Document v1eLRZQrBK7Qqmpa5j0Ewy128

Scientific Associates 0 INC. 6200 S. LINDBERGH. ST. LOUIS. MO. 63123 TELEPHONE: 314-487-6776 SUBJECT Contact Sensitization Study in Guinea Pigs July 28, 1977 SAMPLE DESIGNATION Alfonic 1412-A Ether Sulfate Sample No. 8126J (3/28/77) S. A. Number 228618 SUBMITTED BY: Continental Oil Company P.O. Box 1267 Ponca City, Oklahoma 74601 SUMMARY AND CONCLUSION: Alfonic 1412-A Ether Sulfate, Sample No. 8126J (3/23/77), was assayed for allergenic potential utilizing the Guinea Pig Maximization Test of Magnusson and Klegtnan (J. Inv. Derma., _52: 1, page 268-276, 1968). An initial range finding study was performed on twelve (12) guinea pigs to establish suitable concentrations of test material for intracutaneous injection and for topical application in the subsequent maximization test. The second study was performed for definitive testing of sensitization, utilizing ten (10) guinea pigs for the test group and four (4) guinea pigs for controls. No signs of sensitization were observed in any of the test animals at twenty-four or forty-eight hours following application of the initial and final challenge patches. Histopathological examination of skin tissues from each of the ten test animals of the maximization study showed no changes in any indicative of allergic reaction. Alfonic 1412-A Ether Sulfate, Sample No. 8126J (3/28/77), was found not to produce sensitization of guinea pigs under the extreme conditions of the test. EXPERIMENTAL PROCEDURE: Alfonic 1412-A Ether Sulfate, Sample No. 8126J (3/28/77), was assayed for allergenic potential utilizing the Guinea Pig Maximization Test of Magnusson and Klegman (J. Inv. Derma., S2: 1, page 268-276, 1968). VVV 000014485 9 Scientific Associates, me. 6200 S. LINDBERGH. ST. LOUIS. MO. 63123 TELEPHONE: 314.4S7.6776 EXPERIMENTAL PROCEDURE: (continued) -2- An outline of the procedure as employed for the testing of this compound is as follows: Healthy young female albino guinea pigs of the Hartley strain, purchased from a commercial breeder, were allowed a period of twenty-eight days to adjust to laboratory environment. The animals, which weighed 250 to 300 grams at the time of receival were group housed (four to six per cage) in wire-bottomed cages, elevated above the droppings, with Purina Guinea Pig Chow and tap water provided ad-libitum. Following the acclimation period, the animals were randomly selected from the stock supply, as required for each step of the experimental procedure. Preliminary Range Finding Test: Twelve animals were used in a range finding study to establish a suitable concentration of test compound for intracutaneous injection and for topical application in the subsequent maximization test. Eight of the animals were injected intracutaneously with Alfonic 1412-A Ether Sulfate, Sample No. 8126J (3/28/77),in concentrations ranging from 0.001% to 5.00% w/w in Sterile Water for Injection, U.S.P. The diluent was used as a control injection (0.1 ml. volume/site) on the opposite side of the test dosage site of each animal. The four remaining animals received the test compound in concentrations ranging from 10.00% w/w (aqueous) to undiluted under an occlusive gauze patch on the clipped skin of the back. Maximization Test: (April 13, 1977) Based upon the results obtained in the preliminary test, ten guinea pigs weighing 410 to 566 grams were administered intracutaneously an induction injection at the selected concentration of 1.00% w/w of Alfonic 1412-A Ether Sulfate, Sample No. 8126J (3/28/77) in Sterile Water for Injection, U.S.P. Paired injections were made within a 2 X 4 cm. closely clipped area of the shoulder region of each animal according to the following scheme: WV 000014486 S. A. Number 228618 This it confidential report which may not he used in any form of advertising without written permission. Scientific Associates INC. 6200 S. LINDBERGH. ST. LOUIS. MO. 63123 TELEPHONE: 314-487-6776 EXPERIMENTAL PROCEDURE: (continued) 3- Maximization Teat: (April 13, 1977) (continued) Dorsal Aspect Left Side of Induction Site Induction Site______ Right Side of Induction Site 0,1 ml, of Freund's Adjuvant Anterior Portion 0,1 ml, of Freund's Adjuvant 0,1 ml, of 1,00% w/w aqueous Alfonic 1412-A Ether Sulfate Sample No. 8126J (3/28/77) Middle Portion 0.1 ml. of 1.00% w/w aqueous Alfonic 1412-A Ether Sulfate Sample No. 8126J (3/28/77) 0.1 ml. of 1.00% w/w Alfonic 1412-A Ether Sulfate in Adju vant and water emulsion (50% v/v). Posterior Portion 0.1 ml. of 1.00% w/w Alfonic 1412-A Ether Sulfate in Adju vant and water emulsion (50% v/v). Maximization Test: (April 20, 1977) Seven days following intracutaneous injections, the second stage of the induction procedure was performed. The same area of each animal was reclipped closely and a 2 X 4 patch of Whatman No. 3MM filter paper, saturated with a 40% w/w aqueous dilution of Alfonic 1412-A Ether Sulfate, Sample No. 8126J (3/28/77), was placed over the area of injection. The test patch was secured under an occlusive bandage and left in place for forty-eight hours (April 22, 1977). (May 4, 1977) VVV 000014487 Fourteen days following topical induction, a challenge patch of Alfonic 1412-A Ether Sulfate, Sample No. 8126J (3/28/77), as a 10% w/w aqueous dilution, was applied to the closely clipped flank of each of the ten test animals and to four animals of the same age and size, which had not under gone the induction regimen (controls). The challenge application (same procedure to effect complete occlusion as employed above) was allowed to remain in position for twenty-four hours, after which the patch was removed and the area washed with water. The skin of the challenge area of each test and control animal was evaluated at twenty-four and fortyeight hours following removal of the patches of test material. (part which may not be used in any form of advertising without written permitiim. Scientific Associates, INC. 6200 S. LINDBERGH. ST. LOUIS. MO. 63123 TELEPHONE: 314.4S7.6776 EXPERIMENTAL PROCEDURE: (continued) -4- Maximization Test: (May 9, 1977) Five days following application of the initial challenge dosage, the animals (test and control) were rechallenged as above, using the skin of the opposite flank* The rechallenge area of each animal was evaluated at twenty-four and forty-eight hours following removal of the patches* Following the forty-eight hour reading, the animals were sacrificed and the skin of the rechallenge site of each was removed and preserved in 10% formalin. The tissues of all test and control animals were submitted for histopathological evaluation, which was performed by John D. Bauer, M*D*, Pathologist. RESULTS: The results of the initial range finding study (preliminary evaluation) of Alfonic 1412-A Ether Sulfate, Sample No. 8126J (3/28/77), are found in Table 1. The results of the final evaluation of the initial challenge are found in Table 2. The results of the final evaluation of the final challenge are found in Table 3. The results of the histopathological evaluation of the skin tissue from the final challen site are found in Table 4. S. A. Number 228618 vvv 000014468 This is confidential report which may not be used in any form of adverliaing without written permi*>. Group A: Intracutaneous Injection Page 1 of 3 Ani- Test mal Concen No. tration 24 Sites OBSERVATIONS (hours after injection) 48 Sites Test Reaction Eryth ema Edema * Control Reaction Eryth ema Edema * Test Reaction Eryth ema Edema * Control Reaction Eryth ema Edema * 72 Sites Test Reaction Eryth ema Edema * Control Reaction Eryth ema Edema * WV 0000X44 1 5,00%w/w 3 0 - 1 0 - 44-20-43 - 1 0 l.OOTw/w 2 0 - 1 0 - 32- 20-31B1 0- 0.10%w/w 1 0 1 0- 1 0-20- 10-20- o CD 0.017ow/w 1 0 - 1 0 - 20-20-10 -20- 0.001Xw/w 1 0 - 1 0 - 20- 20- 20 - 20- 3 5.00%w/w 4 0 N 1 0 - 42N10-42N1 0- l,00%w/w 2 0 - 1 0 - 32- 10- 31 - 1 0 - 0.10%w/w 1 0 - 1 0 - 1 0- 1 0- 1 0 - 1 0 - O.OUw/w 1 0 - 1 0 - 1 0 - 1 0- 10 - 1 0- 0.007ow/w 1 0 - 1 0 - 1 0 - 20- 1 0 - 20 - 5 4.007,w/w 3 0 - 2 0 2,007ow/w 3 0 - 2 0 1.007ow/w 1 0 - 1 0 0.507ow/w 1 0 - 2 0 0,25%w/w 2 0 - 2 0 - 7 4.007ow/w 3 0 - 2 0 2.007ow/w 2 0 - 2 0 l.OOTWw 2 0 - 2 0 0.507,w/w 2 0 - 2 0 0,257ow/w 2 0 - 2 0 - 32-10- 33-1 022-00- 20 - 1 0 20-00- 20 -1 020-10-10 -1020-20-10 - 104 2 - 1 0-44N0020- 20- 20 - 0021 B1 0- 21 B 1 021B1 0- 20B 1 010- 1 0- 10-00- *0ther reactions, S.A. Number 228618 Code? 1 Slight 2 * Moderate 3 Marked 4 *= Severe B = Blanched N *= Necrosis Jroup A: Intracutaneous Injection (continued) Page 2 of 3 Ani- Test aub-- mal Concen roup No, tration 24 Sites OBSERVATIONS (hours after injection) 48 Sites Test Reaction Eryth ema Edema * Control Reaction Eryth ema Edema * Test Reaction Eryth ema Edema * Control Reaction Eryth ema Edema * 72 Sites Test Reaction Eryth ema Edema * Control Reaction Eryth ema Edema * 3 15 2.007ow/w 4 4 - 2 0 - 1.00%w/w 2 1 - 1 0 - 0.507ow/w 2 1 - 1 0 - 16 2,007ow/w 4 2 N 2 1 - 1.00%w/w 2 1 - 2 1 - 0.507ow/w 2 1 - 2 1 - 17 2.007ow/w 2 3 - 2 1 - 1.00%w/w 2 2 - 2 1 - 0,507ow/w 2 1 - 2 1 - 18 2.007w/w 4 3 N 2 1 - 1.00%w/w 3 2 - 2 0 - 0.507w/w 2 1 - 2 0 - 43N20-4 3N 1 031 - 1 0- 3 0- 1 020- 10- 2 0- 1 040N20-4 0N1 020- 20- 1 0- 1 020- 20- 20- 1 031 - 20-3 1 - 2031 B20- 30- 1 031B 20- 3 0- 2043N20-43N1 031 -10- 30- 1030- 10- 20- 10- *Other reactions. S.A. Number 228618 Code: I * Slight 2 * Moderate 3 ** Marked 4 - Severe B = Blanched N = Necrosis o O' o o o o Sroup- B: Topical Application (continued) Sub group Animal No* Test Concen tration 24 Hours Erythema Edema Other 1 9 10.07ow/w Undiluted 2 11 25.0%w/w 50.0%w/w 12 25.0%w/w 50.0%w/w 3 19 30.0%w/w 40.0%w/w 0 03 21 02 12 03 12 03 2- Code: 1 = Slight 2 - Moderate 3 = Marked 4 - Severe D = Desquamation L = Leatheriness T = Thickening W = Wrinkling 48 Hours Erythema Edema 00 32 00 20 20 31 20 31 Other . - Page 3 of 3 72 Hours Erythema Edema Other 00 3 2 L,W 0 02 0T 2 03 0 T,W 0 0D 3 3D o* sr * O o o o > > > S. A. Number 228618 Table 2: Evaluation of Challenge Sites - Maximization Test, Initial Challenge Page 1 of 1 1 ,`Vernal Condition 1A Test 2A Test 3A Test 4A Test 5A Test 6A Test 7A Test 8A Test 9A Test 10A 11A 12A 13A Test Control Control Control 1 Control 24 Hours Erythema 0 B.P. Approx. 25% of area 0 B.P. Diffuse, entire area B.P. Approx. 75% of area B.P. <25% of area 0 0 B.P. Approx. 50% of area 0 0 0 B.P. 2 foci, <10% of area 0 Edema 0 0 0 0 0 0 0 0 0 0 0 0 0 0 48 Hours Erythema 0 B.P. <107. of area 0 B.P. <1/4 of area 0 0 0 0 0 0 0 0 0 0 Edema 0 0 0 0 0 0 0 0 0 0 0 0 0 0 Code; 1 = Slight 2 = Moderate 3 = Marked 4 Severe 0 Negative reaction B.P.= Barely perceptible S. A. Number 228618 VVV 000014492 Table 3: Evaluation of Challenge Sites - Maximization Test, Final Challenge Page 1 of 1 1 ' al Condition 1A Test 2A Test 24 Hours Erythema 0 B.P. <25% of area 3A 4A 5A 6A 7A 8A 9A 10A 11A 12A 13A 14A Test Test Test Test Test Test Test Test Control Control Control Control 0 0 B.P. Approx, 25% of area 0 0 0 B.P. <50% of area 0 0 0 0 0 v i S 1 = Slight 2 = Moderate 3 = Marked 4 * Severe 0 * Negative reaction B.P.- Barely perceptible Edema 0 0 0 0 0 0 0 0 0 0 0 0 0 0 48 Hours Erythema 0 B.P. Approx. 25% of area 0 0 B.P. Approx. 25% of area 0 0 0 B.P. <25% of area 0 0 0 0 0 Edema 0 0 0 0 0 0 0 0 0 0 0 0 0 0 S. A. Number 228618 ooooi^93 Table 4: Histopathological Evaluation of Skin Tissue From Final Challenge Site. ______________________________ Page 1 of 1 | --tmal Condition Microscopic Findings 1A 2A 3A 4A SA 6A 7A 8A >A 10A 11A 12A 13A 14A Test Test Test Test Test Test Test Test Test Test Control Control Control Control Superficial perifollicular infiltrate not exceed ing that seen in controls. Scattered lymphocytes and large mononuclear cells were seen within the dermal papillae. Scattered lymphocytes and large mononuclears were seen within the dermal papillae. Slight focal subepithelial infiltrate by lympho cytes and some perifolliculitis, not exceeding changes seen in controls. Some infiltration by lymphocytes and large mono nuclears was seen within the papillae. Slight focal subepithelial infiltrate by lympho cytes and some perifolliculitis, not exceeding that seen in controls. No changes seen. No changes seen. Some infiltration by lymphocytes and large mono nuclears was seen within the papillae, occasion ally involving basal layers of the overlying squamous epithelium. No changes seen. Negative Negative Negative Negative { j ] = Except for occasional slight j 5 ' perifollicular lymphocytic infiltration. Comment: There were no histological changes present in any of the ten test skin tissues which were indicative of allergic reaction. The findings for each of the four control specimens were negative except for occasional perifollicular lympho cytic infiltration. S. A, Number 228618 WV 000014494 n Scientific Associates, ?nc. 6200 S. LINDBERGH. ST. LOUIS. MO. 63123 TELEPHONE: 314-487-6770 CONCLUSION: -5- The test material, Alfonic 1412-A Ether Sulfate, Sample No* 8126J (3/28/77), was found not to produce sensitization of guinea pigs when assayed by the Guinea Pig Maximization Test (op. cit.). SCIENTIFIC ASSOCIATES, INC. Robert H. Moulton Vice-President and Director Biological Research Services S. A. Number 228618 cs VVV 000014495 Thi* it confidential report which >nay not be used in any form of advertising without written permisaiOT.