity of TBN is convenient and comparable to any of the other available methods to determine total body nitrogen. Overall, prompt gamma neutron activation analysis is useful for the evaluation of long term sequential changes in TBN and can be used to study the patients outside the confines of a metabolic unit. Total body potassium measured by “OK counting has been successfully employed by several investigators for body composition studies.‘8,‘9 Pierson et al.‘” used this technique to determine the changes in adiposefree mass in five obese subjects maintained on approximately 800 Kcal (3200 Kj) daily for a period of 46 days. Of the 17.4 kg mean weight loss achieved by these subjects, 66% was adipose tissue and the remainder, (34%) adipose-free mass. The mean loss of lean tissue based on TBK determination in the present study was 3 1%. It is important to point out that the different components of lean tissue, i.e., water, potassium and nitrogen, may respond or adapt differently during weight reduction. In this regard, nitrogen balance, as measured by prompt gamma neutron activation analysis, was negative for both groups during the 12 wk of dieting. However, the nitrogen loss was minimal and failed to reach statistical significance for diet A. Whether this finding was due to individual variability in the loss of
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nitrogen or the absence of dietary carbohydrate awaits further confirmation. It should be noted however, that the measurement of absolute values of any of the components of lean tissues was advantageous, since the alterations in these parameters could be expressed as a percent change thereby normalizing the individual values. In this regard, there was no significant difference in the percent change between the two diets for any of the measured parameters. Moreover, multivariate analysis for the loss of two important intracellular components, potassium and nitrogen, indicated no overall difference between the two diets (f-value 0.9314,p-value 0.419). It is also interesting to note that on the basis of skinfold thickness measurement, the loss of total body fat was estimated to be 6.75 f 1.98 kg and 7.51 f 2.90 kg for diets A and B respectively. When these values were compared to the TBF calculated from TBK determination, (Table 1) the skinfold technique underestimated the TBF loss by 20.5% and 12.5% for diets A and B respectively. However, since skinfold thickness actually measures adipose tissue rather than “fat,” and fat accounts for about 80%-85% of adipose tissue, the difference between the two techniques could be easily reconciled. The biochemical changes during the 12 wk of weight reduction reflected the adaptive changes to the two diets. Normoglycemia was maintained, while hyper-
ET AL.
uricemia occured, as expected, with normalization of uric acid by the end of 12 wk. The reduction in T, was greater for group A subjects, however, the changes in glucose, uric acid and triiodothyronine, expressed as a percent change from the baseline values, were not significantly different between the two diets. In summary, it can be concluded that, (1) the technique of prompt gamma neutron activation analysis was used effectively to determine long-term changes in total body nitrogen during weight reduction, (2) of all components of lean body mass, total body nitrogen showed the maximal resistance to change, however, the loss of lean tissue, expressed as a percent change, was comparable for the two diets, suggesting that caloric restriction rather than dietary composition was responsible for the observed changes in the measured parameters, and (3) estimation of body fat by TBK determination is probably the best available technique. ACKNOWLEDGMENT The authors wish to acknowledge the contribution to the study of the following persons: M. Stravino. CRT, LRT, and J Rothmann, who performed the total body potassium measurements; A.F. LoMonte, M.S., who performed the total body water measurements; S. Minton. B.S., A. Kronenberg, B.S. and R. Skelaney for their participation in the measurement of total body nitrogen; V. Brooks, R.N. and Nursing Staff: J. Matkovitch, Patient Coordinator; Kathleen Roberts and F. O’Brien for typing the manuscript.
REFERENCES 1. MacLean WC, Graham GG: The effect of level of protein intake in isoenergetic diets on energy utilization. Am J Clin Nutr 32:1381-1387,1979 2. Bistrian BR, Winterer J, Blackbur GL, et al: Effect of a protein-sparing diet and brief fast on nitrogen metabolism in mildly obese subjects. J Lab Clin Med 89:1030, 1977 3. Jourdan M, Margen S, Bradfield RB: Protein-sparing effect in obese women fed low calorie diets. Am J Clin Nutr 27:3-12, 1974 4. Vaughn BE, Boling EA: Rapid assay procedures for tritium labeled water in body fluids. J Lab CLin Med 57: 159- 166, 196 1 5. Cohn SH, Dombrowski CS, Pate HR. et al: A whole-body counter with an invariant response to radionuclide distribution and body size. Physics Med Biol 14:645-658, 1969 6. Vartsky D, Ellis KJ, Cohn SH: In vivo measurement of body nitrogen by analysis of prompt gammas from neutron capture. J Nucl Med 20:1158-l 165, 1979 7. Durnin JVGA, Rahaman MM: The assessment of the amount of fat in the human body from measurements of skin-fold thickness. Br J Nutr 21:681-689,1967 8. Cohn SH, Ellis, KJ, et al: Comparison of methods of estimating body fat in normal subjects and cancer patients. Am J Clin Nutr 34:2839-2846, 1981 9. Cohn SH, Abesamis C, Yasumura S, et al: Comparative skeletal mass and radial bone mineral content in black and white women. Metabolism Vol 26 No. 2, 1977 10. Marliss EB, Murray F, Nakhooda A: The metabolic response to hypocaloric protein diets in obese man. J Clin Invest 16:468-479, 1978 11. DeHaven J, Sherwin R, Hendler R, et al: Nitrogen and
sodium balance and sympathetic nervous system activity in obese subjects treated with a low caloric protein or mixed diet. N Eng J Med 302:4777482, 1980 12. Howard L, Dobs A. Chodos R, et al: A comparison of administering protein alone and protein plus glucose on nitrogen balance. Am J Clin Nutr 31:226-9, 1978 13. Zollner N, Keller C: A 300 kcal (1.2MJ) diet using conventional food. International Journal of Obesity 5:217-220, 1981 14. Contaldo F, Di Biase G, Fischetti A, et al: Evaluation of the safety of very-low-calorie diets in the treatment of severely obese patients in a metabolic ward. International Journal of Obesity 5:221-226, 1981 15. Yang M-U, Van Itallie TB: Composition of weight lost during short-term weight reduction. J Clin Investigation. 58:722-730. 1976 16. Yang M-U, Barbosa-Saldivar JL, Pi-Sunyer FX, et al: Metabolic effects of substituting carbohydrates for protein in a low calorie diet: a prolonged study in obese patients. Intl J Obesity 5:23 1. 198 I 17. Hegsted
DM: Balance Studies. J of Nutr
106:307,
18. Cohn SH, Vaswani AN, Zani I, et al: Changes chemical composition with age measured by total-body activation. Metabolism. 25: 1, 1976
1976 in body neutron
19. Jourdan MH, Bradfield RB: Body composition changes during weight loss estimated from energy, nitrogen, sodium, and potassium balances. Am J Clin Nutr 26:144-149, 1973 20. Pierson RN Jr., Wang J, Yang M-U, et al: The assessment of human body composition during weight reduction: Evaluation of a new model for clinical studies. J Nutr 106:1694-1701, 1976