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Room air was circulated at 23 room-volumes per hour atid filtered by .... EXPERIMENT E. EXPERIMEN". (V/mffi. (V/m). GROUP 1: C. -GROUP 4z 20. (. (v/- !sJ.
EXTREMELY LOW F QENCY ERTA r~ll 45G,- ELECTRIJC :FIELD EXPOS-URE of RATS A SEARCH FOR GROWTH, FOOD0, AN ATER -CONSUMPTIO-N, BLOOD' METABOLITE, HEMATOLOGICAL, -AND PATHOLOGICAL CH4ANGES del

N.Mathewson G. M. Qosta S. G. Levin S. S. Diczmond M. E. Ekcstrom

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N. S., 4 vlathewson, G. M./9osta, S. G./Levin, 's. s. 'DiamondaW' M. E. /Ekstrom 9PEFRIGORGANIZATION NAME AND AOL.RESe,

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4'rhree hundred eighty-four yoting male rats were exposed to 45-Hz, vertical electric fielda in nonmetallic cages, in an attemot to detect alteration of growth, food and water consumption, alterations in selected hematological and seruim biochemical values, and pathologicpd changes. Three experiments were performed, each using six groups of 16 aniilas exposed to field strength;a of 0, 2, 1G, 20, 50, and 100 1/in (RMS). All variables were statistically analyzed for difforences and the possible ex .stence of a "dose effect relationship". A further experiment (C)

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ABSTRACT (continued) used 48 control animals and 48 animals exposed to 20 V/m (RMS) to minimize the possibility of missing a true alteration. Although some differences were found in three experiments (E, F, and H) neither a dose effect relationship nor a biological effect due to exposure was observed. iriment G, no statistical differences (p< 0. 05) were observed for any variables~it was concluded that no alterations in growth, food consumption, or water consumption resulted from exposure to extremely low frequency (ELF) electric fields. Neither serum or plasma concentrations of total protein, globulin, glucose, cholesterol, triglycerides and total lipid nor hematological values for red blood cells, white blood cells, segmented neutrophils, lymphocytes, monocytes, eosinophils, hematocrit or hemoglobin appear to be influenced by Z.LF fields. ,41 addition, necropsy and histopathological examination of tissue from 16 organ dtstems did not reveal any changes that could be attributed to electric fields. Projdct was sponsored by the U. S. Naval Medical Research and Development Command, contract number XSB09.

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PRE FACE The authors gratefully acknowledge the valuable assistance of S. A. Oliva for designing and maintaining the electronic systems of the exposure facility, C. A. McIntire III and W. E. Jackson III for performing the computer programming and statistical analyses, J. E. Egan for the hematological analyses, and G. D. Lee for preparation of the tissue specimens.

We further acknowledge

the invaluable efforts of A. L. Miller, A. E. Cummings, and P. W. Jones for the care and timely manner in wh'ch these experiments were performed.

This

project was sponsored by the U. S. Naval Medical Research and Development Command, contract number XSB09.

I"1 1'

TABLE OF CONTENTS Page Preface

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Introduction...................................

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Materials and Procedures ..............

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Biochemical analysis Hematology . . .

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Animrals.................... Irradiation facility..................8 Experimentalidesign

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Pathology.....................12 Statistical analysis...................12 Results.........................13 Analysis of growth, food consumption and water consumption during 45-Hz electric field exposure............13 Analysis of biochemical and hematological. results versus 45-Hz field strength.................19. Pathology versus 45-Hz field strength............26 Two-way ANOVA for 45- and 60-Hz effects ......... Discussion and Conclusions

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References........................32 AppendixA......

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AppendixB.......................43

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Nat

LIST OF FIGURES Page Figure 1. Graphical summary of the average body weight and daily growth for each group during exposure . . . . . Figure 2.

Figure 3,

Figure 4.

Figure 5.

Figure 6.

Figure 7.

Figure 8.

Graphical summary of food consumption data during exposure ...... ...............

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Graphical summary of water consumption data during exposure ..... ..................

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Summary of the mean growth, food and water consumption data versus 45-Hz field strength ........... .

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Summary of median total protein, globulin and glucose data versus 45-Hz field strength ...........

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Summary of median total lipid, cholesterol and triglyceride data versus 45-Hz field strength

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Summary of median red blood cell, white blood cell anid segmented neutrophil data versus 45-Hz field strength. Summary of median lymphocyte, hezratocrit and hemoglobin data versus 45-Hz field strength .......

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LIST OF TABLES Page Table

1.

Experimental Design and Animal Usage

Table

2.

Electric Field Strengths Applied to Each Chamber for Each Experiment

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The Number of Animals in Each Group of the 45-Hz and 60-Hz Two-Way Analysis of Experiment G.......1

Table

4.

Statistical Summary of the Analysis of Variance of Growth, Food Consumption and Water Consumption

Table

5. Statistical Summary of the Kruskal-Wallis Analysis of the Blood Biochemistry and Hematology Data . .

Table

6.

Statistical Summary of the Two-Way Analysis of Variance on 45-Hz and 60-Hz Field Strengths of ExperimentG................27

Table

7.

Statist-rnal Summary of the Kruskal-Wallis Analysis on 60-Hz Field Strengths of Experin'ent G .........

Table A-i........

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Table a-2........................37 Teble A-3..........

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Table A-4.......................39 Table A-5.......................40

Table A-7

Table B-1.

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Table B-2.......................46 T able B-3.

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Table B3-4............................48 5

LIST OF TABLES (eontlnued) Page

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INTIODUCTION Extremiely low fretquency (ELF) radiation generally denotes ele tronmgtietic radiation having frequencies'from a few hertz (sometimes including zero hertz or de) to several hundred hertz. The natural or ambient levels of ELF radiation have been reviewed by Polk;" 1 however, the extent to which these dlelds may interact with bl';logical systems is still the subject of considerable research.

The largest source of man-made ELF radiation comes from com-

mercial electrical power lines operating at 50 or 60 lIz.

With the recent devel-

opment of the Navy's ELF Communications System for operation at 45 or 75 Hz, yet another man-mt'de source of this radiation will come into existence. It has become apparent that at least certain biological systems can sense and utilize ELF fields.

For example, some birds are affected by weak mag-

netic fields, 18 and it Is proposed that they may use terrestrial magnetic fields to aid in orientation. 20 Kain-djn has presented data suggesting that sharks and rays can locate prey by the weak ELF electric fields they produce, and that some fish may also be able to use terrestrial fields in obtaining orientational and navigational information. 5 Other biological effects from ELF fields have been reported by Goodman et al. on Physarum polycephalum 3 and by GavalasMedici and Day-Magdaleno on monkeys. 2 Recently, large 60-Hz electric fields have been reported to reduce the growth of rats I 0 and mice. 9 Noval et al. 13 have reported that 45-Ilz vertical electric fields at field strengths of up to 100 V/m (RMS) produced reduced growth, reduced abdominal body fat, and altered brain and liver enzyme activities.

The present work represents an attempt to verify the existence of low-

ered growth rates for rats exposed to similar 45-Hz, vertical electric fleld3 and to determine if food and water consumption, serum metabolite concentrations, or the parameters of a complete blood count were perturbed. liminary report of this work has been submitted elsewhere. 12

7

A pre-

MA'I'RIALS AND PROlCEDURES Animals.

Male Sprague-Dawley rats, approximately 180 g, liur(SD)

(I), obtained from llilltop LAb Aniimals, Inc., Scottdale, Pennsylvania, were used for all experiments.

Animals were quurantined, evaluated for health

status, and then randomly assigned to individual cages within the six exposure chambers of the irradiation facility.

Hody weight, Icod consumptaon, and water

consumption data were obtained three times each week during the exposure period and the 5-day preexposure acclimatization period.

The diet was a stand-

ard commercial rodent feed (Wayne Lab-Blox, Allied Mills, Inc., Chicago, Illinois) obtained in pulverized form for food consumption measurements.

Food

and water were provided ad libitum. Irradiation facility.

The irradiation facility has been described completel;

in a previous report. I I It was contained in a typical laboratory room maintained under slight positive air pressure, minimizing outside contamination. Access was restricted to personnel concerned with this research, who wore clean gowns, masks, and gloves.

Room air was circulated at 23 room-volumes

per hour atid filtered by IIEPA and activated alumina filters.

Illumination was

provided 12 hours each day from room and chamber lights, beginning at 6:00 a. m.

Temperature was controlled at 220 + 20 C and continuously recorded;

relative humidity was not controlled, but a continuous record indicated it remained between 25 and 55 percent. There were six identical exposure chambers contained in three racks,

each rack consisting of an upper and a lower chamber.

Each exposure chamber

contained 16 nonmetallic cages described previously, 11 providing food from aI 250-cm 3 glass jar and water from a glass sipper tube and 250-cm 3 glass bottle. Chambers were horizontal, parallel plate capacitors with an upper plate of aluminum screen, a lower plate of aluminum sheet, and an average plate separation of 46.4 L 0.3 cm (S. D.). A signal generating and signal monitoring system provided the sinusoidal 45-11z voltage, which could be independently varied from a nominally zero

- l l

l

field strength value up to 1000 V/m (RMS) for each chamber.

This system con-

tained two signal generators to provide a backup generator If the primary signal generator failed and circuitry to sound an alarm if both systems failed. Voltage and frequency were routinely measured to within t 0. 5 percent. The 45-Hz exposure field and existing ELF fields were measured by the Illinois Institute of Technology (lIT) Research Institute, Chicago, Illinois. Data were obtained for electric and magnetic fields at 15, 45, 60 and 180 liz, and were presented in the report of this facility. 11 The largest ambient field (45Hz fields turned off) was found at 60 Hz.

The average value of the 60-Hz elec-

tric field per cage was obtained for each animal position. Experimental design.

Four experiments (lettered E through H) were per-

formed using the six exposure chambers, each containing 16 animals housed one per cage (Table 1).

Six groups of 16 animals at field strengths of 2, 10,

20, 50 and 100 V/m (RMS) plus a control were used in experiments E, F, and H in an attempt to determine whether a dose versus effect relationship exists. It was assumed in the experimental design that any dose effect relationship that might be found would be monotonic; i. e., if an alteration were observed at one

Table 1.

Experimental Design and Animal Usage Experimental group F G H

Exposure conditions (V/m)

16

16

2

16

16

16

10 20

16 16

16 16

16 16

60

16

16

16

100

16

16

16

96

96

Animals/group Total animals

J9

16, 16,16

16

Controls

16,16,16

96 384

96

field strength, the alteration produced by a higher field strength would be at least as large.

The field strengths chosen cover the range of field strengths

reported by Noval et al. 13 as having produced the growth alteration previously mentioned.

Experiment G was designed to maximize the number of animals

for one field strength by using three chambers as controls and three chambers at a field strength of 20 V/m (RMS) (see Table 2).

This configuration mini-

mizes the chance of making a type-iI error, i. e., the failure to declare a result significant for a fixed level if a real effect is present.

The field strength

of 20 V/rn (RMS) was chosen because this value produced the effects observed by Noval et al.,

13

and it is approximately one hundred times greater than what

the ELF Communications System would generate.

Because of the large number

of animals in each 45-Hz groap of experiment G, a further classification into four 60-iz field strength groups can be made (Table 3).

TIjs classification

was analyzed using two-way analysis of variance to test for 45-Hz and 60-Hz effects, ond to determine if the 45-Hz and 60-Hz fields interact.

Chambers

were permanently numbered 'om one through six for reference, and chamber Table 2.

Electric Field Strengths Applied to Each Chamber for Each Experiment

Exposure chamber

45-Hz field strength (V/m (RMS)) Experiment G F

Position

Number

E

H

100

20

50

2

10

C

20

U

10

2

20

C

4

L

20

50

C

10

5

U

50

20

C

100

L

100

C

20

2

1

U

2

L

3

6

C

-L

16 animals per chamber C denotes control group (no 45-Hz fields applied) U denotes upper chambers L denotes lower chambers 10

'77"=

Table 3.

The Number of Animals in Each Group of the 45-Hz and 60-Hz Two-Way Analysis of Experiment G Frequency (Hz)

Field strength (V/m(RMS))Tol

_____

60

.21-. 42

.53-. 84

1.1-1.7?

24 12 36

8 4 12

12 24 36

2.6-3.8

animals

Field strength (V/ra(RMS)) 0 20 Total animals

4

48

8 12

96

48

bias was minimized by utilizing each chamber as a control or for a given field strength only once (Table 2). After the exposure period, animals were

ithdrawn from the exposure

room one at a time, given an intraperitoneal injection of chloral hydrate at a dose of 36 mg per 100 g of body weight, and placed in a clean cage.

When one

animal from each group had been obtained and all six had become unconscious, the animals were taken to a separate room where euthanasia was completed by heart puncture and exsanguination.

Three milliliters of blood anticoagulated

with EDTA at a final concentration of 8 mM were used for hematological analysis.

The remainder of the sample was allowed to clot for 1 hour at room

temperature, then centrifuged at 600 x g for 15 min to remove the clot, separated into aliquots, and frozen at -60 0OC for later biochemical analysis.

For

experiments G and H, this procedure was modified in three ways to reduce the variability caused by recent food consumption and aggressive behavior exhibited by the six animals when placed together in one cage. withdrawn at 4:00 p.m. on the day before euthanasia; kept in separate cages until euthanasia;

First, food was

second, animals were

and third, plasma was obtained from

blood containing 5 mM potassium ethylenediaminetetraacetate by centrifugation at 600 x g for 15 minutes, 11Y

4

Biochemical analysis.

Total protein and globulin assays were performed

on the AutoAnalyzer (Technicon Corporation, Tarrytown,. New York) by the technique of Sobocinski etal. 17 Glucose, cholesterol, triglycerides, and total lipids were assayed using commerial reagents and standards from Boehringer Mannheirm Corporation' (New York, N. Y.) wvith the following catalog numbers: 15715, 15738, 15989, and 15991, respectively.

Unknowns and quantitative

serum controls (Monitrol I and II, Dade, Division, American Hospital Supply Corporation, Miami, Florida) were assayed simultaneously in a random sequence. Hematology.

Red and white cell counts were obtained using the Coulter

Counter Model B (Coulter Electronics, Inc., Hialeah, Florida).

Hematocrit

values were obtained by reading capillary tubes after centrifugation, and hemoglobin was estimated by the cyanomethemoglobin method employing Drabkin's solution (Hycel, Inc., Houston, Texas). Pathology.

Necropsies were performed on a minimum of four randomly

selected animals from each group.

Specimens from the skin, brain, salivary

gland, lung, heart, stomach, duodenum, cecum, colon, liver, spleen, kidney, urinary bladder, adrenal gland, pancreas, and testes were fixed in 10 percent buffered Formalin for histopathologic examination. were weighed when removed from the body.

Adrenal glands and spleen

Tissue preparation and staining

were performed according to accepted methods as outlined in the Armed Forces Institute of Pathology Manual. 8 Hematoxylin and eosin were routinely used, but selected samples were submitted to Gomori's methenamine silver stain, BrownBrenn tissue gram stain, and the periodic acid-Schiff (PAS) reaction. Statistical analysis.

Histograms of individual observations were u3ed to

determine if nonparametric statistical tests should be employed.

Based on

this evaluation, body weight, food consumption, and water consumption data were analyzed using the t-test or analysis of variance (parametric methods) followed by the Neuman-Keuls test 1 9 (nonparametric method) if significance was observed (p< 0. 05).

j

For biochemical and histological analyses, significance

*1

12

S~.,.

P~7~-

(p< 0. 05) was tested using the one-way analysis and multiple pairwise comparisons of the Kruskal-Wallis tests 4 or the Mann-Whitney test 1 6 (nonparametric methods). RESULTS During these experiments the exposure facility operated without failure. Routine monitoring of the signal-generating systems (at least three times a week) revealed that the chamber voltages remained constant to within +4 and -2 percent, and the 45-Hz frequency did not vary more than k 0. 5 percent. In this research, growth is defined as the net change in body weight of the test animal.

Since the duration of these experiments was short (28 days) and

since all animals used were approximately the same age, the average values of growth per day, food consumption per day, and water consumption per day are valid measures for comparing groups within an experiment.

These values

were calculated by obtaining the total change in body weight or the total weight of food or water consumed for each animal and dividing by 28 (number of days in the exposure period). Analysis of growth, food consumption and water consumption during 45-Hz electric field exposure.

The average body weight and the average growth

per day for each group and each experiment are summarized in Figure 1. The cumulative food and water consumption and the food and water consumption per day for each group and each experiment are summarized in Figures 2 and 3. The daily summary of body weight, growth, and cumulative and daily consumption of food and water are presented in Appendix A.

The strictly monotonic re-

lationship observed for growth, food consumption, and water consumption showed that these experiments were well controlled.

The small depressions observed

in the body-weight change/day curves (e.g., in G experiment on days 10, 18 and 25) always occurred after the cage bedding was changed, and may have resulted from the animal's attempt to mark his new bedding by defecation. 15 Notice that

13

EXPERIMENT E

EXPERIMEN"

(V/mffi

(V/m)

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EXPERIMENT H

GROUP 1 COMPOSITE OF THREE CONTROL

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GROUP 1: 50 oGRCUP 2: 20

GROUP 4: 10 -GROUP 5: 100

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Figure 1. Graphical summary of the average body weighlt and daily growth for each group during exposure

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1

EXPERIMENT F

EXPERIMENT E (mi(V/.r)n)L-

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25

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Figure 2. Graphical summary of food consumption data during exposure

30

EXPERIMENT E

EXPERIMENT F

(V/m. *GROUP 1: C

/GROUP "GROUP 4: ;tQ

GROUP 2: 2 *GROUP 3: 10

GROUP 5: 50 *ROUP 100

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GROUP I: COMPOSITE OF THREE CONTROL GROUPS

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Figure 3.

SEQUENTIAL DAY

SEQUENTIAL DAY

Graphical summary of water consumption data during exposure

16

these readily visible depressions in growth per day are the result of a virtually indistinguishable displacement in the average cumulative body weight curves. In three experiments (E, G and H), no statistically significant differences were found between any experimental or control groups for growth per day, food consumption per day, or water consumption per day when these parameters were compared at the several 45-Hz exposure field strengths used (Table 4). In experiment F, statistical analysis revealed no difference in water consumption per day between any experimental or control groups, although differences

Table 4.

Statistical Summary of the Analysis of Variance of Growth, Food Consumption and Water Consumption Experiment Variable

-

E

F

G

H

a BW/A day

-

**

-

-

h Food/& day a Water/& day

-

**

-

-

-

Not significant (p> 0.05) ** Significant difference (p< 0. 01) -

in growth per day and food consumption per day were observed.

For growth

per day, significant differences (p< 0. 05) were found between the 100 V/m group and the 2, 20. and 50 V/m groups, but not between that group and the control or 10 V/m groups.

For food consumption per day the only difference

(p< 0. 05) was between the 20 V/m, 100 V/m, and 10 V/m groups but not between the control, 2 V/m, or 50 V/m groups.

These differences are not bio-

logically important, as suggested by the findings that (1) the differences observed in experiment F were not observed in experiments E, G and H, i.e., the alterations were not reproducible, and (2) no dose versus effect relationship was apparent from these differences (Figure 4).

Thus, all four experi-

ments indicate that rat growth, food consumption, and water consumption

17

77

MEAN DAILY BODY WEIGHT CHANGE VS FIELD STRENGTH

1

.%

J= It1 SE

MEAN FOOD CONSUMPTION VS FIELD STRENGTH

________________

29

2

0

\

. .......

Q 26 25:

Z

241*

SE

MEAN WATER CONSUMPTION VS FIELD STRENGTH

44z

44-

.~40z

IEXPERIMENT j 32-k

F 0.--.e

z

02

10

so100 20

FIELD SIRENGYN (V/mi

Figure 4.

Summary of the mean growth, food and water consumption data versus 45-Hz field strength

j :I~

Ij

l

Jtt I

were not altered by exposure to 4b-Hz ELF eleutric fields of up to 100 V/m for up to 28 days. Analysis of biochemical and hematological results versus 45-Hz field streigth. In Appendix B are values for each biocheni-ial and hematological parameter measured on all control and experimental animals and a statistical Analysis of these data

summary for each control and experimental group.

(Table 5) showed that no consistent alterations were found in any of the parameters measured in any of the four experiments performed. vidual parameteri

Table 5.

Findings for indi-

are summarized below.

Statistical Summazy of the Kruskal-Wallis Analysis of the Blood Biochemistry and Hematology Data

Variable

Experiment ....... F G H E

G.,OB

*

'

GLU

**

TL

*

-

*

-

*

(HOL

-

-

-

-

TRIG

-

-

-

-

RBC

**

**

-

-

.

.

WBC POLY

. ..

LYHS

.

HCT

**

HGB

* ..

.

i -

-

.

MONO

within normal limits

EOS

within normal limits

Not significant (p> 0.05) Significant difference (p< 0.05) *Significant difference (p< 0.01)

19

A

Total protein: In experiments E, F and G, no groups were statistically different from the contrr" or from each other. the control and 100 '/m

In experiment H, only

group .Jlffered. No trend or dose relationship was ob-

served (Figure 5). Globulin: In experiments E, F and G, no groups were statistically different from the control or from each other. In experiment H, only the control and 100 V/m group differed.

No trend or dose relationship was observed

(Figure 5). Glucose: In experiment G, where the statistical test should be most sensitive, the control and irradiated groups were not statistically different.

In experiment E, only the control and 2 V/m groups differed.

In experi-

ment F, the control group, 20 V/m group, and 100 V/m group were statistically different; and in experiment H, only the control and 100 V/m groups were statistically different.

No consistent pattern of statistical difference or dose re-

lationship was observed throughout these experiments (Figure 5).

In addition,

some of the apparently large variability observed in experiments E and F may be due to the euthanasia procedure used (see below). Total lipids: In experiments F, G and H, no groups were statistically different from the cont:ol or each other. group and the 100 V/rn group differed.

In experiment E, only the 2 V/m

No trend or dose relationship was ob-

served (Figure 6). Cholesterol: In all experiments (E, F, G and H), no groups were statistically different from the control or from each other, and no trend or dose relationship was observed (Figure 6). Trigly, erides: In all experiments (E, F, G and H), no groups were statistically different from the control or each other, and no trend or dose relationsldp was observed (Figure 6). Red blood cells:

Ir, experiments G and H, no groups were statisti-

cally different from the control or from each other.

In experiment E, differ-

ences were found between the control and the 10, 50 and 100 V/m groups, but

20

84

-'-

'

"-r-

MEDIAN TOTAL PROTEIN VS FIELD STRENGTH

S......

- ,-.,,,-.,-,....... ....... .....,, ,........ ...r ,T.'"..'..... 1 40i. ............. ............... ,.+,.,,...........• +,,

0 MEDIAN GLOBULIN VS FIELD STRENGTH

z

i2 -

..... *%

........ ,." .....,..,."...............

,.. 2,03

MEDIAN GLUCOSE

:

FIELD STRENGTH

w2 30-VS

1''''"..

260-I

" ......... ...... ...5.2.. .... .. .. ...

.. ....... .... . . ........ I'

..............

ISO*'

140

2

10

Figure 5.

L

EXPERIMENT E

.i H ,i....... 0

+++ + +

i' +

A ..... ........ ....................

o220--t . %, "p . ,' +lllL,

.. + .....,.....+..

so

20

Summary of median. total protein, globulin and glucose data versus 45-Hz field strength 2

100

I I

I

IJ

I

MEDIAN TOTAL LIPIDS VS FIELD STRENGTH

520-

S/

0

0

440-

XERMN

e.-".MEDIAN TRIGLCERIDE

~o

3

~

.M.............

r

o 10- 0

0

2

MEDIAN CHOLLYETEEPEN VS FIELD sSTRENGTH

I I

ls

SFIELD STRENGTH /n

v210

VS FIELD STRENGTHm *

0w5011,

Figure 6.

E ,

n

i

,

'

Summary of median total lipid, cholesterol and

triglyceride data versus 45-H1z fteld strength 22

not between the 2 and 20 V/ni groups.

Additionally, statistically significant

differences were found between the 2 V/m group and the 10, 20, 50 and 100 V/rn groups.

In experiment F, statistically significant differences were found

between the control and the 2, 10 and 50 V/m groups.

However, since these

differences were not reproduced in experiments G and 11, the pattern of statistical dliference in experiments E and F may be related to the euthanasia procedure (see below).

Considering all experiments, no trend or dose relationship

was observed (Figure 7). White blood cells& In all experiments (E, F, G and H), no groups were statistically different from the control or from each other, and no trend or dose relationship was observed (Figure 7). Segmented neutrophils (POLY): In all experiments (E, F, G and H), no groups were statistically different from the control or from each other, and no trend or dose relationship was observed (Figure 7). Lymphocytes:

In all experiments (E, F, G and H), no groups were

statistically different from the control or from each other, and no trend or dose relationship was observed (Figure 8). Hematocrit: In experiments F, G and H, no groups were statistically different from the control or from each other.

In experiment E, only the

2 V/m and 50 V/m groups were different from each other.

No trend or dose re-

lationship was observed (Figure 8). Hemoglobin:

In experiments F, G and H, no groups were statisti-

cally different from the control or from each other.

In experiment E, only the

2 V/m and 50 V/m groups were different from each other.

No trend or dose

relationship was observed (Figure 8). In experiments G and H, an improved euthanasia procedure was used in which plasma was obtained from control and irradiated animals that had fasted for approximately equal periods of time, and that were not excited since the anim.as had been caged separately until euthanasia to prevent stress.

As a

result of these improved procedures, marked reductions were noted in the group

23

MEDIAN RED BLOOD CELL COUNT VS FIELD STRENGT1H

7.0.A.

6.6

~

*................

S6.2-

5.4 7600

1

MEDIAN WHITE BLOOD CELLS VS FIELD STRENGTH

,.

~6000%.

.~

520

.. do*t

144

g

do.P*%.

4400-

9

.r .MEDIAN SEGMENTED NEUTROPHILS ........... VS FIELD STRENGTH

S.0-

U

*..........

EXPERIMENT

&

E

*

GU--

5010

20s

100

FIELD STRENGTH IV/1m)

Figure 7. Summary of median red blood cell, white blood cell and segmented neutrophil data versus 45-Hz field strength

24

-94

~

MEDIAN LYMPHOCYTES VS FIELD STRENGTH

a'

........ ....

W

...

.................

V *

a.

88

MEDIAN HEMATOCRIT VS FIELD STRENGTH

542_______________ 48-

J

40

MEDIAN HEMOGLOBIN VS FIELD STRENGTH

EXPERIMENT E

F *-

16.1-

*-

..... *...... ................... ..................... ~~~.

-h*..............................................

14.0-

02

10

20

s0 FIELD STRENGTH (V/rn]

100

maori

n n Figure 8. Summary of median lymphocyte, eaoci hemoglobin data versus 45-11z field strength

for glucose, to group variability (Figures 5-8) and the within group variability hematocrit, and hemototal lipids, cholesterol, triglyceride, red blood cell, that there were globin values. It can be stated with a high degree of confidence variables used no differences in the plasma or serum values of the biochemical 25

in this study.

After completion of serum triglyceride assays for experiments

E and F, it was observed that free glycerol was present in the commercial standards used.

To correct the triglyceride values in experiments E and F, the

values reported in Appendix B and the group medians of Figure 6 must be multiplied by 0.66. analysis.

This constant factor does not alter the results from statistical

The accuracy of this correction factor and of the measurements in

experiments G and H was verified by using two independent standards (Precilip, Boehringer-Mannheim Corp., New York, N. Y., and Triolein, Sigma Chemical Company, St. Louis, Missouri) as well as the known molar extinction coefficient of the reduced form of nicotinamide-adenine-dinucleotide (NADH). Pathology versus 45-Hz field strength. were coded and analyzed in a blind manner.

All animals used in this analysis No gross lesions or differences in

adrenal and spleen weights were observed at necropsy, and no significant microscopic changes were present in any of the tissues. Two-way ANOVA for 45- and 60-Hz effects.

Detailed analysis of field

map data after completion of experiments E, F, G, and H suggested that the level of the contaminating 60-Iiz electric field had been underestimated in the initial field map. 11 In an attempt to determine the effect of this field on the results of the 45-Hz analysis, the data from experiment G were separated into four groups according to the average 60-liz field strength per cage for two-way analysis of variance as shown in Table 3.

Since the experiment was not initially

designed to examine possible 60-Hz effects, the two-way classification did not yield groups of equal size.

In addition, random animal positioning at the begin-

ning of the experiment happened to allow the group exposed to the highest 60-Hz field strength to start with an average initial body weight slightly heavier than the other groups. As a result, care must be taken when interpreting results of the two-way analysis of variance for growth per day, food consumption per day, and water consumption per day shown in Table 6.

Despite these restrictions,

certain statistically valid comparisons can be made.

There were no statisti-

cally significant differences (p< 0. 05) between the control and 20 V/m groups

26

Table 6. Statistical Summary of the Two-Way Analysis of Variance on 45-Hz and 60-Hz Field Strengths of Experiment G V

Factor one

bl

Factor two

tet

8f0 HzI

45z

ABW/c day--

Food/A day77 AWater/& day

-*

significant (p> 0. 05) Significant (p< 0. 05) Significant (p< 0.01)

-Not

* **

at 45 Hz for growth per day, food consumption per day, or water consumption per day.

Neither was the interaction between the 45-Hz and 60-Hz fields found

to be significant for growth per day, food consumption per day, or water consumption per day.

Further, there was no alteration of growth which is attrib-

utable to the 60-Hz field.

The statistical differences observed in the data on

food consumption per day and water consumption per day occurred only between the highest and lowest field strength groups.

Thus, the statistically significant

results obtained - ,.re most likely not biologically important because (1) the animals in the group exposed to the highest field strength started with an average initial body weight slightly heavier than the other groups, and (2) the food and water consumption correlated strongly with initial body weight.

This conclu-

sion is reinforced by the expectation that a biologically important alteration in -food and water consumption would be reflected in a growth alteration.

To com-

plete the 60-Hz analysis, the biochemical and hematological data were tested for significance; results are presented in Table 7.

No statistically significant

diffcrences were observed for glucose, total lipids, cholesterol, triglycerides, white blood cells, segmented neutrophils (POLY), lymphocytes, hematocrit, or hemoglobin.

Significance occurred in total protein and globulin data only be-

tween the highest and next highest field strength groups, and in the red blood cell data only between the lowest and next lowest field strength groups.

27 ,p-."

Table 7.

Statistical Summary of the Kruskal-Wallis Analysis on 60-Hz Field Strengths of Experiment G Variable

Sigidflcance

TP

*

GLOB

*

GLU TL CHOL TRIG RBC

*

WBC -

POLY LYHS HCT

-

HGB

-

Not significant (p> 0. 05) Significant difference (p< 0.05) •* Significant lfference (p< 0. 01) -

*

Because no.significant differences were observed for any variable between the highest and lowest field strength groups or between the two field strength groups with 36 animals each, no biological significance was attributed to these differences.

DISCUSSION AND CONCLUSIONS Theoretically, ELF radiation can be separated into terrestrial and manmade radiation.

The terrestrial field strength in the exposure facility was not

measured, but is considered to be negligible compared to the experimental field strengths.

Man-made fields at frequencies from 25 to 60 Hz are present any-

where electric power is used. 21 For example, under high-voltage power lines,

28

electric field strengths of thousands of volts per meter are found; I and in the laboratory, every operating electrical appliance is an ELF irradiator;

The

man-made 45-Hz fields in our exposure facility were documented by the lIT Research Institute.

They also documented a contaminating 60-Hz field which

was present in all chambers, i.e., both irradiated and control. No biologically important differences were found for any of the variables studied.

No statistically significant differences were observed in any experi-

ment for the following variables: water consumption, serum or plasma cholesterol, serum or plasma triglycerides, white blood count, segmented neutrophils, lymphocytes, adrenal weights and spleen weights.

No significant differ-

ences were observed in any of the histopathological analyses.

For the follow-

ing variables, the only observed significant statistical differences occurred between irradiated groups: growth per day, food consumption, serum or plasma total lipids and hemoglobin.

Significant differences between the control and

irradiated groups occurred only for total protein, globulin, red blood cells, and hematocrit.

Further, no dose-relationship was observed between the exposure

field strength and any of the variables studied.

The most unequivocal results

are from experiment G, in which 48 animals were used in both control and 2C V/m exposed groups.

No significant differences were observed for any of

the variables measured in this experiment. The data from experiment G were subjected to two-way analysis of variarce to test the possibility that the contaminating 60-Hz fields in the exposure facility affected the results of the 45-Hz analysis.

No significant differences (p< 0.05) were found in growth per day for the 45-Hz or 60-Hz fields; further,

no significant interaction was observed for growth per day, food consumption, or water consumption.

These findings together with the fact that the two 60-Hz

groups (of 36 animals each) were not significantly different from each other led to the conclusion that the 60-Hz fields in this experiment did not produce a biologically important alteration.

From this analysis it is surmised that the data

from the 45-Hz fields also were not affected. 29

A

A -ilot experiment gives further indication that ambient fields did not perturb the negative finding of the 45-Hz analysis. '9

In that experiment the average

growth rate for 96 animals was found to be 7. 8 grams per day per animal. These animals were fed and handled using procedures identical to those in experiments E, F, G, and U; however, they were housed in rat cages of standard No. 2 mesh and sheer -tainless steel. At our request, the lIT Research Institute determined that these cages provide better than -40 dB of electric field shielding at 60 Hz (Appendix B of reference 11).

Thus, these animals (in the pilot ex-

periment) were grown for 28 days in an ambient 60-Hz electric field which was less than 0. 010 V/m (RMS).

If the ambient electric fields present in experi-

ments E, F, G, and H had produced a growth reduction in all experimental groups of at least 20 percent (the smallest growth reduction reported by Noval et al. 13), then these shielded animals would have growth rates greater than 9. 2 grams per day per animal.

Instead, the growth rate of the animals in this pilot

experiment (7. 8 grams per day per animal) is well within growth rates measured for the six groups of animals in experiment F (Figure 4) which were the same age as in the pilot study. The findings of this research are consistent with the work of Knickerbocker et al. 6 and Krueger and Reed. 7 Knickerbocker et al. exposed male mice to a vertical electric field of 157,000 V/m at 60 Hz for 10-1/2 months (6-1/2 hours each day), and analyzed for differences in growth, reproduction, gross pathology, and histopathology.

Although they noted that the unexposed male progenies of

the exposed animals did not grow to be as heavy as the male progenies of the control animals, no other statistical or biological differences were observed. Krueger and Reed exposed female mice tc horizontal electric fields of 100 V/m at both 45 Hz and 75 Hz.

No statistical differences were observed between ex-

posed and control animals in rate of growth, serotonin levels of blood and brain, or susceptibility to challenge by influenza virus. After exposing 384 young, male Sprague-Dawley rats for 28 days to 45-Hz vertical electrical field strengths of 2, 10, 20, 50 and 100 V/m (RMS), no 30

biologically significant differences were observed for any of the measured variablk

,

Further, no dose relationship was found for any of these variables ver-

sus the applied 45-Hz field strengths.

31

nil

REFERENCES 1. Bracken, T. D. Field measurements and calculations of electrostatic effects of overhead transmission lines. IEEE Trans. Power Appar. Syst. PAS-95:494-504, 1976. 2.

Gavalas-Medici, R. and Day-Magdaleno, S. R. Extremely low frequency weak electric fields affect schedule-controlled behavior of monkeys. Nature 261:256-259, 1976.

3.

Goodman, E. M., Greenebaum, B. and Marron, M. T. Effects of extremely low frequency el.;ctromagnetic fields on Physarum polyc. halum. Radiat. Res. 66:531-540, 1976.

4.

Hollander, M. and Wolfe, D. A. Nonparametric Statistical Methods, Ist ed., pp. 114-129. New York, N. Y., John Wiley and Sons, 1973.

5.

Kalmijn, A. J. The electric sense of sharks and rays. 55:371-383, 1971.

6.

Knickerbocker, G. G., Kouwenhoven, W. B. and Barns, H. C. of mice to a strong AC electric field - an experimental study. Trans. Power Appar. Syst. PAS-86(4):26-33, 1967.

J. Exp. Biol.

Exposure IEEE

7. Krueger, A. P. and Reed, E. J. A study of the biological effects of certain ELF electromagnetic fields. Int. J. Biometeorol. 49:194-201, 1975. 8.

Luna, L. G., editor. Manual of Histologic Staining Methods of the Armed Forces Institute of Pathology, 3rd ed. New York, N. Y., McGraw-Hill Book Co., Inc., 1967.

9.

Marino, A. A., Becker, R. 0. and Ullrich, B. The effect of continuous exposure to low frequency electric fields on three generations of mice: a pilot study. Experientia 32:565-566, 1976.

10.

Marino, A. A., Berger, T. J,, Austin, B. P. and Becker, R. 0. Evaluation of electrochemical information transfer system. I. Effect of electric fields on living organisms. Electrochem. Technol. 123:11991200, 1976.

11.

Mathewson, N. S., Oliva, S. A., Oosta, G. M. and Blasco, A. P. Extremely low frequency (ELF) vertical electric field exposure of rats: irradiation facility. Bethesda, Maryland, Armed Forces Radiobiology Research Institute Technical Note TN77-2, 1977.

32

12.

Mathewson, N. S., Oosta, G. M., Oliva, S. A., Levin, S. 0. and Blasco, A. P. Effects of 45 Hz electric field exposure on rats. In: Biulogical Effects of Electromagnetic Waves, Johnson, C. C. and Shore, M. L., editors, Vol. 1, pp. 460-471. Rockville, Maryland, HEW Publication (FDA) 77-8010, December 1976.

13.

Noval, J. J., Sohler, A., Reisberg, R., Coyne, H., Straub, K. D. and McKinney, H. Extremely low frequency electric field induced changes in rate of growth and brain and liver enzymes of rats. In: Compilation of Navy Sponsored ELF Biomedical and Ecological Research Reports, Vol. III. Bethesda, Maryland, NMRDC Report EMRPO-1, Department of the Navy, National Naval Medical Center, January 1977.

14.

Polk, C. Sources, propagation, amplitude, and temporal variation of extremely low frequency (0-100 Hz) electromagnetic fields, Chapter II. In: Biologic and Clinical Effects of Low-Frequency Magnetic and Electric Fields, Llaurado, Sances, and Battocletti, editors. Springfield, Illinois, C. C. Thomas, 1974.

15.

Russell, P. A. Open-field defecation in rats: relationships with body weight and basal defecation level. Br. J. Psychol. 64:109-114, 1973.

16.

Siegel, S. Nonparametric Statistics for the Behavioral Sciences, 1st ed., pp. 116-127. New York, N. Y., McGraw-Hill Book Co., Inc., 1956.

17.

Sobocinski, P. Z,, Canterbury, W. J. and Hartley, K. M. Automated simultaneous determination of serum total protein and globulin. Bethesda, Maryland, Armed Forces Radiobiology Research Institute Technical Note TN73-11, 1973.

18.

Southern, W. E. Orientation of gull chicks exposed to Project Sanguine's electromagnetic field. Science 189:143-145, 1975.

19.

Steel, B. G. D. and Torrie, J. H. Principles and Procedures of Statistics, pp. 110-111. New York, N. Y., McGraw-Hill Book Company, Inc., 1960.

20.

Walcott, C. and Green, R. P. Orientation of homing pigeons altered by a change In the direction of an applied magnetic field. Science 184:180182, 1974.

21.

Westmann, H. P., editor. Reference Data for Radioengineers, 4th ed. pp. 929-931. New York, N. Y., International Telephone and Telegraph Corporation, 1956.

33

_

_

_

.

.

.

..

.

.-

,,..

APPENDIX A Daily Summary of Body Weight, Growth, and Cumulative and Daily Consumptlon of Food and Water The following seven tables (A-i through A-7) summarize the actual raw growth and consumption data for each experimental group of the four experiments (E, F, G, and H).

In addition, the dates were obtained trd a summary

of the temperature and relative humidity on those dates is provided.

The

field strength and chamber position (either upper or lower) for any group can be obtained from Table 2 of the text. The headings for each column are defined (proceeding from left to right) as: DATE:

date these data were obtained; expressed as day/month/last digit of the year

DAY:

the number of days these animals have been exposed

TEMP:

the average - the range of room temperature ( F), taken over the interval from the previous data day to this day

HUMIDITY:

the average + the range of relative humidity (% RH), taken over the interval from the previous data day to this day the number of animals in each group, either 16 or

N:

48; or when an animal's food or water consumption could not be accurately measured due to an accident; (e. g., bottle was spilled), it then becomes the smallest number of animals used for any one of the calculations for this date BODY WEIGHT: XBAR

the average mass (g) per animal for this group

SD

the standard deviation of XBAR (g)

34

CHG. BODY WT: XBAR

average change In mass per day (g/day) per animal for this group, taken over the interval from the previous data day to this day

SD

the standard deviation of XBAR (g/day)

FOOD CONSUMED AND WATER CONSUMED: XBAR

average food (water) consumed per day (g/day) per animal for this group, taken over the interval from the previous data day to this day

SD

standard deviation of XBAR (g/day)

TOTAL TO DATE

average of cumulative food (water) consumed (g) per animal from day zero to this day

SD

standard deviation TOTAL TO DATE (g)

ASTERISK (*)

this symbol is used to note that the data from all animals could not be used; where N = 16 for experiments E, F, and H, or N = 48 for experiment G.

Further information

is provided in the next paragraph It was necessary to delete three animals from experiment F because they accidently went without water over a weekend. This accident occurred early in this experiment, and animals resumed normal drinking, eating, and growth values; therefore, these animals were not deleted from the biochemical and hematological analyses.

Because one animal was deleted from group two (Table A-3)

and two animals were deleted from group six (Table A-4), asterisks indicate that fewer than the usual number of animals were used for each day.

35

.. . ... .......... ... ..

'I II

i

i

TABLE A-i GRcOUP I

DAT!1 DAY TtpC

HUMIDITY

N

BODY.WRIGHT 460 Sb

CMIG.BODYWT. 1103 so

)M3AN

FOOD CONSUMED TOTAL so To DATE

SO

I.ATtl CONSUMED TOITAL SD TO DATE

X303

SD

7, 44

a

64 1- 6 16 12k.-3

12.20

6.00

6.60

6060

6,6.00

6le .66

6.66

69.0

9,4,3

& pz.6-0 204 6- 1 16 l3r.66

13.17

7.03

1.16

26.40

1.131

526

3.03

42.43

4.953

Ito,443

4 Pt.*-* 34* 1- l? i6 533.6

14.60

?-11A

1.60

Z6.3@

I.as

1@5'5?

"r0

44.51

1,42

IMP0

16.11

16m6

6.6

1.63

P6.19

3.79

150.14

q.15

43.15

6.14

303.43

31.44

9.29

1.13

Z16.75

1.s4

r4'.s4

16,3

44.16

9.67

m-1.71

45.%4

04-a

14 4' 12 70-I II'

30" 1- a to 660.99

760646

II4448 7

14 1 11604.1 4110.4

6l.60

I 04,4"

6.66

46- a is 311.44 *-

10.54

7.34

3.56

ea.@%

1,69

303.60

14.67

46,1

6.16

4.53

a to1332.03

13.11

6.07

1.67

25.60

6.50

363.43

1%,sr

4,3.14

6.63

60.4

6 it 346.99

66.43

5.75

3.21

36.9?z

2,?3

4"4. 9

24.20

4,1, Os

?.14

6914.64 %d1.02

25e 4.. IS 70665r4 0- I Itj 355.61

j$ 64.9I

.4q

Z5. c k

43* - 0

.03

703.31

26, 445 61 71.0-0 43* 0- 3 16 369.?'

26.63

4.C,5

,32 'w

51 1-0 1;11. 1

30, 4

23p10-e n 53+ 6- 1 16 379.63

65.14

4.7l13.07

2'

14 ?9 1+0-0 54408- 1 14 385,1%

66.66

2.56V

ll,

4

14 pk40-0 464

230' 44 14 74-6 Q+46

,

3.43

7

3.43

6?i3

3,1-5 517.114 .V0

33.044

'..' I5.922

1.43

40N. 4.1 3..5$

5 4

1 -1 *27

379

43.546

-. 34.07

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412.

4,9 4

003.t2

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495 6.l

1IW..

I13. 3 '

10 u 104 193.54

GROUP a

DATR DAY Tily

N

HUMIDITY

BODY WRIGHT 81603 sD

6 661to7I.44

6MC. OOY WT, 8143 98

nap

7' 445

6

14.00

0.66

so 4"

a ?24-4 14. a- I It 633.59bl 14.56

0,64

1.62

20.32

It,' 44

4 11+-11344 1-6aI

6.66

3.31

14.' 44

F pf0-1 34+ 1- a It z27353

16 .'s

s 024-6 464 1- 1B66

6+2-0

64*

264P.24 14.04 21.1's

.0 6

FOOD CONSUPTD TOTAL SD TO DATE7 SD

60

0..6.0

6.0

0.00

5s.64

7.56

42.a3

6.13

-%-56

43.06

4.6?

CONSUMED TOTAL TO DAT!E SD

6.66

1.60

100.56

1'6.56

16.62

66.33

6.06

193.94

22.44

41.'31 11.59

?S5.06

30.74

?,5

61 53.61

9.60

19.11

0.0

3.26

69.?4

7.63

1.6?

Z6.3.1 9.44

all' 4.9 14 ?146-0 40+ a- a 16 325S.43 10.63

66.59

43361

6.25

362.20

305.65% ?4.06

42.06

S.48

466.36* 65.26

?.37

1.04

31.50

2.67

400,14

24.46

43.?7

5.64

1@.%@

P.05

1.9O

10.63?

2.05

461.48

28,44

44.53

C.02

Z51' 415 Ia e+2-8 57+ 61- I 16 353.64

10.0?

4.31 4,4 ".$)

341 10

3

22 .71

as*.01

42.03

5.,34

7140-l 43- a- 3 16 370.27

15!. 22

5. 68

1.81

32.14

1

419. 'l 12

'!L .37

45.14

0.3-1

23 71*0-S 53+ 6- I IS 3711.1aZ0.61

3.fl

1.111 31.1?

681.46

3',7. G

45.,3

16,018

3.1

1.55

741.41

30.451

41.87

to 7640-o 4t+ G- 6 Is 341.50

?se

.21

a.,4

5Z. 5 7140-S 54+ 6- I It 364.93

0.669

.66 12.2?

25.!56

18.56

a3.,4n

6 64.45

%69 5.06

s 14 s

R1 6-

sD

9.59

606.

30r.34

160044 It ?1+1- 64

WIT!3 81307

.

2,.57

5.!'0 R.54

99.16

00e.58

6eq.1065.7 1

002

"7.54% 120.180 .9

135,74

(4.14 1007.34 153,9

GROUP 3

DATE DAY T". MUIIDINY

r., "-

a

640-6

0' 44" 6 7246lie 445 10104"

N

4603l

04 6- 0 to 611.7

BOYWRGT00.1OY'T IOYCMI.NOY 1617 T.*TOTAL so 1104 SO XtAl

FO CONSUMED SD' TA) DA0TE sD

5.67

6.66

ea* 11- 1It 2626.6s 17.39

6.5?

6.6-4

24.81

4.0k)

3.5'3

,c 0.06

6.66 00

WATER 813

sD

8.0

ol06

0. 0

45.h63

811?

36.33

9.69

$.$1

CONSUMED TOTAL TO DATE SD

6.06

6.66 6.6 79.07

6 15.36

4 72*0-6 34. 1- 6 It 244.11

12.14

0.61

Z5.64

1.5,2 too.%$

43.35

3.63

113.36

10.36

? ?*46-1 30+ 1-

K 6704M

16.71

0.514

1.69

27.76

1.49

104.86

ll.?

46.03

3,66

?63.49

Z5.36

1It1 20461

42,41

5 72+1-61Q+4 2-

M334

T.13

t,77

20.93

3.34

241.13

16.64

3%55

12,.61

36.54

16-4

It ?1+1-0 41-+6- a Is 3611.31 14.95

?. n

1.62, 67.64

1.64

6546.66 16.23

41,45

4,66

651.44

.9.3

21.' 4

14 ?1+0-0 46469- 2 It 32a.71

16.?6

7.13

1.16

a9.1?

1.94

304.34

62.30

41.651

3,7?

577.11

57.69

?I.,

16720-a 46*4is

16.65

4.61l

1,43

T,1

Z?.3 .49

426

63.15

lie' 44S

a Is "9. 13

ne 445 le rkw.-o 5r+.0- I IS 3.4.3

17.15

'00 41-2E1 71+11-0 .30- 3 16 35Z.04

?2.65

3. 49

:;V. 4'5 23 71*6-0 33 0- 1 16 3 e,6.3 5

20.63

4.19

1.0

5 !19n9 7.-0 54- 0- 1 16 3R73R.7 70.80~f 3.,"N

4431.10

2n.-.!

.10.08?

.4?

20.1 El~

5.3110.7

25.4I0L

413.35

.5'1

0.10

4.135 945 .03

41.31,

00.0,

3,23 .7 1.04

24.7

2.7*

G44. 4 ' 44.58

'le. 04

lo41

"n1,31

36

IT L .

Jt

4C.-5

0,1,3 34

4 1.07

4.20

74',,c-, V,.5.33 7'k 901

i' 7 54 11.8

103-1.61945.05

TABLE A-2 IMPIMSI4T t GICJP

4FODCMM&WAE

DATE bay Itto

6

To Ales

H U1811VV

16Y'bE114T KIRE to

1

too, .1

MI V TOTAL TO DATE

so

141

So

68L4

e.s6

6.75 ,1 6.31

6c84 47 c.ps

25.41 13 25391

4.8% 41-a1 45 31

56.!4 6.113 36.63 191141% 1.4.05 41.36 117 .4 46

Ma.l 6.66 .3

6.-6 414 1- 1 to 291,64 f

1646

?764

1,05

17.66

2.T0

R40.13

P2.31

41.46

3.7f30666.1

53.sr

ato 303.60 It

I.66

6.12

1,21

16.11

1d

o"7.3s

114.31

44,61

6.16

47e. S4

63,00

10.11o

GIGS

1.2648.7s

3.1

316411t 32.76

.13J0

694

6363

6

11..14

6,46

I.151

Z57

3N.Z% .10

443.04

31, 14

.4.,05

"l.3

(0

2Z.72

6.74

1.7?

2?". Iq

"-"

5n". 1"

4'. Om

03.Q.

-1,.M

77,7.84

M4.4 384

4,81

1.58

44.Sk

5I. Zt 57A

4q.7,"

41,37

8

24.56

1,73

1.3

4.'

s.37

24.22

4.13

2.20

Is 724-0 4z+ S- a 0; 318.75s

25.' 4e% te

-6r+- oT - 1 1183

. 21

M8' 4.5 a1 ?1+00 43+ 0- 3 It 4.15 23 r190-0 53+ a- 1 16 11.4.33 71 -0-M 3440-

Z.''4" ^ V

0.se

.4

6.86

1s a£1.16

ato' 4^' 14 14-6 40+ a- a to 323.6

10.1

61

113117 1.6 16.41

64 2. a

? 1+1-4 42* .

230' "'

TOTAL To DATE

)Mfkp

11.34

610-0

it' f1+636-2 11646- I to 236.46 ii ' a4.6 14 7e-I 6. p4- a to 10.41 ISO' 4.1

CHG, BODY W~T. 486~ so

1 16 1384,5

I'.S -el.3

6.66

6.6o

6.1 .66 6

C9.19 II.1 11,1tl

2.43

7'.39

",10.31A0)

44, 14

6.06

6.6

?S.N6 M640 1.0 96.33 666 3

I'S1 14.!Y7 li-. 37,

451.7 3'S,r414.7

4,.12 1 Mi73.S3 140. 4

GROUP 6 FWD1

bat

To, 4^S SO,

BODY WEIGHT 14 14686 so

DAY TEMP HUM41IITY

"

6

0+6-6

C140. BODY*WT, Xb68 SD

CONSUMED1 TOMA TO DATE

)K686

5D1

6+ 1- 6 it 214.40

12,5a

6.@6

6.66

6.05

600B

672+5-6 29+46- 1 14 ?31.05 r

Me?0

6.2ai

11

.15.57

1.0

6su. 33

14.1t

6.14

1.73

26.F62

2.13

103,1

OATIP ID

14686

oleo1 6.66 51.13

O.01

SD

C04SUMED TOTAL TO DA4TE SD

0.13

0.66

4.52

64.16

3.75

.4,1

7.GLI

42,13

131!1'

6.66 S.43

11lo, 4.5

4 ra46-6 344 1- a

14' 4AS

t

304 1- 3 14 073.4%

17.36

4.1 1

i.18

27.32

3,7

165,14

15.411

43.4?

5.24

M1.26

16.04'S

6 ?24-0 42+ 1- 1 1t M1,45

11.67

6.46

1.64

26.M

2.

41,54

0.30

41.60

4.41

3%0 . I

19.13

r.5.1 2.40

20,a55

'11

It 325,16

r2.4?

s.at

1.7?

26.24

6 1 N3.3I

4% 4.ke

6.73

1.43

e64."'

61.34

1.,

24

Ito, 4eS 11 711-Q

4,4%- a IS 305.46

ale 4^S 14 ?1+0-0 46.6aale 4^S 16l720-6 42+4L3' 4'S16 to I"Ae 4'21

2

44-z0 574 13- 1 11 Ml^1' 714-

3 15

4340 A*-

411!5s toI3Il

4.3

.4.,40

4,5%

41-5.31 $6. ?.1

3.02

36.!.?

30.5

.l..

9,5

613.1"

%3.0 4,4

445.4?

34.3

4.19

7041

l."

5044.k-4 373

2 .: 3a9 30.. cl~

71

1

.1

1

1 1Ir 373,27

26. VI

3, 44

1.. 24

.3

17,74

34K

ze 511 5n71+6-0 544 a- 1 16 38k.44

27.1m

-. 7'

1.4-1

.69

3. C

111

340' 4'! Z3 ?1-0-a 33+ 0-

30.4

33,.4

7t.,

Z4.4 44

108.es

.4 4

1.3 -. Q~[..

71

7.3

~7l4~,.

,

4..

173,

ll[~37'lI. .447

5 4.4

7,114 7

4

11c.'

114.7I

GROUP7 6 pool,

DATE DAY TEM

HIIY I1'6

7.e ".

6

Se des

2 746-0 as+46-

644-6

M1

B0DY 6MIGHT 14606 so

G+ 0- 6 16 11.4 1 16 232.64

C1447 BODY WT. so Sb 14686

14

D

0.08

0.00

16.76

0.30

1.11

26.416

1.S9

16.4 M0

,1.0

401114140 TOTAL TO I-ATE

0.86

11.68B

6ATER 1436

6D.

,0 1D 0

.60

3.1$P 44.45

"4.31

%D

4684130 TO7A L TO ZATE

SD

0.08

15.00

4.66

8o.8

6a .21

S.8 " 5,50

8.327.43

40.853 34,17

8.60

li.'415

472e4-0 344

41.6,4

16.46

a.6

1.I.18,27.23

2.12

t4, 4e5

7 pa41-I 36+ I - 2 16 275. 41

26.45

8,71

6.66

26.44

3.130

IV' 44

6a+6-0 42431- 1 It6211.56

226.6'

6.7r

1.66

21,.66

3,77

350.71S 31.%6

4r.11

1.73

42 1.6

44.54

16' 4

11 ? 1+1-6l4Q 16- 2 14 305,5?

23,61$

7.61

1.10

Z8.50

1.94

307.7rl

2AI

4r,38

5.11

$16.01

57.4's

ate 4'

14 7140-6 464 a- 3 Is 3"--S

Me.6

?.,)1

1.61

369

3.36

3".4.6

33.2!'

41,04

7. ?6

663.07?

61.'445 16 72+q-8 4a4 $- 2 Is 34.q4

27.33

6.118

1.51

11.55

!,.%1

'w..34

33,53

4k,.44 .1c, 6.(

"7.1

43-. '

05 4 1 ?3V 4.1521

1- 3 t

76+2-0 Sr740- 1 14 352,54

5. I;.3

2.94

3;07,--

4.44

3

4. 4.30 ,

1172

3Z-711

27

1 16 374.61

34.0)3

4.17

I 1.

31.l:

544 0- 1 16 384.46

35. 44

4.04

1.,45

V8.7

1-0-0 434 0- 3 16 3 C.1.47

36' 443 23 ?1+0-0 '534 0V. 54'1 VS

0-

11)7.3i& ?.$j M37,4

C7~0

. 15 43.7 4.31

1q.1.3 46 .85

" I

1111ki.44

'44. 41

-.

48,C2

70.B3

V'. 0' 1 .18.4

4%.U,

4,-

.MAN"

11.14

5,17

,r31 3'0 10.35 ').43

%G. 3. S c.3 11.77 14.'

I15915Z 161.4%) 11 8G,041657.

TABLE A-3

I

EXPERIMENT F GDOUr I

HUMIDITY

CHG. BODY WT. XBAR sD

FOODCOGSi1ITD TOTAL TO DATE SD

(.as 8

.8 .8

8.66

6.68

3.84

24.,-8

2.33

49.97

5.87

79.99

6.39

29.44

2.85

188.86

£6.68

2.68

147.13

12.30

29.25

2.8

196.62

1.33

2.29

195.09

16.37

32.46

2.82

261.54

23.63

24.59

2.2?

244.8?

26.52

39.26

4.43

346.11

31.83

25.76

2.54

32.16

27.25

37.86

4.29

451.29

43.53

1.62

25.79

2.13

373.75

30.76

38.77

4.72

528.63

51.80

6.11

1.83

26.11

2.00

425. .8

34.34

40.15

3.78

669.73

58.40

39.??

38.88

4.34

?2-.54

68.3.'

)GAR

a 6-

-- n 6 1127.24

10.85

0.8

6.86

8.0

8 .88

14' 54

2 72+1-I 624 2- 2 10 152.69

12.76

12.43

1.66

18.73

1.52

37.49

I/ 5."S

4 72 L-6 56

1- 1 16 176.14

12.85

9.83

1.69

21.2$

1.94

IS/ 515

7 72t1-6 57+ 1- 1 16 196.42

£5.63

8.76

1.41

2Z.38

21' 55

9 ?3+0-6 58+ 6- 6 16 212.57

15.29

0.07

1.04

24.26

23e54 It 74.6-1 63. 6- 3 IS 23052

£6.96

6.96

1.79

as- 5/5 14 7546-3 70.16-IS 14 257,69

£6.56

9.66

|.57

26."5/S 16 78+6-1 04+ 4- 4 16 272.65

26.27

7.4

8.0

SD

CONSUMED TOTAL TO DATE

SD

I2 3-5

£6 2618.06 26.49

Wittl XNAS

DAY TEMP

30)- 5e5 £8 720-6 70+ 2-

H

BODY WEIGHT XBR SD

DATE

SD

0.8

2-. 6,-5 Z1 72.1-0 61+ 3- 0 15 311.38

22.80

7.21

1.6£

27.??

2.10

50..

23 7t.O-t 61+ £- 6 £5 327.02

22.92

7.62

2.56

20.58

3.42

567.33

41.7£

40.90

4.41

81(.34

75.24

£6 338.44

24.57

5.7£

1.8

26.48

2.45

624. 12

41.0K

41.48

4.51

85,.

02.832

c')+ 8- 0 16 357.97

25.58

6.51

8.97

27.09

1.%6

?7.0

49.G1

36.96

4. 1?

4..6"

6/ 645 25 6C+2-0 6+.0- 0 9e G*4 28 ?I+eGROUP

DATE

£0083.1 9I.k .

2

DAY TEIM HUMIDITV

H

BODY M4IGHT CHG. BODYWIT, XBDK so XMAR SD

XBA

FOODCONSUMED TOTAL SD TO DATE

WTER SD

XBAR

SD

CONSUMED TOTAL TO DATE SD

£2- 5,'5

6

£29.65

3.94

8.66

8.86

e.e

6.66

8.66

6.6

8.0

6.66

6.66

14' 55

2 r2.1-1 424- 2- a £5 £48.28

9.'3?

9.32

3.68

*5.18

1.85

36.35

3..?

25.87

3.45

50.13

6.G6

1/

4 721-6 56+ 3- 1 £5 £46.44

11.22

9.66

2.86

E1.35

1.66

79.05.

6.3£

38.63

4.28

111.48

14.12

7 72+1-6 574 I-

54

6+ 6-

6*8"

i

e.8e,

£2.6.

6.43

2.43

23.68

1.75

148.30

£6.84

29.81

3.5?

280.83

23.98

21/ 5"5 9 131e-8 50+ 0- 0 £5 288.2£

13.04

8.24

2.56

24.82

2.86

197.95

13.47

32.48

3.58

265.79

30.45

6346- 3 15 224.54

14.68

8.17

2.76

24.09

2.10

247.72

17.11

48.15

4.86

346.09.

38.78

16.12

9.24

2.??

25.70

2.6

?24.5

72.85

37.97

4.S2

£1676+8-I 64 4- 4 15 267.53 36' 55 £8 72+0-8 3.+ 2- 8 £5 282.30

34.57

7.63

2.34

26.26

2.10

2.5C

27.68

3.12

26.54 20.71

36.78 43.41

4.23 5.51

58.1.

7.43

3774 3 112.9

537.55

17.97

64.37

6 .9

65 21 72t1-0 61+ 1- 0 14 .72.05

18.81

?.62

3.9

00.2£

1.92

517.3_4

33.,1 E

9. 8

6.21

7-1,. 17

63.;

1- 0 15 319.69

2.99

0. .5

4.38

2$.41

2.%9

576.16

38.12

41.2;

4.47

85.71

91.61

e/5 25 66*2-8 60+ 8- a 15 330.33

2Z.97

5.32

1.29

28.26

1.73

G32.68

40.52

41.84

:2.92

187.79

90.04

t5 1 358.78

23.55

6.79

2.38

28,39

2.70

717.46

48.12

36.77"

5.28

19e 55

23/ 54 It ?4+6-1

1 Is 191.72.

26e. 5,5 14 754-3 76+18-10 1 252.26

"

29e

2,

4" 645 21 7t. 6,

2

91 6-5 23 ?1*83-u GO+ GOUP

DATE

DAY TEMI' HUMIDIT

N

BODYWEIGHT XAIIS SD

12.23

6.88

6.86 068

4.9P

1.55

16 54

4 ?2-1-0 364 1-

)9B 5' s

6+0 8-6

6+ 6- a £6 1£2.56

FOODCONSUMED TOrAL SD TO DATE XBAR

CHG. BODY £WT. SD XAS

a'S 72.-1 162+ 2- 2 10 142.90 12.37

5

459.',9 51.30 4.5 5

£018.09 111.71

3

14/

It/

*

72+1-6 574

-

£6.51

SD

XBAS

WATER CONSUMED TOAL SD TO DATE

8.68

SD

6.88

e.80

6.6

8.68

6.69

8.66

1.45

33.8i

2.69

22.67

3.4£

4S.-3

.08

72.59

6.49

28.83

2.20

I0 95

9.27

5?

£92.40

14.82

1£6 £59.89

£2.76

8.16

1.84

1£6 £84.63

16.13

8.5£

1.46

21.39

2.34

'36.78

£2.85

29.82

6.82

25G86.24 18.82

58+ 6- 6 15 266.9)

£7.26

8.14

6.84

23.62

3.11

184.4£

16.41

32.88

4+4-I 63+ 0- 3 16 217,14

16.66

6.12

1.35

24.28

3.76

232.96

25.43

36.59

..6

333.43

25.48

44746-3 70+10-10 16 243.45

21.62

8.?

1.36

24.59

3..3

386.73

33.72

37.Ir

4."

444.91

36.42

269, 5,5 £6 78+6-1 64+ 4- 4 16 255.63

501.22

43.94

(.04 ?

51.Ho,

21e.'5 23" 54It 5

26

22.81

6.69

1.32

25.13

3.12

356.95

39.36

48.35

£6 72+0-0 91),- 2- 6 I 2A9.54

24.56

6.57

1.61

25.8

3.72

413.95

46.P3

41.49

4.63

6'5 21 72+1-0 61+ 1-8 £6 292.56

?7.2?

7.1.

1.39

26.7£

3.6

4F;6.09

55.16

39.03

.1.86 7Z1.30

0.Z5

1.94

26.86

3.46(

5-7.81

59,90

37.60

4.20

1.78

26.31

2.94

595.4

19 6.1.

48.5£

7.0.

1.97

27.46

3.30

A77.9

7' .4a.

3.40

'8, 5,4 2,

9 73.-0

4//'5

23 ?1+0-1 $,+ 1- 8 IS 389.05

6, S's 25 66+2-80

8.e.46

+ 8- 8 £6 317.0GO 30.41

9'h6'- 28 ?1.8-0 6 + a- 0 £5 338.6?

34....

38

k>

.4?

Al. BA

796.51

73,18

c.0

8,7 .53

63.44

5.4

9

,.30

1.73 97.9£

;r

.

-!-*a

I

. -

I Jl .. . _

.........

~, ....

.. ,.++.,

...

...

.

TABLE A-4 1 ou 4 EFOODT

i

DATE

DAY TEMP

I?, 54s

PO.*IiE

HUMIDITY

8"-a

BODYWEI1HT kBAt SD

H

0+ 8- a t8 131.01

CHG. BODY WT. XBHR 'i)

XBAR

S1

WATER CONSUMED

TOTIL 1"0EnITE SD

XPAR

61D

T'5."L TO DATE SD

22.46

8.8l8 8.88

OO

8.08

0.88

8.88

8.88

8.88

2- 2 16 141.32

12.,9

5,16

1.53

16.35

0.97

32.7

1.94

22.55

2.26

45.11

4.S2

I26/5.5

4 72*1-0 54* 1- 1 16 158.34

13.V?

8.5,

1.26

14.82

1.5?

72.33

4.75

28.?

2.98

282.66

B.8G

IS/. 5

9.10

14, 5,'S 2 2+1-1 6e

0.88

P ?2 1-9 57+ 1- 1 16 182.19

15.46

7.95

1.16

21.53

1.69

13.93

21 5.-5 S ?3#0-0 58+ 8- 8 13 L9O8.5

17.38

.43

13,4

23.80

1.9

182.92

23/

19.86

P.12

2.54

23.62

2.0

30.15

8.')

2.39

24.2?

2.23

302,.5

6.73

1.24

24.4s

2.14

351.07

70- 5.S 10 72.0-M S8, 2- A 2I 20?6.4? 21.0 7-15 7- 4-), 22 72+1-0 .+ 1- 0 1 235j.4% 5.01c 7.,'

2.27 1.01

25. 02! 25494

2.'40

) .').I2 '?.3 47.r7 .1. 14.A2 I

2.0'

24.33

e.3 "32.

1. .;

1.57 e3

2.2

9523".3-1 !-4 I.?.7X"".2

26.70

1.03

1114..:5

22 34*6-1 6300- 3 16 212.30 11

26, 5." 14 75+6-3 7?+10-18 16 238.836 212.41 ,5 "

14 P8.0-1 644 4- 4 16 251.'2

4" 6'S 23 ?+8-

52

V. 6'5 25 £-3+2-0 5' 6.-S ,8

2?.0-

DAYTlMP

I22 5-5

8

2- 4 IG 305.05

25.61

8- 8 16 316.38 0'

'5.23

.03 0-

' 16 3363.71

n5.28

8. I:'

-1.7:1

0. $1;

29.28

2.67

190.26

15.50

22.'.5 32.13

3.52

254.92

21,92

14.9?

38.41

4.432 331.4

29,52

20.14

37.56

4.93

441.41

43.0.4

20.t33 30..;

b.85

528.83

53.7%

r,.33 .,5

607.21f 30.7 72,..-' 75.,',

3 O ".

3:, ...

.

'3.2I3

1

..

4.1.

-,.7'3 7 7. I

457

HUMIDITY

81S-8 00

BODYWEIGHT XBAR SD

H

e- 8 1G 132.58

CHG. BODY W4T. >bAR SD XBAR

FOOD,CONSUM'1ED Tu2i. 51D TO DATE SD

..

f:;

.23.-'

WA.TER COiINi ED TOTAL SD TO DATE SD

(3,13

t.38

8,88

8.88

08

0.88

8.08

24' 8,-S 8 72*1-2 62 - 2 28 146.89 11.75

6.9$

2.59

27.87

1.15

34.4

742.37 4.27

0.80

o,8o

0.08

6.88

0.00

2."10 N6.54

1.50

45.88

3.01

20.8!3

1.52

101.26

6.042

Z76.25

2.92

2619.72

22.29,

247.38

14.817

2IV 53J

4 Pal-O 56" 2- 1 15 163.86

12.22

8.49

1.12

28.!6

1.39

29' 545

71+1.-n 974 2- 1 26 188.34

24.23

B.216

1.25

22.34

2.45

23 . 38

73-8-0 584 8- 8 26 283.84

2.82

205.f"4 22.65

21/ 5/5S

42.27 . .7i

5

GROUP DATE

23.42

8.83

8.07

24.46

7.76

2.32

23£?,

- 3 16 19.1

15.92

7.62

1,46

Z2.805 Z.2I5 231.33

24.8

26' 5.5 14 ?5.6-3 76418-10 16 245.?4

28.45

13.88

1.57

24.02

Z.81

33.39

20.24

35.65

3,58

427.73

30,.0C

28, 5-5 i6 7a+801 64* 4- 4 16 257.33

19.65

5.70

1.6,5 24.01

2.15

1'31,1

24.10

35.67

:.05

49'.48

37.93

25n72.o-o sR* 2- A 26 272W.

'0.38

. ?.f) 4

1.0.0

.223,5

70.'.2

3S.73

.'-

2272.2 0 ;1+ I2- 0 h4 262.54

0.54

6., 7

2.22 .

4/ 6,5 23 ?12.-1 4P+ 2- 0 16 344.,l

21.98

7.

1.61,5C

2a.? N..3

4.7"

1,3

...2

21.85

5.38

2,37

5.C0

23.-51U 11 74+0-1 63+

"0,8 5,5 ..

,' e.5 Z5 G.36 -8 GO+ 0- a02 3 9' 6-'52

?120-8 £2I+0-

DATE DAY TEMP

0

08 88

.

HUMIIDITY 0

83 0-

H

BODY WEIGHT X6BR SD

129.84

1

14' 545. 2 72*1-1 J2+ 2- Z 14 1.91 -

CHG. BODY WiT. >2363 Sb2

248R

4,.

22

927.13 54,3'

2.02

.

'1.30

:,7. .2 42 53."

47.0?

1

.

320,.79 21.u?

...

,3,.0

4.1.7

.:5. 1

.:,S

'.-w X

-. 37

32.,4

:3.04

.

'.

5'."

1;.*11,..1 "-1 127.'

6 .:

!8o3

SD

CONSU'EtD ToTAL To DATE £SD

8.88F

O.88

.88

8.88

8.88

8.84

8.88

8.o

-so

e.e

7.45

3.89

17.51

22 9

35.03

2.22

23.66

3.53

47.31

7.32

11.35

9.38

3.74

28.9?

2.54

74,6,

4.75

29,a5

3.69

ll,4Q

13.70

1.03 1.66

1.11.71 7.92 192.35 20.41'

24.53 32,0G

3.71 3.58

194.02 25b.14

23,70 25.43

0

19-'55 21 5/5

? ?2+1-0 57+ 1- 2 14 188.42 9 73'0-8 58+ 8- 8 14 285.28

11.8? 13.0?

0.30 8.43

3.43 3.51

22.58 23.,2

124 163.54

4TER SD

22.2

4 ?231 0 93;

1-

FOODCOMSUMED TOTAL SD TO DATE

12.81

26. 5.-5

227'1 5.5

.J.,4

2.4' .

71 1

2.01 . 10

-36.7a 3.05

6

GROUP

12 5"

IG 332.43

.1')

3i..o3 P.24

1 ?14+8-163+ 8- 3 14 2,81

11.50

7.86

3,39

24.53

1.71

241.41

22.67

38.71

5,.28 3^.,21

14 ?5+6-3 70+110-2 14 24824

27.99

.38

3,95

25.91

2.61

329. 13

21.13

37..5S

5.Ol

449,4

51.2

20.' 5,5 26 ?646-2 64* 4- 4 14 2G4.25

28.35

7.56

3.4?

26.210

2.59

31,32

23.55

36.81

5.02

52.1,0)

60.11,2.

';A, 515 1s ?2+0-0 5.,l+2- 0

22.64 1.86

7.00

2.42

2G.41

1,3

I 2.54 i

29.l2

41.,.3

23.43

8.22

37.9

2.30

7.',

3.75

5.;

2.75

26, 3'

2. 9'

4-3 I.-G-

13

2l 7221-83 421+ I- 0 24 3124.4 23 ?10-1 G2+ 1- 0 14 310,3G 25 GG30-0 fli+0 - 0

14 32.6*.2V.22 +0

i

4,2, 'f

2 0.

K?

221,5r7 . 7.5,.'

.

.

19'

.9

,11.4

"

31

''2

17

45

5.57

3:4..

41 421.25 1.?b,3. .2'

i

9

.

2,

4,7:3 l 99.,

1 4.

illii3

70..,

02.2. 9 73.4' C01..03.

4D-

,.

.,.~ 2

..............................

-. 45

39..-

m

TABLE A-5 EXPERtIMMlT G FMDO CLOi'SUMD DIATE

2.3."41"3 ,

6,3

DARYTEMP

0- 0-

0 4N

03.".?

10.40

1 4e

lO

I .50

0-0

2? ,347r2.0-04 31.)

6.15

7

2

?21-

2,51

5

13

44 4I

14,o ti

8,54

1.05

15.58

$3.00

16.37

4.85

.

0-

1- 0 40 3

I4 -2.k

22'

7/5

I8 72.0-I1

24'

7,3 22

1'l

?'5 Z23 7II-) -',

'1

I8'

-6

03'

5

74.0-2

4 48

,l+ 1)-

?$.+o-n c7+ 3

3

-1I

311.28

4.

3 13'.41

4$

355.33 1

' 9 '!,27,55

l 1-

-;I- M-

0 48. 17, F

(9.00

0.00

1.7,6

47.14

1.21 3R

.85

2. 41

20.6k

2.03

2.34

5'~

.5

.,

2.217

.Z

2.0 0.25

8.00-

1.45

21.55

4.54 31,

18.05

3.21

4).'TER

TOTALE

TO I-ATE

S1,

.00O

1.2

8.14

'0. 1*73.4

);Blip

0.01'

l2.'S

14 72ll-e .+

7e5

.8,A

4$ 3'3 .C5

-,8+-0Ga. a-

5'

CM0

0

3' ?'S 26 72*2-2 62' Z- 0 48 27.,32 7-

CHG. BOD' 2J. XbI:r, 1,

BODY WEIGHT NBAR SD

H

73-0-0 0,3+ 0-

a 2

'

HUMIEII.

X BAI

SD

9.00U S.53

16.39

22-. 6Q3

U.83

.

*-

TOTAL TO DAT'IE 51)

0.00

.00

0.8O

a.80

33.44

3.35

66.08

7.30

3.83

137.30

14.52

-. 1.4

3.82

241.82

R5.44

70

3. 53

325.q0.

31.58

9e4.81').2

.()4 04.86

¢CC,SUM'ED

SD

248.4?

U3.45

410.06

4.45

355.88

?5.56

2..17

357.56

27.41

"-.'.'0

3.

4M'.561

48.5

77.5,

2.03

413.55

32.4*)

"ii0,'4

5.33

'51.48

5.,11

1.--.3

7.70

3.'10

-168.71

31 .01

12.a" :

7',

3.'.'

55c,'

45.k::I

Z4..7 77',.2'.22

2$,?

24.77

4.1

2.2.

17,11

5.00

2.24 -

'.i

"..

27.4-1

2.23

.1-11

....

:I'.T4 05.I

1.

5,.q;,

.,.5

52."

.42

4.'

.,7 4.34

9"9.79

9513,0 "

.

".',I'

5.70

%I1.7 02.4.7'

1.;":'

''

5.,)

05?.5: 7

,

e'

7,5 273 9 7410-A 4'- 9 0

DATE

23'

"5

2" Z?7

A/5

1Ci.'615 Z.

?,3

DAY TEMP

8

Iq 35n.n8

NUMIDIT'

ti

8 48

" OD' IEIGiT X12R f.1

2)4.55

r',i. 812D',' WiT. 412P. S1D

FOnI'

1% 750 .'

"T8AR

C.ilSUrIE D TO'TALI TO DATE

S

i.%TEP 'E

3.79

-2.7'

3.7

3.7, 0

3.1,5

233.54

14.751

3. ' .4

2'~.4.1

25.28

4.321

71.03

33.72

343.,4

37.34

9,18

e.00

.3

1 471 221.13

11.57

8 .21

1.5

22.72

1.5

4

1 "48 aZ.75

I2.35

?.$

2.25

23."I

a.29

.IF.28

7.%4

7 72'20e A3' 0- 1 43 151."4

23.86

8.403

2.2'S 24.Z-5

2 .32

2305,03

14.2

3

2'.5 4.)-9

28.53 17.7

".114

0+

8-

7+0-0 62. 0

S 709I-0

2

0-040

o 0.210

U. cc

M. 00 45.44

15.55

8.84

2.5 4

a- 6 40 273.43 2

15.14

3.73

2 .75

-15.03

2.44

24X, ..)

14 ?I+1-0 622

0- 0 48 3)5.94

18.5!51

.2

j.(25

.?'.67

2.4

75.2;

9.43

16 72'1-0 '3

21-

.7

.3.2

2.':.

27M50

2."-.,

405.4

4.70

1. t.

'31 7/5

I0

7' 715 9'

'5

11

-"5 28 ?72+'-

14' 75227Zit

2

I-I.1

C'*

I4V.143

00 0.

48 32,.3i

0- 6 4S 332.,7

-'-

21,17

2477I5"

9.310 .5..,

.4a

1

0.00

65.167.29

,

2 ...

.2.'Z

.21

4,8

439

3.,,0

41.40

5.50

5,7."3

(5.42

.3-,0-24 3:1.41

41."5

,

C5.2 01

5.4 4

.J'.'.

.-.

,

, ?3-'3 235

g3. I-A

4$' 340i.7V

2.3..

5.9

:.'. ."

,"''2.3'

.,:4

t"'',,,) 44t.'7

25 74+0-1

63+ 0- .

,4b 34.6,55

24.97

4.13

2.0

:"

,.:5

,014.22' 53.44

4? 393.55

?.:'2

4.,0

2.-I. ,

28 ?4-0-0 64- 0-

5.t00

5D

3q.3

1I."

71S

1.00

) 120 5.25

20.90

10' 7', 1'

CON5U'MED' TOTAL. TO DTE

SU

2 ?3*8-0 e3+ 6-

0-0-0

228.8 ",

"'

40IAI.Ij

735.24

61.,')

'

,55,

'":'10 I,

.7.44 7.1,

$.:

5.73 -'r 4

'

:"

"I.'l . I

2I,"?

10".12P.741 2.73

IL

TABLE A-6 Ef EP IMUHT W I

GROUP

FOOD CONSUMED DAYS

5 5-

m

6

,'5

8e. 8a It

ma

£3." 9ea

DAY TEl'M

HUMIDITY

86.i-9

6+ 0- 6

s's

£6 197.81

WATER

TOTAL TO DATE

'D

XiAII

sD

CONSUMED TOTAL TO DATE

SD

8.68

6.86

8.60

0.6U

6.00

6.08

6.66

23.59

2.35

23.3

2.35

34.29

3.56

34.29

3.56

3 71+1-0 63+ 0-

1 £6 220.42

13.15

7.88

2.23

?4.31

2.33

72.24

6.54

33.8

4.70

£02.85

12.49

6 ?2+6-6 65+ 8-

16 243.53

£5.3t

?.s

1.58

25.28

2.60

14.89

14.23

32.98

4.54

28.?4

25.57

*

16 262.65

1£.05

9.56

1.92

25.63

2.0

19.36

19.61

34.33

5.33

269.39

35.47

1 16 276.2

19.65

E.78

.I?

?5.54

2.6

2560.44

24.64

35.97

4.83

341.33

44.87

I- & £6 36e.40

1.74

8.84

1.35

27.e

3.62

331.59

33.41

35.46

$.'IS 447.52

61.75

22.52

0.47

1.66

2.91

2.95

387.41

39.12

35.35

5.45

518.22

72.11

22.52

5.58

1.35

27.13

2.73

441.67

43.81

37.03

5.49

592.20

V,'-!

6.6J3

1.67

10.54

2.60

527.2

51.81

31,4r

5.9.%

701.501

28.32

2.58

543.82

56.64

34.26G 6.14

7.91

Z.17

639.74

66.25

237.46

5.94

29.8"

L.46

755.23i. 6.81

26.67

5.17

736-0 64+ 8- 6

68+

8-

8-

15 ?10-06

2?/ 815 22 72+0-1

8

£6 31?.34

67+ 6- 8

0*0-6

GC'JU

e+

9.7,4,6

8- 1 £6 34.'.0l1 25.29

2701-0 66+ 0- 0 24

911 26

DATE

SD

2.97

25, 8/5 26 72+--0 7i+

2

*3A3

7.84

22e- 6'5 £7 ?2+6e8 69+ 0- I 15 329.5£

29e"

CHG. BODY .T?. XDAR sD

£8.65

6,e, -1 £3 ?4+666-'

BODY hSIGHT XBR S.

1 F3+03-044+ 8- 0 16 285.85

6.-3 £6 ?4+a-e 6r.

IS.

H

9-

0.8

V.6

16 362.L"

25.32

6.08

1.39

16 371.7

295.86

4.86

1.33

£6 313.6

26.62

5.47

1.8

6.150

. lI..4

774.02

:e..94 122.77 62

17i

2

DAY TEMP

HUMIDITY

6 +.0-8 6

N

BODY iZIGHT N8AR SD

CHG. BODY WT. X2AR SD

X9AR

FOOD C ISUMED TOTAL SD TO DATE

WATER SD

XBAR

SD

8.e

COiSUMED TOTAL TO DATE

SD

6- 8

£6 18.16

13.94

6.86

8.88

6.08

8.66

6.80

8.85

Se'

I r3+01-8 64+ 6- 6

£6 197.I?

14.40

9.8?7

2.89

23.06

2.09

23.06

2.09

33.?6

4.45

33.76

4.45

Se 0'5

3 71.1-8 63+ 6-

1 16 211.74

16.29

7.28

1.29

23.69

1.69

70.81

5.67

33.84

4.27

10!.44

12.57

16 237.37

18.2e6

6.54

1.24

25.18

1.78

146.33

1.67

32.59

4.31

199.22

25.19

16 2t,5.64

26.44

5.14

1.65

25.67

2.17

197.68

14.35

32.56

4.76

265.13

34.09

268.87

21.01

6.GI

1.33

25.76

2.13

249.68

18.1£

35.10

4.92

335.32

.43.57

16 29 .52

23.16

7.89

1.31

26.86

2. 5

325.64

24.,79

34.58

4.82

439.84

57.32

25.64

8.686

1.4e

27.93

2.58,

3531

28.74

34.3!t

4.65

507.03

1591

6 324.11

25.87

?.14

1.48

27.35

3.38

440.0:

33.7?

34.81

5.8£

579.84

76.'t.l

1 18 343.56

26.74

C.C1

I.0"

?.9.04

3. I

51527.70

41.71

7123

4..A3

*5.

n .

£6 '3159.01 28.11

7.52

1.35

26.23

2.25

584.3C

45.66

7,4.15

4.6i7

7.,"3

16 368.44

26.94

4.72

2.86

22.19

2.40

640.74

4.5.50

25.67

A.

16 3".2.89

31.55

5.90

1,30

29.12

2.45

757.49

59.15

35.84

4.78

TOTAL TO DATE

SD

43R

SD

72.6-0 65.

£.e 8/5

6- 8

ma3 6 73.M-6 64+0-

13. 1£5

.5

IS/ 065

as-

16 74-e-8 G7

6-

1 i

13 74+6-Z 68+ 1- 2

ma5 £5 ?1+6-0 66+ 6- 6 16 365-.84

?Pen05

? ?+O-6 69. 8-

25, 8-5

20 72+8-3 ?1+ 0-

£

67+ 0-

27,

8a -s ?2.6-I

29

8'5 24 70.1-0 66+ 8- 6

2/ ',-3

208 8+6-9 GROUP

DATE

Se Ba 6

0.86

8.80

SI

38.4

5,0; .2 I47," -'..51,'24.

S

3

DAY TESP

6

6+ 8- 8

.808

HUMIDITY

6+0-6 80

"85 1 738-6

6-

8

H

£6 192.44 16

64+6 0-

BODY WEIGHT BARN SD

CHG. BODY W'T. HIAR SD

WATER CONSUMED

FOODCONSUMED XPAR

9.1?

6.68

6.80

8.66

08.21

£1.04

?.??

2.91

23.84

SD

6.6 ?.09

TOTAL 10 DATE

SD

0.86

8.66

6.68

6.6O

9.60

0.0

23.e4

2.89

34.18

5.15

34.18

5.15

8e Se5

3 71+1-8 63+ 8-

1 16 216.59

£1.69

6.19

1.51

24.69

1.68

72.43

5.89

34.57

4.44

103.31

13.C.8

Ile 05

72+068 656 0-

16 243.94

14.28

9.12

1.35

25.72

1.64

149.59

$.65

34,34

3.80

206.34

24.99

8 ?3+0-6 64+ 6- 8

16 261.6?

16.54

8.87

..71

26.22

1.99

2802.83

13.42

3S.68

4.2?

277.7£

32.04

I

£6 276.44

19.32

?.36

1.85

25.93.

2.15

g53.89

£7.41

36.35

3.68

358.41

38.73

16 362.32

21.86

8.63

1.23

27.96

2.29

337.58

21.73

36.31

4.29

459.34

50.62

16 326.33

23.78

9.61

2.1?

28.19

2.'46

393.97

25,90

36.47

4.45

532.28

54.11

333.37

24.39

6.52

1.46

20.53

2.22

451.02

38.17

39.79

4.18

611.07

1 14 353.78

27.05

G.60

1.49

09.45

2.711

531-.30

37,2*

30S.10

4.42

72.1. 1

79.917

16 ,64.41

29.37

?.G2

2.3

8. .32

2.83

598.01

42.41

37. ?

4.54

-. 1.71

ES. Q

IS 31

7

31.52

3.'3

,.

Z8.76 216

2,Z8

655,53

4A.16

39,33

5,40

c74"'..A

95 "

398.50

34,21

5,31

1,56

7'9.11

2.73

771,9C

55.:?

37.9

5.75

13

8"S

IS.

S'S

16 r4+-0

s?+ 8-

".S 13 ?4+6-2 66+ 1- 2

16' e9 "

15 7t.6-8

66- 6-

22a ma

1? 72+-8 69+

251 9'5 28 ?2+6-8 27" 8'S 22 ?2+8 29/ Z

1

71+ 0-

0-c-0

1 1

?+ 0- 6

815 24 rOI-D 66+ 9,'5 26

-

6

o

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i6

|~~~~~~

mil

41

j~~r;--

>.

a wII

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:

TABLE A-7 GROUP 4

HUMIlDIT

DATE DRY TEf?

24

BODYWEIGHT SD AR

CHG. BODYWT. sD REAP

HOAR

FOODCONSUMED TOTAL TO)DATE sD

WATER sD

REAR

SO

CONSUMED TOTAL TO DATE SD

V. S'S

8

a. 0- 0 i0 188.15

22.36

0.05

6.009

6.08

0.08

8.00

6.00

8.668

41 84s

2 73+0-8 S6t. 0- 6 16 186.34

11.4?

7.99

2.42

21.80

2.08

21.88

1.60

31.1

3.06

31.51

3. Ole

11- 6/5

37?1+1-8 63+ a- I 20 210.63

12.6?'

e. 14

1.02

23.21

2.98

68.90

5.73

32.45

3.16

%.,40

9.00

11, 84S 6 72-e-8 65* 0- 8 26 Z35.61

14.38

8.33

2.30

24.53

2.75

142.49

18.42

31.59

3.49

192.38

26.90 25.43

0+0-0

0.60

8.0608.86

8l-6 10 252.63

25.66

8.52

2.30

24.83

2.55

192.1$

135.33

32.98

3.3?

250.35

150 845 ID ?4+0-0 67. 8- 1 26 264.;15

15.95

5.62

1.160

24.65

1.67

241.46

16.29

73.28

3.31

324.74

18' 645 F) 74*0-2 60. 1- 2 16 k87.48

2756

7.74

1.40

25.12

1.84

319.24

2,1.81

33.69

3.42

4,15.83

20--0415 7*0-86 64*8- 8 16 303.99

29.74

0.25

2.04

26.20

2.63

371.79

24.62

33.98

3.93

493,63

ARIA

19.58

4,51

2.90

27.1?

3.98

426.12

3123

35S.22 3.6?

563.08

5-.17

PS/'085 28) 72--0-43 71+ 8- 1 16 33 2.82

23.53

C,.6.4

2.030

?7.24

7-4.7,5

4.53

66A. 32

Ve 045 22 r2*8-1 67* 0- a 16 34C.43

24.91

6.76

1.42

27.19

2,01

561.32

40.77

3.C2

7.62

7346

73.1,.2

3.62

1.67

27.1?

2,07

6251 .65

42.74

36.43

4. l

205. 14

74.22

41.75

1.19

27.$

7' .19

?24;i%:

$0 or,

b4.2c*

3.9 5

13e Boos 0 73+0.6

4*

8- 1 14i313.02 I?720-6 ',f)* 1?5

??e

0/.56 24 '0.1-0 66480- 8 15 7353.06 26.23 2' D's 28

8+0-0

04 0- 0 26 .372.66

26.59

6."I

.. E1

*

507.02

*

*

32.64 40.99

*

1.5

'G

007

GROUP 5

DATE DAY TEMP HUMIDITY

H

BODYWEIGHT REAR SO

0+ 6- 0 10 292.62 22.20

CHG. BODY61T. SD REAR

ROAR

WATER

FOOD CONSUMED TOTAL SD TO DATE

SD

ROAR

SD

CONSUMED IOTAL TO DATE SO

3.063 8.80

5e Ties

a

0.80

8.138

00

0.28

8.06

8.86

8.06

6.06

0' 8/5

1 73*0-8 64+6-0 816 290.6f

22.56

6.66

8.03

23.88

3.43

23.60

3.43

33.54

3.48

33.54

0e8"

3 7141-663+ 0- 2 26 224.36

23.24

7.84

2.32

24.22

2.66

72.2J9 5.12

34.26

3.90

282.6?

8.AI

It2,-9546 72+4-8 45* 6- 0 16 240.99

24.66

6.66

8.95

25.76

2.89

149.96

20.69

34.24

3.2

264.48

17.50

64+ 0- 6 16 259.17

25.96

9.89

2.57

24.73

3.44

263.02

27.13

35.94

4.56

2767

25.63

67+ 8- 2 25 2172.88 26.48

5.96

2.44

25.89

1.92

2S4.82

29.69

35.38

4.65

346.668 34.39

13e S'S

0+0-0

8 73+06

2-5'85 20 ?4.60

3.48

17.1080.29A

2.28

27.53

2.40

337.42

27.A2

35.75

5.32

453.93$

48.22

29e 8415 72.8-6 664 8- 0 !6 322.56

27.65

7.02

2.23

27.*8

2.54

393.3C

31.76

35.27

6.20

524.4?

C2.17

21' 8,1517 72-0-8 69'.0- 1 26 2-23.27

27.95

3.85

2.72

V7.47

2.34

443.32

35.2?

38.22

5.0 11)

5e A/!% 28 72.0-0J72* 0i- 1 26 3 41.139 2801?

4.2

2.7$1

3.32

SV 2. 19

425.1

:A7 3 q

0,.49 712.3'

2.0 l

%..O

594.

33

13 74+6-2 68+ 1- 2 26 295.93

28-'8

27'e 6e'

22

20l 64 24

12. - +70- 0 26 359,.23

16.27

7.6?,

68. 0- a 15 317.214

20.65

4.060 I.27

I'8.04

CHG. BODY WT. ROAR SD

R

9*-

GROUP

'ATE

DAY TEMP. HUMIIDIT'

I 73+6-0 64V 8- 0 16 3 71-1-

BODY WEIGHT ROAR SD

64 - 6+IS 192.32

8

4, Ile e 015

6.-0

H

5+68,6

I' 8"

99.56

63+ 6- 2 16 225.02

6 72.*- 65* 0-

9.55 26.48 169

5 16 242.56 13.05

/2 73.-0 64+ 0...-0 A5 k tB8 .. 15.23 6,4 . 2 259.29 3/ 8e5 07,0-..".....s

25' 8'S 10 74--80670- 2 1 267.42 Joe 9I' 13 740-2 68 201. S

2- 2 15 295.49

5s' 720-0 66* 0- 8 26 322.72 s8 1 ?12.-86*6-

5'

72 O' 720-0 7.

/

1'54 22 72-0-2 67+ 0-

79'

B'S

263124.45 1

A- 1 16 3

0+0-0

15.56

6 2; 7,5F,

0+ 0- a 15 390.37

7..19

2. 05

0.00

O.0E

2.206

23.24

2.51

7.73

1.39

26.37

6.4

.65 12.3

25.84

5.45

5,96 6.02.

6.22

.RI

SD

CONSUMED TTAL TO DATE SD

8.8

0.00

0.0

2.51

32.54

3.60

3,54

75.98

24.26

33.03

4.26

98.61

153,48

29.87

31.919

vAA 4.42

94.66

.B 0. 6.6 23.24

0.8 3.68 22.75 24.9

4.1 9. 5, 92?04.47 35,.37 33.9 3

454.96

322,.42 .

3.9 34.29

39.5?

34.02

4,76

332.03

42.94

4.64

339.27

53.0

35.0

5.48

437.24

!:.S3

339 1376 f

6.2

63 0

3.13

5.A4

;.6

27.3(

1I.

2.4-4 205

3194.22

2157

440.E9.

2.41 4.2

7.

5.49

WATER SO

121; 1'3.2

257,32

1tIA

15.19

6.42

, 3.50

2.72

41.34

30.21

91t?

7R6..9.1 l0~

2 .63

8.62

23.76

4.93

.4 ,8 25.49

. 1.32

19.90

fl'3 2

*

r 3 3.50

.0717

1,.71 2.43 .46

Z-.,54 64

852I.66 59.22

¢QOM ED FOOD TOTAL 70 TO DATE

7.24

6.o t.tO

0.0Z

2?.22

141.4 :0.72

24 ?0-1-0 6C,- 0- 0 16 303.43

2' 9,15 ?a

.30

,

2.

70.17.

?

.9

2.87 2.2' 0

42

66.A6

361,2

7

3.32 .4

594

701"

01.9

.A6.47 760. 3,4

12.3'

5,3.2

7. 5X7"

33. 3t"3..403 36.,4

42 67,9 , -7,17,-

.711 .2 117 4.

..2 4.49

I-.', .

.12,:

.;

1,'11 .; r.,.0

APPENDIX B Data and Statistical Summary of the Biochemical and Hematological Analyses Tables B-1 through B-11 present the raw data and a statistical summary for the biochemical and hematological analyses for each experimental group of the four experiments (E, F, G, and H).

The average per day growth rate over

the 28-day exposure period and the final weight are presented for convenience. The field strength and chamber position (either upper or lower) for any group can be obtained from Table 2 of the text. Note that in experiment F, animals numbered 231 (Table B-4), 687, and 689 (Table B-6) were deleted from the growth analyses as discussed in Appendix A. A 999. 0 was used in the final weight (FNL WT) column and a 000.0 was used in the growth rate column (WT/DAY) to identify the deleted animals.

When

a 0. 0 is encountered in the rest of these tables, it indicates that the biochemical and hematological determinations were not performed.

This only occurred as

a result of an insufficient quantity or a clotted sample. Histograms of these data indicate that nonparametric techniques should be used for their analysis.

However, the first quartile (Q-1), median (MED), third

quartile (Q-3), the number of animals (N), the average (AVG), standard deviation (S. D.), and standard error (S. E.) are presented for the statistical summary. The headings for each column are defined (proceeding from left to right) as: ID#

three-digit code randomly assigned to each animal of an experiment.

The digits for units and tens specify

where each of the 96 animals was positioned.

The digit

for hundreds specifies the chamber number (1-6) T. P.

total serum or plasma protein (g/dl)

GLOB

serum or plasma globulin (mg/dl)

GLU

serum or plasma glucose (mg/dl)

43

i.........................................

T. L.

serum or plasma total lipids (mg/dl)

CHOL

serum or plasma cholesterol (mg/dl)

TRIG

serum or plasma triglycerides (mg/dl)

WT/DAY

(final body mass-initial body mass)/28 (g/day)

FNL WT RBC BC

final body mass (g) red blood cells (cells/m 3 x 106) we blood cells (cells/mm x 103

WBC

white blood cells (cells/mm 3x 103)

POLY

segment neutrophils (%)

LYHS

lymphocytes (%)

HCT

hematocrit (%)

HGB

hemoglobin (g/dl)

II

II

44

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