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TABLE 1. Characteristics of the Men

Subject

Age (years)

Height (cm)

Initial Weight (kg)

Body Mass Index (kg/m)

Weight Change (kg)

5801 28 181.2 62.6 19.06 - 0.8
5802 29 181.5 84.2 25.56 - 3.9
5803 31 175.3 64.8 21.09 + 0.3
5804 22 180.2 58.6 18.03 - 2.0
5805 30 190.4 89.1 24.57 - 0.6
5806 27 181.5 89.5 27.17 - 2.6

3. Most of the subjects lost weight during the study. The rate of weight loss diminished as the experiment proceeded. The nature and significance of this remain unclear.

4. There were no demonstrable biochemical or physiological (work performance) changes, except for a marked fall in BUN.

5. The time required to reach a steady state varied from seven to more than 28 days. The reasons for this variability are not clear.

Reference

1. P.V. Sukhatme and S. Margen "Models for Protein Deficiency," Amer J. Clin. Nutr., 31: 1237-1256 (1978).

TABLE 2 Summary of Initial and Final Laboratory Values

Parameter Subject
5801 5802 5803 5804 5805 5806
Initial Final Initial Final Initial Final Initial Final Initial Final Initial Final
Total protein, g/dl (6.0-8.3)a 6.9 7.2 7.1 7.5 8.0 8.1 7.4 7.6 7.3 7.1 7.2 7.4
Albumin, g/dl (3.5-5.1) 4.2 4.5 4.0 4.8 4.6 5.2b 4.7 5.3b 4.6 5.0 4.2 4.2
BUN, mg/dl (6-25) 12 3b 10 5b 9 2b 15 6 10 4b 11 5b
SGOT, mu/ml (8-45) 24 29 37 23 37 48b 18 26 32 26 31 17
SGPT, mu/ml (3-40) 13 12 24 5 33 44b 11 24 25 11 24 14
GGT, mu/ml (7-51) 18 15 35 16 16 22 11 13 20 18 15 21
LDH, mu/ml (104-235) 134 159 168 180 181 198 163 175 156 148 170 143
Creatinine, mg/dl (0.5-2.0) 0.8 0.9 1.0 0.9 0.9 0.9 0.9 0.9 0.7 0.8 1.1 0.9
HCT, % (37-52) 47.3 42.7 46.8 43.4 44.7 44.1 45.9 44.1 46.9 46.7 47.2 46.0
Haemoglobin, g/dl (12-18) 16.2 15.0 16.0 14.6 15.2 15.1 16.2 15.1 16.5 15.7 16.3 15.7
RBC count, 106/mm3(4.2-6.2) 5.3 5.07 5.17 4.96 5.06 4.95 5.2 4.96 5.42 549 5.47 5.37
WBC, 103/mm3 (4.8-10.8) 7.5 6.9 7.5 6.8 5.6 5.3 5.6 3.9b 5.5 5.6 6.5 6.8
Segs, % (50-70) 61 64 38 43 40 48 57 44 59 51 66 66
Bands, % (3-5)                        
Lymphs, % (20-40) 35 29 49 50 49 46 43 51 34 31 22 25
Monos, % (2-10) 1 4 6 5 3 3   2 4 3 1 1
Eos, % (1-4) 3   5 1 8 3   1 3 5 12 9
Basos, % (0-2)   1 2 1       2        
Atypical lymphs                        
T3 uptake, % (25-35) 28 27 30 26 27 27 27 25 27 26 33 25
T4, m g/dl (4.2-12.5) 7.1 7.6 7.8 7.5 6.1 6.6 10.4 10.1 7.6 8.2 7.1 7.3
Free thyroxine index 2.0 2.1 2.4 1.9 1.7 1.8 2.9 2.5 2.1 2.2 1.9 1.9

a. Reference values in parentheses
b. Values outside reference range

TABLE 3 Diet Composition

Basic Formula

g/subject/ day

Kilocalorie Supplement

Parts by Weight

Egg albumin, dried 17.788 Cornstarch 2.0
Maltodextrins 111.945 Maltodextrins 2.0
Cornstarch 111.945 Sucrose 0.5
Sucrose 22.389 Cottonseed oil 1.0
Cottonseed oil 13.609 Deionized water 7.4
Shortening 38.786    
NaCl 3.556 Total solution 12.9 ml
KCl 4.444 Egg Albumin Supplement Parts by Weight (mgN/g)
CaCO3 0.765
Ca (H2PO4)2H2O 2.568
MgO 0.489
Methylcellulose 3.000 Egg albumin dried 1 126.8
Biotin 0.0002 Deionized water 2  
Deionized water 268.720 Egg albumin solution 42.27 ml
Total 600.000 Trace Mineral Solution g/ subject/ day Mineral mg/ subject/ day
150 g/meal
 
FeCl36H2O 25.52 Iron 10.00
CuSO4 4.43 Copper 3.00
ZnSO47H2O 40.31 Zinc 15.00

TABLE 4. Protein and Energy Intake

Subject No.

Study days

cal/kg

cal/day

Total Protein Intake/Day (g)

Calculated N Intake (g)a

Non-protein N from Extra Cal Solution (g)

Calculated Total N Intake (g)

Laboratory Analysis of Total N Intakeb

Kcal from Protein/Total Cal %

5801 1-6 40 2502.4 22.31 3.57 0.08 3.65 3.68 3.40
7-77 42 2627.5         0.03  
5802 1-77 40 3368.0 30.00 4.80 0.13 4.93 4.96 3.56
              0.05  
5803 1-17 40 2590.0 23.06 3.69 0.08 3.77 3.82 3.47
18-77 41 2654.8         0.06  
5804 1 -14 40 2344.0 20.88 3.34 0.06 3.40 3.43 3.40
15 77 41 2402.6         0.04  
18-77 42 2461.2            
5805 1-6 40 3564.0 31.75 5.08 0.11 5.19 5.24 4.07
7-14 38 3385.8         0.04  
15-77 36 3207.6            
18-77 35 3118.5            
5806 1-9 40 3580.0 31.88 5.10 0.17 5.27 5.30 3.39
10-77 42 3759.0         0 07  

a. 57 mg N/kg body weight
b. Mean of four values standard deviation.

TABLE 5 Individual Formal Exercise Regimen

Subject

Bicycle Ergometer

Treadmill

No.

kg m/min

Minutes

Mph

Grade (%)

Minutes

5801 500 20 3.5 10 20
5802 1,200 25 4.2 10 20
5803 500 20 3.0 10 20
5804 600 15 3.0 10 15
  800 5 6.0 10 5
5805 700 25 4.0 12 20
5806 700 20 4.0 10 20

TABLE 6 Faecal N (g/24 fur): Average of Four Analyses of Two Samples from Pool (Range of Eight Samples)

Day

1

2

3

4

5

6

6-14

0.346

1.070

0.664

0.692

0.826

1.016

0.342-0351

1.058-1.085

0.655-0.675

0.673 0.705

0.815-0.835

1.011-1.024

15-35

0.391

1.000

0.714

0.700

0.812

0.896

0.389-0.395

0.993-1.012

0.705-0.720

0.695-0.713

0.798-0.876

0.888-0.904

35-56

0.523

0.891

0.753

0.730

1.065

0.918

0.516-0.527

0.886-0.895

0.745-0.758

0.724-0.733

1.055-1.078

0.916-0.921

56-77

0.577

0.858

0.577

0.808

0.816

0.863

0.570-0.583

0.850-0.859

0.572-0.578

0.805-0.812

0.807-0.825

0.860-0.868

TABLE 7 Average N Balance (g/day) in Five to Ten Days and during Last Six Days

Average N Subject No.
balance for: 1 2 3 4 5 6
5to 10 days - 1.255 0.21 - 1.125 - 1.782 0.406 - 1.845
For last 6 days - 0.36 0.44 0.38 - 0.168 0.696 0.408
Period of 15 to 77 15 to 77 22 to 77 22 to 77 8 to 77 28 to 77
stationarity (63) (63) (56) (56) (70) (49)
Average N balance (in stationarity) - 0.41 0.70 0.003 - 0.20 0.75 0.18
Expected weight loss or gain (kg) (using 32 as factor) - 0.826 1.411 0.005 - 0.358 1.68 0.282
Observed weight change - 0.40 - 3.30 0.400 - 0.800 - 2.30 - 0.900

4. Investigation of a short-term procedure to evaluate protein quality in adult human subjects - a preliminary report


Nevin S. Scrimshaw, William Rand, Zhi-chien Ho, V.R. Young, and E. Murray

Department of Nutrition and Food Science. Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

It is recognized that the use of experimental animals and biochemical methods of evaluating protein should be verified and calibrated by nitrogen balance studies in human subjects. Because the traditional procedure is costly and time-consuming, it is essential that a shorter and more practical method for evaluating protein quality be developed for worldwide use. This report presents some relevant findings from ongoing experiments in our laboratory.

Experimental Design

Study 1

In 1974 Dr. Myriam Puig of our laboratory conducted an experiment to explore the usefulness of a short-term evaluation of proteins, using the same 12 healthy young men for both a short-term and a standard protocol (1). Her short-term design involved a one-day protein-free diet and six days' adaptation that served as a priming period of low 0.3 9 prot./kg for the ascending design, or high, 0.6 for the descending design, and then ascent or descent in three one-day steps (0.4, 0.5, 0.6. or 0.5, 0.4, 0.3). After approximately a two-week break her subjects completed the standard protocol. Two diets were used, chick-pea and diluted beef, with six subjects on each, three ascending and three descending. The data from this experiment (table 1) suggest that the nitrogen balances for the successive single-day intakes of different levels did not represent a consistent response (see fig. 1). Either the single day was insufficient to reflect the change in intake, or three periods were too few or too close together. Nevertheless, pooling of all data, ascending and descending, gave a zero nitrogen balance intercept not significantly different from the mean of the same subjects studied in the standard manner.

Study 2

Four healthy young men participated (table 2) in the next experiment consisting of five different levels of protein intake for two-day periods each. They received medical and nursing supervision throughout the entire study and maintained relatively constant physical activities from day to day. No clinical changes were noted during the experimental period. Protein was supplied by a soy protein isolate (Supro 710). Caloric intakes were 45 to 48 kcal per kilogram body weight, as estimated to maintain body weight from their intake before the experiment (table 3). Additionally, the subjects received daily vitamin and mineral supplements. and a relatively constant water intake. This diet was provided in three meals a day plus a small snack in the evening.

FIG. 1. Time Course of Urinary Nitrogen (g) for Subject 3 on One-day Ascending
Design. (Circled Data Represent Responses to Intakes of 0.4, 0 5, and 0 6.)

TABLE 1. Urinary Nitrogen (gN)

Contents 0.3 g/kg 0.4 g/kg 0.5 g/kg 0.6 g/kg 0.7 g/kg
Supro 710 23.91 31.9 39.9 47.8 55.8
Polycose 192.0 188.5 185.0 182.0 179.0
Corn oil 118.3 116.7 115.0 113.0 125.5
Sod. chloride 2.0 2.0 2.0 2.0 2.0
Calc. phos. tribasic 2.0 2.0 2.0 2.0 2.0
Potassium phos. tribasic 6.1 6.1 6.1 6.1 6.1
Avicel 5.0 5.0 5.0 5.0 5.0
Water (ml) 400 400 400 400 400

TABLE 2. Physical Characteristics of the Subjects

Subject Weight (kg) Height (cm) Age (years)
W.Z. 77.30 177.8 22
R.S. 61.50 163.8 19
H.L. 69.13 179.7 25
P.M. 65.55 177.8 24
Mean 68.37 174.77 22.5

FIG. 2. Four Subjects on Two-day Ascending Design.

FIG. 3 Four Subjects on Two-day Descending Design.

TABLE 3. Composition of Formula Diet with Soy Protein Isolate (Supro 710) for a 70 kg Man (in grams)

Contents 0.3 g/kg 0.4 g/kg 0.5 g/kg 0.6 g/kg 0.7 g/kg
Supro 710 23.91 31.9 39.9 47.8 55.8
Polycose 192.0 188.5 185.0 182.0 179.0
Corn oil 118.3 116.7 115.0 113.0 125.5
Sod. chloride 2.0 2.0 2.0 2.0 2.0
Calc. phos. tribasic 2.0 2.0 2.0 2.0 2.0
Potassium phos. tribasic 6.1 6.1 6.1 6.1 6.1
Avicel 5.0 5.0 5.0 5.0 5.0
Water (ml) 400 400 400 400 400

The duration of the experiment was 30 days with 15 days on ascending protein intake and 15 days on descending protein intake. Each 15-day period began with a 5-day priming period at a constant level on either a low (for ascending) or high (for descending) level of protein intake. Following this, a different level of protein intake was provided every two days as follows:

Day Priming Period (g/kg/day)
  Ascending Descending
1-5 0.2 0.8
6-7 0.3 0.7
8-9 0.4 0.6
10-11 0.5 0.5
12-13 0.6 0.4
14-15 0.7 0.3

There was a three-day break on an adequate ad libitum diet between the two dietary phases. Urine and faeces were collected daily for analyses; food colour (blue) and cellulose capsules were used as markers for faecal samples. All samples, including food samples, were analysed by the Kjeldahl method.

Results

Figures 2 and 3 show the time course of the nitrogen balances for all subjects. Table 4 shows the balance data for all subjects.

TABLE 4. Nitrogen Balance (gN)

  Ave. Ave. Ball (g pros.)   Inter Req.
Subject FN (g) 0.3 0.4 0.5 0.6 0.7 Slope (mg N/kg) (mg N/kg)
Ascending
1 0.783 -1.83 -0.88 -0.13 -0.44 -0.21 0.30 31.9 107.9
2 1.023 -3.55 -1.84 -0.55 0.66 0.25 0.93 89.3 96.0
3 0.065 -1.38 -0.43 -0.57 -0.10 0.65 0.45 39.9 89.4
4 0.728 -1.06 -0.47 0.41 1.05 1.28 0.60 46.4 77.0
Descending
1 0.918 -003 -0.66 -1.21 -1.57 -2.31 0.44 49.8 113.8
2 0.787 -1.81 -2.01 -2.04 -2.32 -2.47 0.15 43.5 290.0
3 0.641 -0.00 -0.39 -0.88 -1.07 -1.92 0.46 48.3 105.6
4 0.893 -0.71 -0.90 -1.45 -1.54 -1.73 0.36 52.1 145.7

We find:

1. Balance (nitrogen output) responses can be adequately fitted by a straight line, ignoring the priming phase.

2. The ascending design gives consistently lower requirements. and is usually a steeper response leading to a better defined requirement.

3. There is little agreement between results from the ascending and descending phase of the experiment.

Conclusions

A protocol based on a five-to-seven-day priming phase with two days on graded intakes should be explored as a possible alternative to the standard protocol. The critical additional experiment now required is to repeat the study using the same individuals for both the two-day and standard protocols.


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