Document QJzjo6nXpGMx0qZkvMZvdpRJ6

>r. SrtdusUiai BIO-TEST XaiviatoMM, 9*C. k^ rfe* i ( I* MONTAOI IOA Hooatoc*. tuNoa ;; February 11. 1961 I ; if fc < * r- Mr. Don G. Fowltr, Project Manager Deed Induatrice Aaeociation 292 Madiaon Avenue New York 17. New York Dear Mr. Fowler: We are endoalng our propoaal for range-finding toxicity teate on materlala in which your aaaociatioa ia inter a a ted. The coat of thcae teata will be $600.00 per material taated. If you have any qucationa on theae procedurea pleaae do not hccitate to call ua. Thank you again for permitting ua to aubmit our propoaala. JCC:par r-n^i. Sincerely, ) 2. Za/auAv J. C. Calandrn Director i $ I I iii w p w g y "4 st-^la/w: 'J>*'s $. tr Ur" Vs & Sr It' *--\tr [ i Al Z f A > , -i 'i ;! ' 'i.a . `',1 9 ... J . ..3 a '.1 RANGE-TINDING TOXICITY STUDIES L Ran^e-Finding Acuta Ortl Toxicity - Albino lUti A. Acute Oral Toxicity - Albino Rats Healthy, young albino rata of the Sprague-Dawley strain with an average body weight of approximately 100 grams are used as test animals. The rats are divided Into groups of four animals each for dosing purposes. All animals used are Vept under observation for five days prior to'-eperlmental use, during which period they are checked for general physical well-being and homogeneity. The animals are housed Individually In stock cages and permitted a standard laboratory rat diet* plus water ad libitum until 16 hours Immediately prior to oral intubation. On the morning of the first test day. after a 16-hour fast (water permitted}, the selected dote groups of four rats each are intubated with previously calculated dotee of the test material. All doses are administered directly Into the stomachs of the test rats using a hypo dermic syringe equipped with a ball-pointed intubating needle. * Rockland Rat Diet, Rockland Farms. New City, New York. 0 W LUlw-wy 11 w >iti a^'iwia njsme im,^.y. . "-'V. v. . j* * , f s- >. 5-r- N 184.01 z Following oral administration of the test material, the rats are returned to their stock cages end observed for the succeeding Id days. AU significant reactions And mortalities which occur are recorded. Particular attention ia given to the observation during the flrtt four hour* After intubation. At the end of the obeervatioa period. All data are collected and the range-finding acute oral mean lethal doee (RFLDjg) of the test material ia calculated using the techniques of Weil*, Thompson**, and Thompson and Well***. --tfc * Well, Carrol S,: Tables for Convenient Calculation of Median* Effective Dose (LDj q or EDj q ) and Instructions In Their Us*. Biometrics, Sspt. 1952. ** Thompson, William R. 1 Use of Moving Averages and Interpolation to Estimate Median-Effective Dose. Bact. Rev., Nov. 1947, * Thompson, William R. and Well, Carrol S, t <5n the Construction of Tables for Moving Average Interpolation, Biometrics. March 1952. 9 21. Range-Finding Acute Percutaneous Toxicity - Albtno Rabbit* Young adult. New Zealand etraln male albino rabbit* are employed a* teat animal*. The body weight* of the rabbit* uied range from two to three kilogram*. All rabbit* ar maintained under ob*rvatloa in 4 the laboratory for five day* prior to testing. During the pre-te*t period i the animal* are examined with respect to their general health and suitability a* t**t animal*. The rabbit* are housed in stock cage* and maintained on the standard laboratory rabbit ration*. Food and water are permitted ad . -i libitum. Twenty-four hour* prior to the dermal application*, the back* of the rabbit* ar* shaved free of hair with electric dipper*. The animal* ar* then returned to their stock cage* to await testing on the following day. The 24-hour waiting period allowed recovery of the stratum cor ncum from the disturbance which accompanies the dipping procedure and dso permit* hsailng of any microtcoplc abrasion* possibly pro duced during the process. On the teeting day, the dermal dose for each test rabbit is weighed accurately on an analytical balanca. Applications of the test material ara mad* in solid form on the exposure sites of the rabbits. Three i 4 dose levels employing four rabbit* each ara used. After each application, % the exposure site i* covered by wrapping the trunk of the animal with an 1 ' * Rockland Rabbit Diet, Rockland Farma, New City, New York i `J .1 *: t ?f -' *. i. i >fc: s s>. f: . 1 l -J '-I 't 1 i 7. 3! 4 impervious plastic sheeting which U securely uped in piece. This pie Stic wrep insures Intimate contact of epidermis end test material. The test materiel remains in contact with the skin for 24 hours. Be havioral reactions ere observed end recorded during the contact period, after which the plastic sheeting is removed from each test rabbit. The exposure sites are examined for local reactions and the animals returned to their stock cages. At this time it will also be possible to obtain an estimate of the primary skin irritating properties of the test materials. Observations for mortality, local reactions, and behavioral abnormalities are continued for a total of 14 days following the skin applications. At the end of the observation period, all data were collected and the range-finding acute percutaneous mean lethal dose (RFLDj q ) of the test material is calculated using the techniques of Well*, Thompson**, and Thompson and Weil***. A * See footnote *, page X See footnote *, page X *** See footnote ***, page X |i<ippwMWp^ypp i * r< -J -< ) t .. S * *<v , 'j tl` -r-bt t'r ' ,'$ .:V*S v>-'- 5 QI. Range-Finding Acul Inhalation Toxicity - Albino Rat* Young, adult albino nti of the Spragus-Dcwlsy strain were employed a* the test animal*. Pre- end post-test cere end obeervetlon of theee rete ere eeeentieUy the eeme ee thet elreedy deecribed for rete ueed in the range-finding ecute orel toxicity teet ( eee p. 1). However, enimele ueed , for inheletion expoeure were not feeted prior to teiting. Five group* of four ret* eech ere utilised in the *tudy of the teat materiel. The everege body weight of the teet enimel* ere approximately 2S0 gram*. The Inheletion exposure* ere conducted in e multiple inhalation chamber designed for eimultaneou* expoeure to atmoephere saturated and fractionally saturated with the test material, e. g, 1/2 saturated, 1/4 saturated. 1/8 saturated and 1/16 saturated at room temperature (2S*C). The teat is designed to run for a four-hour period. At the end of the exposure period, the rats are observed for the succeeding 14 days. < .1 i 1 . IkS. A -1 6 IV. Range-Finding Eye Irritation Tot - Albino Rabbits A group of flvs Albino r>bblu r used to evaluate lbs eys lrrlutlng properties of the test material. Young adult, Nw Zealand strain animals are selected lor the test. The test method employed Is patterned alter that ot Dralse et al*. Exactly 50 mg of solid teat material la Instilled Into the conjunctival sac of the right eye of each test rabbit in a group of five animals. The left eye of each animal serves as a scoring control. One, 24, 48, 72, 9b hours, and 7 days following the initial Instillations, the cornea. Iris, and palpebral conjunctiva are examined Individually and graded for Irritation and Injury according to a standard scoring system*. The maximum possible score at any one examination and scoring period is 110 points which indicates maximal irritation and damage to all three ocular tissues. Zero score indicates no Irritation whatever. After the completion of the test, the scores are analysed and a descriptive eye Irritation rating is assigned to the test material. The criteria used for assignment of the descriptive rating are the frequency, the extent, and the persistence of irritation or damage which occurred to the three ocular tissues. In the classification system used, special emphasis is placed upon irritation or damage to the cornea. Corres pondingly less stress is placed upon conjunctival and iridlal effects. "Methods for the Study of Irritation and Toxicity of Substances Applied Topically to the Skin and Mucous Membranes", Dralse, John H., Woodard Geoffrey, and Calvary, Herbert O,, J, Pharm. It Exp. Ther. 82, 4, December 1946, 7 Th* daacrlptiv* rating aulgned ar* on* of th following, aacb of fI which charact*rix* * particular level of ophthalmic irritation and damage; j Non-Irritating Practically Non-Irritating Minimally Irritating Mildly Irritating Moderately Irritating S*vrly Irritating Extremely Irritating Maximally Irritating i 5 ; * 1 Jj 4 r - ; /* i>a -A-^5tL * PILOT STUDY 4 Effect of lead acetate as a radioprotective agent Materials and Methods Wistar rat* of both sexes weighing 100 gma. and on an ad libitum diet were used. Animals were divided into 6 groupa at follows: Croup 1 100 mc/hg A (4) 200 mp/h| B (4) 2S0 mg/hg C (4) Croup Q A (4) B(4) C (4) Croup in A (4) B (4) C (4) Animals 12 12 12 N*a Pb{ AcU AgNO, Animal* were Injected with NaCi (negative control). Pb (Ac)g (experiment) or AgNOj (heavy metal control) in aterile distilled water. Route of adminiatration wae I. P. and dote* were divided into SO mg/hg . equal doee*. Following the laet doee. 400 r of whole body X-lrradiation wa* administered 24 hour* later. Oote was corrected for skin level and no shielding of scatter was employed. All animal* were returned to cages for routine care. At death, complete autopsy examination* were carried out including microscopic sections. J * i Result* Table t shows the death pattern following the X-ray exposure in all groupa. i_ i 1 *~ .................. .............. - Day* Poat X- ray 1234 Group I A l 01 2 B1 1 0 2 V * V*. .;: lf .1 "' j : v C -r F V*. * ' fc>* ' ,: ..- r ^f t . ' .> ^^ " fci\A'V.- .-? fetor--j . ^ -a * -.ii --^a r? j 3>.v ".'9 Is.'" * ` g r""' r , tg $ * 5 C2 Group U A 2 B1 C2 Group III A 1 B0 C0 Total 10 Cumulative % 27.7 Expected % 30 1 0 0 0 '0 0 0 2 33.2 . 1 0 0 0 0 0 0 2 38.7 . 0 1 0 0 1 0 1 7 38. ' ,' V -Y, t H V *'*!* -* ' 1 vs .1 Table n Croup 1: A B C Croup 11 A B C Croup 111 A B C Expected Pathological Obeervetion# Hamorrhagaa Cytolyala 4444 4444 4444 444 4444 44 4444 4 4444 4444 4444 4*4 444 444 444 4444 444 Bact. Inf. 4 TtUt ! a howa that the expected mortality rata wu not altered by tha agenta admlniatered la this atudy. Hcwavar, Tabla U doaa indicate that tha organ and aaroaal aurfaca hamorrhagaa wara aignificantly raducad in Croup U B and C. Lymphocytolyala and bactaramia with colonktalion m not changad In thaaa groupa. Conclualon A pilot atudy to teat tha radioprotective action o( inorganic ionlzad taad waa carriad out in Wiatar rata. Tha cumulatlva mortality rata waa not altarad. Howavar, tha frequency and intanaity of hamor rhagaa in organa and aaroaal aurfacaa in tha laad acatata traatad animal a waa laaa than in tha control#. Further atudiaa ara indicated to mora clearly aval* uate thia finding. Tha numbera of animal# uaad and tha LD 50 irradiation doaa exposure ahould be modified In future research. Barnard Mj Wagner. M. D. Profeeaor and Chairman Department of Pathology -. fVVki