[PAGE 1]
ADDENDUM

GROOMING ACTIVITY OF ALBINO MICE

EFFLCT OF CHLORDIAZEPOXIDE AND1900

INTRODUCTION

Chlordiazepoxide and1900 were employed to explore the usefulness of a test based on the grooming behavior of albino mice, according to the method of O. Rohte, (Brit. J. Pharmac., 34:236, 1968) for the detection of sedative agents. Rohte states that the main advantage of this test is that it reduces the difficulties in testing weak sedatives by providing a releaser mechanism to the instinctive grooming movements of white mice.Rohte found that administration of certain test substances was associated with a significant inhibition of grooming movementscompared with control group.

METHOD

Apparatus - Observation boxes wereclear plastic with dimensions of 5.5 x 10 x 5 in³. The boxes were covered with wire screening and the bottoms were lined with cardboard. Activatedcarbon, Darco grade S-51 of the Atlas Powder Company, was used to cover the mice. All injections were made.with a 25-gauge needle and glass syringes.

Subjects - The subjects were 60 female albino mice (19-31 grams), HR/ICR strain supplied from theo

[PAGE 2]
Procedure - The test mice were covered with pulverized charcoal and the intensity of blackening and its pattern were noted at 1.5 hour intervals up to 6 hours. The observations were graded on a 3-point scale: 1 = white to light grey; 2 = medium grey; 3 = dark grey to black.

A. Chlordiazepoxide. Five subjects per dose level per day received Chiordiazepoxide (Librium HCl) intravenously between 8 a.m. and l0 a.m. or a saline solution (control).Dose levels of Chlordiazepoxide were 125 mg/kg, 63 mg/kg, 32 mg/kg, 3.2 mg/kg, and 0.32 mg/kg. Saline given at a level of 10 ml/kg (largest drug,volume of solution injected). All animals at a single dose level were injected on the same day.After injection the animal was placed in a jar containing the powdered carbon and covered by shaking the jar. The subject was then removed by tail with forceps and placed into a shoebox cage lined with sawdust for approximately 3 minutes. Subject was then placed into a plastic observation cage with ad libitum food and water. Observations of the intensity and pattern of blackening were made at one hour intervals according to the method of Rohte (1968) (see Figure 1,2). Observations were made on both dorsal (ventral ) surfaces and each body area was noted as either dark, medium, or light (3,2, or 1, respectively).

B. 1900. Two dose levels, 1 mg/kg and 0.1 mg/kg were. tested in five mice each, according to the method used with Librium.

-2-

[PAGE 3]
RESULTS

The "grooming unit" appears to accurately measure grooming activity during the first few hours of test. At later intervals grooming becomes more difficult to measure because the observer is unable to note additional grooming activity on areas which are already light. Therefore, the decreased rate of grooming toward the sixth hour in all groups except the controls (see Figure l0) may be due to the inability to measure grooming rather than an actual decrease caused by the drug.

For purposes of statistical analysis, each subject was assigned a grooming index at each hourly observation (see Table l). The grooming index was obtained by summing the grey values (light = 1, medium = 2, dark = 3) for the six body areas. a-f. Each dose group was compared with the controls in a. Wilcoxon two-sample test (Laurence and Bacharach, p. 80) (see Table 2). The test was not applied to the one-hour data because of the large number of ties.

For Chlordiazepoxide, the grooming indices of the 125 and 63 mg/kg dose groups were found to be significantly decreased as compared to controls throughout the 6-hour period. The 32 mg/kg group showed a significant decrease in grooming at 2nd and 6th hour. The grooming of the 3.2 mg/kg group was found to be significantly decreased as compared to controls from the 3rd to the 6th hour. No difference in grooming was found with the 0.32 mg/kg group.

For900, 0.1 mg/kg was a no effect level. Mice which received 1900 at l.0 mg/kg showed a significant decrease (P = :05) at the l-3 hour intervals and very significant decrease (P = 0.01) at the 4-6 hour intervals, when a one tailed test is considered.

-3.

C

[PAGE 7]
DISTRIBUTION OF VALUES OF THE GROOMING INDEX ATEACH 1-HOUR OBSERVATION FOR 5 DOSE LEVELS OFLIBRIUM HC1 AND FOR SALINE CONTROLS

TABLE 1.

[PAGE 8]
-VALUES FOR A TWO-TAILED TEST

ON 5 DOSE LEVELS OF LIBRIUM HC1

() = Not significant

\* Rumke, C. L., and de Jonge, H., Chapt. 3, Design, Statistical Analyses and Interpretation in Vol. I Evaluation of Drug Activities: Pharmacometrics ed. by Lawrence, D. R., and Bacharach, A. L., Academic Press, N. Y. 1964

4

[PAGE 9]
DISTRIBUTION OF VALUES OF GRÓOMING INDEX AT EACH 1-HOUROBSERVATION FOR EACH OF 2DOSE LEVELS OFS1900e

[PAGE 10]
TABLE 4.

P VALUES FOR 2-TAILED TEST OF 1900 0.1 MG/KG VERSUS1900 1.0 MG/KG DOSE LEVELS

6

22

[PAGE 11]
LD5036.0-28.0-450- MED500.56 -0.18-1。8- RAT10 LD50/MED5063.0

[PAGE 12]
INTRAVENOUS TOXICITY TO MICE

REACTION SIGN

MIN。 DEG- MIN. TO

APPR REE RECOVER

6

[PAGE 13]
LD50 100.0-63.0-160.0-

MED501.8-0.56-5.6-

RAT1O LD50/MED5056.0

[PAGE 14]
INTRAVENOUS TOXICITY TO MICE

MG/KGML/KG FRACT.

REACTION SIGN

MIN。 DEG- MIN. TO

APPRREE RECOVER

C

[PAGE 15]
CS 2367 W 3

[PAGE 16]
LD50100.0-79.0-120.0- MED505.6-1.8-18.0- RATIO LD50/MED50 18.0

[PAGE 18]
.LD50 110.0-89.0-140.0- MED503。2 -1.0-10.0- RATIO LD50/MED50 35.0

[PAGE 19]
2

[PAGE 20]
PAGE 2598

[PAGE 21]
INTRAVENOUS TOXICITY TO MICE

REACTION SIGN

[PAGE 22]
LD5028。0-22.0-36。0- MED50 03.2-4-1.0-4-10-3- RATIO .LD50/MED50 8.8+4

PAGE 2598

[PAGE 23]
LD5036.0-28。0-45。0- MED500.56-0.18-1.8- RAT10 LD50/MED5063.0

24

[PAGE 24]
2717 W 2

[PAGE 25]
LD5045。0-36。0-56.0- MED505.6-1.8-180- RATIO.LD50/MED50 8.0

2778 1

[PAGE 26]
INTRAVENOUS TOXICITY TO MICE REACTION SIGN

MIN. DEG- MIN. TO APPR REE RECOVER

[PAGE 27]
LD5020。0-16。0-25。0-MED500。18-0。056-0。56-RAT1OLD50/MED50 110.0

[PAGE 28]
INTRAVENOUS TOXICITY TO MICE

[PAGE 29]
LD50 100.0 -79。0-120.0- MED501.8-0.56-5.6- RATIO LD50/MED50 56.0

[PAGE 30]
2897 W 2

[PAGE 32]
2

[PAGE 33]
LD50100.0-79。0-120.0- MED500.56-0s18-1。8- RAT10 .LD50/MED50 180.0

C0/20

[PAGE 35]
.LD5063.0-50.0-79.0-MED50 0。18-0.056-0。56-RATIO.LD50/MED50 350.0

[PAGE 36]
INTRAVENOUS TOXICITY TO MICE

REACTION SIGN

[PAGE 37]
LD50200.0-160.0-250.0- MED50.18。0-5。6-56。0- RATIO LD50/MED50 11.0

2 2 i

[PAGE 38]
LD5025。020.0-320= MED50108-0。56-5。6 RATIO.LD50/MED5014。0

[PAGE 39]
LD5025。0-11。0-56.0- MED50056=0.18-1。8- RAT10 LD50/MED50.45.0

[PAGE 40]
INTRAVENOUS TOXICITY TO MICE

2

6

[PAGE 41]
LD50110。0-89。0-140.0-MED500。56.=0。18=1s8-RATIO LD50/MED50 200.0

[PAGE 42]
2

[PAGE 43]
LD50120.0-100.0-160。0- MED501.8-0.56=506 RATIOLD50/MED5069.0

5031 W

C

[PAGE 45]
LD5036.0-26.0-49s0- MED503.2-3-1.0-3=0.010- RATIO LD50/MED50 1.1+4

[PAGE 46]
INTRAVENOUS TOXICITY TO MICE

REACTION SIGN

MIN。 DEG- MIN. TO

APPR REE RECOVER

: S

[PAGE 47]
INTRAVENOUS TOXICITY TO MICE

[PAGE 48]
LD50 160.0-120.0-200.0-MED500.18 -0.056-056-RATIO LD50/MED50 880.0

[PAGE 49]
INTRAVENOUS TOXICITY TO MICE

[PAGE 50]
LD5050.0-40.0-63.0- MED500。56-0.18-1。8- RATIO LD50/MED5089.0

5071 W 1

217

[PAGE 51]
INTRAVENOUS TOXICITY TO MICE

PAGE 507i

[PAGE 52]
.LD50320.0 GREAT MED501.8 -0.56-5。6- RATIO LD50/MED50 UNK

[PAGE 53]
INTRAVENCUS TOXICITY TO MICE

PAGE 5092

[PAGE 54]
LD50 100。0-79.0-130.0= MED500。018-5.6-3=0056= RATIO LD50/MED50 5.6+3

C

[PAGE 55]
INTRAVENOUS TOXICITY TO MICE

REACTION SIGN

MIN。· DEG- MIN。 TO

APPR REE RECOVER

PAGE 5104

[PAGE 56]
PAGE 5104

[PAGE 57]
LD5056.0-45.0-71.0-MED500。18-0.056-0.56-RATIO LD50/MED50 310.0

[PAGE 59]
LD50.20.0-16.0-25.0- MED500.56 -0.18=1.8- RAT1O LD50/MED5036.0

[PAGE 60]
INTRAVENOUSTOXICITY TO MICE

[PAGE 61]
LD50 320.0 GREAT MED501.8 -0.56-5。6- RATIO .LD50/MED50 UNK

[PAGE 64]
PAGE 8638

[PAGE 65]
.LD50 32.0-25.0-40.0- MED500.56-0.18-1.8- RATIO LD50/MED5056.0

[PAGE 66]
INTRAVENOUS TOXICITY TO MICE

2

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[PAGE 211]
OBSSONS

[PAGE 214]
DECIR ADCCI DITY

DEII DECCI DITY

LOSUE URE DE DEC ITY RBE  CS  CSGE DECATO RAROO REAOTE IRPATE RROT RTE ITPBRPOR DPROR PPTH TNCOI RRR PTH

LOSU PURE INCTER RAROR REAOTTE DECED OR ODERTH DECNGRCNG RENCY DER D ADITY

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[PAGE 215]
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DE RR ROTE DES DER DETH DECG RECGGNCY DECR DCSO DSITY

OBSNS

[PAGE 219]
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PAGE A-

[PAGE 223]
$$
\begin{array} { r l } & { \frac { 1 } { 2 } \leq \frac { 2 } { 3 } < \frac { 3 } { 2 } < \frac { 1 } { 3 } } \\ & { \frac { 1 } { 2 } \leq \frac { 3 } { 2 } < \frac { 1 } { 2 } \leq \frac { 3 } { 2 } ; } \\ & { \frac { 1 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 1 } { 3 } ; } \\ & { \frac { 1 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 1 } { 3 } ; } \\ & { \frac { 1 } { 3 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } ; } \\ & { \frac { 1 } { 4 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 4 } } \\ & { \frac { 1 } { 5 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 4 } } \\ & { \frac { 1 } { 5 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 4 } } \\ & { \frac { 1 } { 6 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 4 } } \\ & { \frac { 1 } { 5 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 4 } } \\ & { \frac { 1 } { 6 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 4 } } \\ & { \frac { 1 } { 6 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 4 } } \\ & { \frac { 1 } { 5 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 4 } } \\ & { \frac { 1 } { 6 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } } \\ &  \frac { 1 } { 6 } \leq \frac { 3 } { 2 } \leq \frac { 3 } { 2 } \ \end{array}
$$

[PAGE 224]
NOIO I       ION NO M  MST RESRE PR PP48 HEA E  BUTE

BOU  OUUY TUGRESR RS R 3HEA BUE  BUTE

RES PS PMPO3G HEA T  BUTE

PAGE

PHL  OOL L O OTS

[PAGE 225]
O   E NO M WRS MSST B0OE  BOUUY TS RES RE R PR4B HEA BE U BUTE

NEUML N NMAL AEHAAL L LAHANC NOE M  ST BOE  OU 09 RES PSPUTE HEASN S UT UTE

PAGE A

0

[PAGE 226]
BOUE  U U° 72 RESS PS PUTE HEN BE UE UTE

SCE ER- ER

HE BU RUT RUTE 09
RRES S OTE 8  OU U2

HE T E UTE 09 RESS S MOTE O  OU UU

(A-1) 95E1)

TIMME HOURS D0E ANG SKG CATNO

PAGE

PH C COO C CAOO COTS

[PAGE 227]
NEUMAL TLTMAL BEH O CGALA  MAL OTN SITTN TN

SOL ER- OLER

BOBUYU. RES RES PS PPGG0 HEN E UE UTE

BOY  OUY U.T R RES PS PM6 HE  EU BAUTE

BOE  OUY U.G R RESS RS MO HE/EE BEUE BUTE

HOURS M/KG AND SEX TIMME 305E CAT NO

6

[PAGE 228]
NOO    D C S  ON DEEN MC ML MCCUS OEFE R RA TR IEN RESCO RACCD RCCION

NO OHE S H E NON D A RY BOE OUY TU.2 99 RESS PS PMOUTE HE E  BUTE

HE E EU BUTE 99 RESP PMOTE B0O  OU TUT NOD MN  IST

(-19E2

HOURS K. M/KG AN SEX TIIME 3500 CA NO

2

[PAGE 229]
NEUL  MAL ANL N CMAL ANNEE

H  TE UT 54 RBE PRS CUTE BOOE  BBZ

RPEEPEE P PO4 HE BEE UE UTEE

6BS P PUTGE HES  UTE UTE

PAGE A

O ION D TS

[PAGE 230]
HEA  U UTE RES MOTE 9E BOY  ORUY TU° NO  A RY

HEA T UE BUTE 87 RESS S POUTE BC  OUY O

B0O  TOUY TU8.3 RES RES P POT7 HEN E E BUT

PRE DSOSE SDOY NESLS L NAL BEH GRCE. CALM

HEA BT E BUTE 87 RES S MOUTE BORUU4 NO CL  ST

HOURS M/KG ND SEYX M.IME 3003 CATT NO.

PAGE

[PAGE 231]
SUSSD  - LISNT

N E NHE EF HREAAET MARRER D6E6 BEEE P PO36 EAUE BS UUTEG UTE

HENE EE UE UTE BEE R MOTE 9E BO Y ORUUYU82

[PAGE 232]
SUSO OT- SUNT

NOO   D   D I ON NESOL CL AL NO M IL OIST

HEA  U UTE 9E RESS PS MOTE EOO  UY UT

NO  C    CD C ION TENO    D D       LY ND D

BOE UYTUY RE5 ESS PS PMO3G HEA BUTE U BUTE

BO  UY TUY RES PS P42 HEA E  BUTE

HE E ET BUTE 42 RRESS PS POTE BUTU8°L NOO S O S

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TIMME HOURS TM K MG/KG

PAGE A-

[PAGE 233]
NHC  CS CE CAL BOE  RU TUB RE RE PS PO5 HEBUE RU RRUTE

NHC D CH C CAL BOUE  BOU TUS RERESS PS POS4 HE B BU BTE

NHSCE N HG SCE CAL B  OUUY TUU9 RES RS PS POT2 HEA/N S UIE RUITE

[PAGE 234]
SUSO DONDUSONT

NHC   CE CAL BOE C O T RE EO PS MP36 HEU 1ET  UTE

NH  S CD CAL BO  3OUU U RES RESS PS PO3G HE   BUTE

NHC  H CCS CA B0OU  UY TUS RESPREO PNS PO4 HEAU E U BUTE

[PAGE 235]
SO O  D  D  E  ER

HEA   UTE 6 BEHAA GR NCE RES PRS PUTE BOR  OU U

NO  N E BOUE  3UUU RE E P MUOG6 HEUU  U BUTE

NO  OHE S E IIP B0E  ODU U° RESRESOSS MOT HESSU EE UE BUTE

PRE DSE DRSAY NES  NMAL BEA GRANC BOU UTUA RESPES POTZ HE  E UTE

PAGE A-225

[PAGE 236]
BEL CE BOE C BO UL ESE UE UE UIIE

EASNS BE UT BUTE 78RES PRS POTE BE  CO

BBPAL CE B8 BEOE PSE PAOTB MESS E USE BUTE

NE S MAL BEPAL ANC RBOE  OU3 72 PBEGIOTS P PRPOUTE HEAAE EE UUTE UTE

PAGE A-220

[PAGE 237]
S O      E  ER

NO  O S I P SLOO IA PUPE RY RRETE PU  ED NOI A W MSS

HEA S  BUTE 06 RES PRS PROTE BO  U TU8S

HEATUG BS BU BUTE 06 RES R UTE BOE C BO U.

BOD  YU2 RE4 E P S4 HEA   BUTE

PRE DE SAY NEU  MAL BEHA GRO GRCE ACTTVE

NOS C  5T BO0 Y O °2 RES REE P P48 HE B U BUTE

PAGE A-227

[PAGE 238]
S  H O      R

NEAL  MAL PUPL OMA ALL RMAL PUPR  RRAL PDE ED

BO  U4RESRE RS PT4HEAB BE BUT BUTE

NONS   EI SLH G1A PUPR RY RRAMAL PUP  STED NO M MMS MAIST

BOOE Y ORUY TUB RE PE PR PMO4 HEAG E G BUITE

PAGE A-22:

[PAGE 239]
NOTO C D L   S ON SKIG OOG N NKS SL TNSE A

BOD  RY TUGRE R P I36HEA  U BUTE

SKINN ONG N FNKS I NI N S NI N BOE  RU U° 36 RESS PS PUTE HEA  U BUTE

HE   UTE 09
RESS PS PUTE BO  BUY TU2

PRE DSE SDOYL NEUM MAL BEAL GB GANC BO  ORY O RE5 PRS PMPO54 HEA IN BUTE

T1ME HOURS CATO . O NSE MG/KG AND SEX

PAGE A-229

[PAGE 240]
SUSD DO ENT

NELL  AL OEL ANC BU  8S 78 BNI P PTIRTASUTE EE E UTE UTE

HN/ B  1UT 54 BESO PR PTE BOE  RU8S

TME HOURS T. G DO0SE MG/KG CA NO. NDD S5EX

PAGE A-230

[PAGE 241]
SUSD SUON SUSSONT

06 NO N E SHE E IP RES UTE BOORY Y OU UT SLOA IA

NO     C SCD  ON SLL01OHA 1H1IA

HEU  UG BUTE 78 RRESMPTE BOY  UY TU°

BOUR  OUY TU° RES RES M OT78 HEAG  UTG RUTE

PRE DSOS DPROY NEUL L NMAL BEHA O OANC REG G G GE CALM

HEA   BUTE 90 RES M UTE B0O  OU TU9 HOIA INA

HURS ANMNG SG TIE ODT CATT NO.

PAGE A-231

[PAGE 242]
SUSSUSONT

O OE SE N SLIOHATA PIA BU  UY U°Z R5 P8

NOI     C   ION O   S H P SLH HBA S HIA

B0  UY TU0 RES REO PS PMO9 HE   UTE

[PAGE 243]
NIIO MNG G G ION NO C O ST

BO Y UY TU°3 RES R P PS HE B E BTE

HE U U UTE RES MS MOTE 9€ BO  OAU U3

HEA BUE U RBUTE 87
RES S UOTE BOD  ODY °4
HEA U BUTE 99
RESS PS PUOTE BODE   TUB°3

[PAGE 244]
NHHCE ED CD CCD CAL B  RUYTUZ RESRES PS POT72 HEA  U UTE

NEU NMAL BEHRAL GR GRANC B  U U°B 5.0 RES P PUTE HE BE  BUTE

PAGE 234

[PAGE 245]
IO O S N NF NO A  ST B  BOU TURL. RE R PR PPU448 HEAN B UTE UTE

HEAN UTE EU BRUTE 9€ RES PNS PUTE BODE  BOU TU NOD D R DRY

AO1 N 7O 3SSNT RERE P PRI42 HEAN  U UTE

HOURS MKG N SEX TIME D0E CAT NO.

(A-1) L682

PAGE A-23

[PAGE 246]
SUSG SSN  SUSSONT

NHSD D D SID CA BOD  BORYU°O REREE R UA HE ST E BUTE

NOSCD D  SCD CAL BODE  BO US R2 S MOI2 HE UET E BUTE

BO  U U8°G RESE RSE PE PRO32 HEA   UTE

NOG H   E NO D C RY RERE P PO4B HE BE  BUTE

PAGE A-236

[PAGE 247]
SUSD SSON - SUSONT

(A1) 992 3

NHAHSSC  ONN C CAL BOY CBOUY U3 RESE EE PS PO42 HEASN 8E UE UTE

NCD D H CCD CAL BOY  BOUU TU° RE P PS POA42 HEASE  E UTE

PAGE A-

[PAGE 248]
US    S

ADOO   AL SD CUE C CUE B P PONS P02

96 GS     HT BESOS PS PTE SHD  O OCUE

SHL L O OCUE

THSSRD PDHSING

(W-1) L092

XO OYS  NOYS ON YYS

[PAGE 249]
SLIL S  - LOSE

96 SH    YS ADPOR BE BL BMALRBESS PNS POTE SHU C  CUE

NOHS GES RESS S PS PM2O

APRNL PER RMAL GD LH C CUS

06 RBENS PRS POTE PRE DS SDY

(W-1) L092

一

[PAGE 250]
SUSS C OOE  CUSOS

6 WAE M      YS AP RA BL BAL RES RP PUTE SHU C  CUE

NHANGS GES RESS S PS MM2O

SHUD LH O DCUE

APNL LE AL S   CUE

PRE OSEE OAY 06 RESS PS PUTE

(W-1) L092

一

[PAGE 251]
SUS  C - CUSS

APOD RD B BMAL SHDD L OD CUE

AP  B AL GSH  C CUE

API   AL SHD  C LCUE

APO R L AL SHU SH C OGUE

APO M L AL SHD CO COUE

AP R  AMAL SHD  C CUE

E\*0 ELL 67

OBSONS

(W-1) 65L2

[PAGE 252]
INME ME HGE ES    GH BS PSS PS P88

SHL H  CCUE

ES   OHT 6 PS PS P96

SD   OUE SIG OLY HEA E T UTE

TNC  ITY CGH L C CUE

PRAE DSSE DSE-AY ABOO   MAL HE BE BLE RTE

2°°36

[PAGE 253]
APRSL PRSL RAL SO N MS A  SNG PAN E R    EY RESD T SSIND

RED MD MAMITY TOXICS DXISC DING OBOL CONL ONN

NOUN  OEE N RESU S  SUL GR  RACT EASSLY HILED

(W-1) (

PAGE A-

[PAGE 256]
I S HES SE  IEF NOE   IST BO OUUY TUPL 87 RESS PS MOTE HEA E UE BUTE

IO E S NHH IE NO N WST MSST BOD  UY U° 60 RESNS PS MOTE HEAT  E BUTE

A I N S  IEP BOO Y U UZ 99 RESS MMOTE HEA/ BE BUTE BUTE

PRE DSOSEE SDOY NOL C CL ST BEHA  HANC NEML L NCMAL BO BUYBU RESREE PS PMO4 HEA S E UTE

(A-1) 92

[PAGE 257]
S A O - AO

SLIIO      ON SL 1A NOE E E DOT

BOD 83 RE BPE PR P36 MEAUI E UTE BUTE

BODE CUU80Z BE PE P P3O HEA  UE BUTE

EE E UTE UTE 9E PES P POTE BOD CGU83

HEASU BUE UTE UTE 72RES PEG POUTE BOD U79 NOD   ST

(A-1) £963

PAGE A

[PAGE 258]
SOL  O  U0%

NEUOLAL BELM CLA RRMAL NOOHOTA IA

BOR  UY U RES RES PS PO3 HEA   UTE

NO SE I ED NA I N D DD CC O HE DRTO BOE C UTU°S RESS PS POTE

HEA T UE BUTE 306
RE RS MOTE BODE  BOU TU3

BO  E3UY TUBORE REE P PO42HEA BUET BU BUTE

(A-I) £962 2

PHL  CIOL  C CIATS

PAGE

[PAGE 259]
DILUNT SUSOSUSOD

NHHS CE HCS CE CAL BOU  U TU.G RS RS PS POG6 HE  EUE UTE

NH C CC SCED CA BOY OUYTUT RE E PS PMO48 HE ET EUE BUTE

NHCS C CCS CCE CAL BODE  OUY TU4 R MRES M MUG9O HE   UTE

PRE DOSE DREOSY NE NL NAL BEHAN ANCE. CALM

BO  UTUGRE ESS PS O42HEA   RUTE

OOBSONS

(A-1) 862

3WI1 HOUURS \*1M M/KG 53X

PAGE

[PAGE 260]
AOITOL CL CION

ANOIOL CIL CION

SUSO O USONT

NHCE   C CAL BOC BOU UO RE PE PRS PO36 HEAU 1 UT UTE

NH  C SCE CALBOE  OU U°SRESRES PR PO5HEAB  U BUITE

NHC   SC CAL BO  3UU.B8 BE PBEE PS PTB HENUE BE BE BUTE

OBBSONS

PAGE A-:

[PAGE 261]
SO O- O - 0%

NHS   SNC CAL BODY UU84 RESRE RS POT7 HEA/G E UTG UTE

NH  N CE CAL BO C OU UBPS BE P POG9 HEA  UE BUTE

NO  N CC CN BOE  BOU UU8S 9. PIOE P PO9G HEA EE  UTE

ーー BOE  O UB RBE RENS PN POT7B E/NUE BEE RUTE UTE

HOURS K / EX 1M O0E ·ON

[PAGE 262]
O - O  0O0

ANDIOLL CL ION

NC N SC CAL R PR PR84 HEAUE BEEE BUTE BUTE

NO N C CAL BOD Y BORUBS RES R P OB84 AEANNEE BEE BUE BTEE

N   C  CAL BOEY Y ETL P PT72 HEAUIE EE TEE DUTEE

(A^-1) +662

SS  N   C S

[PAGE 263]
8001 SOL- OOO  OOO

NO   C CAL BODY CU8B8 R R PRS PYTG EAUG PG BUEG BUTE

NU   C  CAL BOBY CCUT RE RA PR PRAI48 HEA E UE UTE

N    CL BO0 UU8.4 PBE P PRS P6G6 E  UUE UTE

PRE DSE DOYY BEHAL, ANCE. NEAL  MAL SLI  SCH SLI LTSH CALM ALERT

P P6 HEA/N BE B TIE

OSSONS

A 5662

IOURS 3053 MKG °ON X39

PAGE A-

[PAGE 264]
SO0E OOOOO

NC  C CS CAL BOR OUTU°4 RSPES PS POO HEU E U BUTE

NS  S SS CAL BODE Y RUY UBE REE PRS 48 HEA   OUTE

NHCD  CD CCD CALBO  BOU UURRE RS MHE  E NTE

OBONS

[PAGE 265]
NHSER  E SE  SN HEE  DET DECT BO  OUY TUL 06 RESOS P POTE HEA  S UTE

N I N S N IN B0  UU°G RESS  PS P0O02 HEA  TE UTE

PR DRS PRESY NEUAL MLL NMAL BENA G GNAN B0Y  UY UG RES RESS PS PMOT7S HEATE 1 E BRUTE

3111 HOUURS
1M
3SC0.ON M/KG S3X

6662

PAGE A-

[PAGE 266]
%001 SOL- GOO  OO

NHOS   S IC BO  ORU UL REREE P PITA42 HEA  U UTE

HEAU  UT BUTEE 54 RES PS PRRUOTE BO CY RUYU°O

NEAL NL NA BEH  BE. NOOL C  IST

HEAB B BIS BIE: 84 RESS P PUOTE BOE Y BOUY UB

TI1ME HUOURS iM D00E MKG °ON

PAGE A

[PAGE 267]
(A-1) 6662

N  H  IP TE BO  O 8B 06 BESIE P PUTE HEA  E UI1TE

SO TER  OTER

NEUOG LG MALBEHAA AANCBODE  OPUBPB78 RES P POITEHEANS B U BUTE

PAGE A

[PAGE 268]
YIL CGL L ION

SO - O  O0%

NHC   C CAI BOY UUBO RES RES PRS PO4O0 HEAUI 1 T UI3

NHS  TNN CES CAL BOY MUU°O RERBE P Z2 HEANE EE BT BTE

HEAN BS BE BUIE RES P POTE 22 BOD BOPY8°O NI O S O N

[PAGE 269]
1000% -  O

N   ST BEHS MAL OYL AN RESS E80S 54 HE P  UTE E   TE

N/E E  TE 36 EA P P UTE RBESE EU8°S S

PAGE A

[PAGE 270]
%001 SOLL OO-OLOO

HEU EE UE BUTE 27 RES PS POTE BOOUYTUL

BOD OUY TU84 RE ES POS PMO4 HEU T BU BRUTE

HENUUT  EU BRUTE RESOS S PMOTE 3 BOD  T TUZ

HE/T E ET UTE RESON P PMOUTE 09 B0 UU°4 AAG ON C SUN

(A-1) IEOS

PAGE A

[PAGE 271]
SISOND DN - DISEN

OI S   TN  IME FLENINS  S ON ELILEE REE RXEN FXTS BS PUPLS PED ENNSIS

BREG DGIEP ABORML NMAL ESY RRRPOIE UNCSOUS DISOG IME AEER

PRAE DSE DSATY NEL SSL  UAAL BEMAA NC CE 13100

H/E E PDUE DUIE 09 PRES PUE PRUTE COI C CO. CSS

(A-1),BS0S

[PAGE 272]
HEAU U U BUTE 90 RESS PS PMUTE BO  TU TU.2 NOSL C  OST

OTHM   MAL COE EL ENT TO RS R R TEO PLLG  G POR RIGG FGGFGIR EX E R E RIND FLERS RS RAL DE PI P  PENT PROAD D S  OAND PUPR R RMAL PUPTE S STED AAKE

PU  S  PN SIRE RE T TCON NO MN MW MS HE TK   UNT BOUY  BOUY TU.2 RS OS PS PM

[PAGE 273]
NEROL   RMAL O O E MAL BOE  RU° 6 REE PR POG HEAS E U UITE

NHOSD   S CA BODE  BORUYTUS 60 BENI P PT9 HEASBE BTEE BUTE BUITE

HE/U  UI UTE 99 RESS PES PUTE BOE  BOU8S

BOE CYBOTUE RE RBE R POS5 HEANNEE EE PE PUTEE

(A-I) ASSAN

PAGE

[PAGE 274]
INN OHN S  IF NOL C C IS BOE  OUUY TUO 9€ RESOS PS MOTE HEAS S EUE BUTE

O  S  IN NO C S OS BO UY TU7B 87 RESS PS PMUTE HEE E E UTE

NO E S N I NO C C R BOY  UY TU°B 87 RESS PS PMOTE HE  E UTE

NOD   ST B0OL  BU TU3 RERE PS PMO42 HE  U UTE

PAGE

[PAGE 275]
NEUOS AL NMAL BEHO GO CGGG GAL NOL CL  ST

N  ES N N NE NOD CD C ORY BOGM  UY T78 22 RES PS PMOTE HEAN E E BUTE

[PAGE 276]
SUSND OUSNT

NOO C       ON NO C  5T

HEAB BS BUTE BUTE 09 RES R PROTIE BOD C BOUUY UB

HEASS I BUS BUTE 09 RESS S PUOUTE BO Y BORTUBT

APIL C O CL OION

NO  S, ST B0 Y BD REEPEE PSE PO42 HEG B UIG BUTE

PAGE A-.

[PAGE 277]
SUSNDDUODDNT

CHCED D CH CD ICA B0O  OU TU. RERES PS POG6 HEASN  BT BUITE

! NOHIO C    C  ION SURS S O S  SOIN SO  D OID D D  IN DDURY B0OY  OUYTBU°.4

RRESRTE HEN  EUE BUTE

PRSE S SRDOY) NEAL  MAL MOAIVE CALM

CAML S LL LAN BOY Y OUY TU HEE UEE  UTE

HOURS M/KG N SX 1ME 3000 CATTT NO.

[PAGE 278]
(A-1) 260s

BODY OUUO. RE P PUG0 HENE EE E BTE

PAGE A-2

[PAGE 279]
BOY  UY TU.G RE8EE M48 HEN  U BNUTE

B  UY TU. RE3SR  M10 HE  UE BUTE

B0Y EUY TU.4 RE E M OG HE BE UTE RNTE

B0O Y UY TUS RES ES  P54 HEA T T BUTE

(N-1) v015

PAGE A-.

[PAGE 280]
N  OH S N I NOIE SE IST BOR  UY TU4 RES  P P102 HEET  RNUTE

NO  C  IN NOD M MW MMST BO  U TUS REE P PSOS HEO E UTE

(W-1) 544

[PAGE 281]
S DER - ER

N O   E NOS M WS MIST BO  3OU U4 RESRE R PTN HEAS S UTE BUTE

I     E NOIO MN WR MSST B0O  BORUU TU·3 P6  PRS PG HEAG G BUUTG BUTE

EG 1G UG UIE 54
RENS RS PRUTE BOD  BO4

PRE DSEE SDAY NEONL   RMAL BAA  OCE NOD CM S S

BOE  ORAUUT. HEA/ E UTE BUTE

(A-In EsIs

6

PAGE A-2

[PAGE 282]
SI OE - ER

PRAE DSEE DSAYI NELG  MAL AAL GIC CE CALM

HEA BEE E UTE 442 BEG RPUITE BOLEY  BRUT NOE N CS ST

SO TIE - TER

NOS   ST BE  BEB.T PES PR PS54 EA E BTE LUTE

NOS D C CRY BE E PRR PI54 EA SE UTE UTE

(-I)EV

[PAGE 283]
NOSD    CA BOL  BOY U°. BEE PEE PNE POT4 HEABUE UTS BUTE

NO  N   CN BOY RUUUP E PEAE R POT3M EASBSB BUTE UTEE

NCD  P C CAL BO  OUB 9E6 BENIE R STTOT3 EAUEE UTE UTE

EA BEE TEE UITE 99 RESS PUN PAOTE BOEY ORU8°Z

NIIT ION   ION NOL E S   ED NOSDG O  RY 06 PBESS PROS POUTE HGTE UT BUTE

PAGE A-

[PAGE 284]
1000% SL- P - O0

N    N NOSD D ARYE RY BOE  B 9€ RBES RUTE HEA  UUE UTE

PAGE

[PAGE 285]
NOS IS N  ICA BO  BOUUUG RES RE  PO3O HEA UTEE AUE UTE

HE  R BUTE 36 RRESS  MOTE BODE  BY TU8S

N  OE SNE N E NOG G O SRY B0OD  Y 84 9€ RES  OTIE HE E UE UTE

TME HOURS °T G 305E M/KG CAT NO AN SEX

PAGE A-

[PAGE 286]
NI O S  NNOD C  STBOE  U T88RES RES PS P48HE BE  BRTE

NO M  IST BO Y ORUY TU8E RES RES P PMO42 HE BTE  BTE

[PAGE 287]
NEAL L NAL RBEO  COL C MAL ANEL TER

BODY Y BO U8Z RERE PS PO4B HEA E BU BUTE

BOE  OUU UUO RESEOS PS PMO2T HENS E U UTE

SN NHIG IEP REO C RUCION BOE Y BOUY TU8.S 9€ RESS  MUTE HEA E UTE BUTE

[PAGE 288]
NO CE S E F NOD ML SL IST B0O  OU TU8S 22 RES P UTE 3GA/ST8 6 HAAY RRATE

HEA A U UTE 84 RESO M MOTE BOO  OUUY TU8°B

PRE DSS DPRSDOSY NEAL  NAL BEHAL  OCE. CALM

AOO A O. ST B0OY  U TU. RESES PS P48 HEA   BUTE

OBBON

KOUURSUR M/KG N SEX 3E T· D0E CATT NO.

[PAGE 289]
SUS USUENT

HEA  S UITE 09 NO   C CA E PR TE BOD Y ORU4

NHS   SC CAL HEA  UT UTE

NOS    CA HEA  U UTE

NO   S CAL B  OU°O HEASG G TG UTE

(A-I) [6-8598

OBSSIONS

IME HOURS T· K. 300E M/KG CATTT NO. AD SEX

6

PAGE A-:

[PAGE 290]
NEA  NAL BEHNAA ARIL RHNAOCE HET T OET OECT REE P PMO4

N  S  P NO M  5T RESRE PS POT HEU G UE BUTE

N  OHE S N NP NO CD C CRY RESREO PS POT HE   TE

1 NOH  DN SHH H HY HET DET DRECT NOSO M AS MST RE3PRES PRNS PPO8 HE UTE RT RUTE

PRE DSSE DSDDAY) NEUAL AL NAL BEHAL BOLHNC NO S  O5T RE5PRES PS PRPO4 HE3E T BNTE

[PAGE 291]
SIUR U UU U AL AP  AES     UMAL APO RA  MAL R RES S OG

TNCI C LIITY RAO   M E   EY

B  R   HT P NS PRS P69

CAPP  TTR  TIHY RTHY EYE Y      EYS EVEUN UUEL UL UE U UAL TN A AIITY

APO RAA BA BAIMAL

APRO RA  AL SD LH O LCUE

PRE DOSE DSDOAY APPT     HT ADOO BO BNL BAL BESBES RS PT8

PAGE A-

[PAGE 292]
PRE DSE DSDAY PUP RL    SACL EXCIE  ING APIO BPAIR L BAIAL RESRE R POT HES BUTE BT BUTE

-1 211

SOL DER  DOER REREO RS OT HEA/E U RUTE

[PAGE 293]
S    AI

AR B B BIMAL

SU  O CUE P PE P PO48

SHU   CUE APR ARE NA BAMAL

ABRR BRBE  BMAL

PRE DSE DSDAY ES   GHT BE P O84 HEA I U BUTE

0°0
0S8
10·0

[PAGE 294]
COY S M CME AO TY TY AIE S10E E 0X CIR ITY SLTOTS TORS NG  GE

GAT H.O OCUE VO BAAE BNALE BAL

GAE 1H17 O UE VOO  N AL

SO T O UE AO BAN BL BMAL

GH H O  UE VON BANARG BNNL BNAL

SHUS SSLY ADOO ROAE E OMAL

(N-I) 201

PAGE A-2

[PAGE 295]
OBSERVATIONS

RESPIRATIONS PER MINUTE 90 PRE-DO5E (SAME DAY) DOSED ONE DAY AFTER NEXT LOWER DOSING

ALONE IN SHUTTLE BOXSITTING QUIETLY

ALONE IN SHUTTLE BOX SHUTTLED SPONTANEOUSLY SOME COARSE PURRING

ALONE IN SHUTTLE BOX SOME COARSE PURRING SHUTTLED SPONTANEOUSLY

ALONE IN SHUTTLE BOXSLIGHT TREMORS

RESPIRATIONS PER MINUTE WAS FED 10 BISCUITS ALONE IN SHUTTLE BOX

SOLUTION - 4.O% ASCORBIC ACID

PAPAGGE

6

[PAGE 296]
OBSERVATIONS

ALONE IN SHUTTLE BOX SHUTTLED SPONTANEOUSLY 13 TIMES 7 MINUTE TRIED TO LEAP THRU TRANSPARENT CAGE WALL 2X IN 5 MINOTES

ALONE IN SHUTTLE BOX SHUTTLED SPONTANEOUSLY 1I TIMES / MINUTE TRIED TO LEAP THRU TRANSPARENT CAGE WALL IZX IN 3 MINOTES

ALONE IN SHUTTLEI BOX SHUTTLED SPONTANEOUSLY 9 TIMES / MINUTE TRIED TO LEAP THRU TRANSPARENT CAGE WALL SX IN Z MINO.ES

APPEARANCE AND BEHAVIOR NORMAL ALONE IN SHUTTLE BOX SHUTTLED SPONTANEOUSLY 16 TIMES / MINOTE

SOLUTION - 4.O% ASCORBIC ACID

PAAP

[PAGE 297]
APOE A AL AL ES RY Y    EYS 84 PBESS PUS POTE

SOL TOO - IER

6 RESS PS POTE NHANNGGS GES

0

[PAGE 298]
BEE SN SIP OP S SINP SR D SING PALUL CURE YG OG ONIE

AR O     O    DE LYMG OOGAMEN

OTOS PD PR O  OVAL RESU ESSIND REISSUVL SSVL L SSSL

SOU  O OCK DIUES   R CCE   NCKS BLLVD HVVD ND

FOMAUR SORMAULT PAUL CPL CRE

(N-I) 0061

[PAGE 299]
SOLI S LLINE

INNC AGE HA O SN  MOD AP BA B BAMAL

CLLG G  LT 7 MOGY S SSH ATEGGG SGGD SHLY FAU BLSH

RESG H  GISH APP  E D   UNO SITG TLY INN C GE

TRE L  LGS ST  O ET WA  D  IAGE RESG G VED GRROP SRTRONG LYGG G SDE

HOURS M M/KG NUER 3IME I DE MOKKEY

PAGE A-L

[PAGE 300]
SO TE - TER

BETI LIG PEITY PALIINL ING

PARL PAURL PURE SITLG OUTLY

SLOT.HH1A ES PS P9

SH CSS CS F CD CEES

S GNS GNN C N GIES

PRAE DDS DSO-DAYI ADPM AER L MAL G P  PNIT10

OBBSONS

W-1i 0061

PAGE :

[PAGE 301]
SOE % - IONE

TOX BG BGIG BN BTTE

HERD OAROP CROSGUNG PURE CLUURE PAAL PIAL

SO YS  SY

PAGE A-2

[PAGE 302]
EXC   UU EL

SKD DS S SRD ED

HICHS PALL PURL PURE

DECD EDEITY DEC AC ITY PALUL PURL PURE

02
622
0\*t

[PAGE 303]
APR D BL BPAL

FAD NS NNU U UED PLUL PUL PRE SING UG UGG UG UND

PALL PLL PRE SING ANG ANG AG AN

N M M M  EYS LYG SG SID

IN ME L R E EYS LYGG G SD

MUC  YV V VOCK SID HOD D  D  HD OOR USUCE   R  CL CKS BETL LN NLS

PRE DOS DRSDOY APO  AL AL R RE P PO

(W-1) 0012 8

PAGE A-

[PAGE 304]
S    CID

ELE AES  ALS MO S  SNDS MOA A AL

SLIUL CUL URE

S G GN   GES

RO E G E  UES DOL R CR B TLY

S GNS G   G UES

PREE DSE PSODAYL PBESS PS P10

(UGNNE) L

[PAGE 305]
SOC   OID

TOXSSDICC ING AT  D   OGS REOIR I OITY

BEL TLS  PHTLLS PALRL PL PRE SITGG DULLY

(UUS)

PAGE A-:

[PAGE 306]
TOXG S BNN BIG TE

GRAING RG RG RACT RESC C C SOCK RESUU S  UNO PHPOTOIIA

GENLLE ON     II D ION COG O  O OG   OG DONE RE5 E S 36

ALO E  OX INCING

PRE DDOS DRSDOY PUP      N IHT RRES

[PAGE 307]
BONNE N NRR SI   K

NNMM M M M  MLI (M FONNEY MNY MES E INUD M PD NND D MK ABA MAPHE WAN      N HYS

LOC S  A SRD SHUC C OC OCK SLT SON LEAAY MRY MOLED

HOURS M/KG NUER TME IM 305E MONNKEY

PAGE. A-

[PAGE 308]
9L RESS MNS MOTE SHU   CUE

SECOU SINS

SO %  S O    INE

WA E E  S   AYS AP BA BA BPAAL PU     : N HT KRE RES PUNS PUOT84 HEA RUE UTE

SHUO  C CUE AP  B MAL RES S P PM102

SHUDD  CCUE RESS S S M02 HE  U UTE

SHUO S  SCUE AP  B MAL

HOURS •1M G D0E UUK MONKEY

(W-1) 862

SHDD  G CUE AP PA A BAAL

[PAGE 309]
SAII

6