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KfjytWo <VUtoT U-t AX ~ r ui-v > ANALYSIS OF BIODEGRADATION DATA d ua lxs dCS Special Study 70-22 Job No. 8908116 Data collected in the biodegradation test program prior to November 16, 1970 were submitted for statistical analysis. The numbers used in the analysis represented the amount of test compound degraded in a 21 hour period. materials representing varying degrees of molecular complexity/were tested. The conditions for biodegradation were similar for all/, except that smaller quantities of the less degradable material^ were used.. Student's t Test w$/s used to check for significant evidence of biodegradation. Control charts were plotted on the amount of the test compound pres at the beginning and the end of the 21 hour period CONCLUSIONS t 1. Significant evidence of biodegradability was found for MJP&7 IIB--40 -wid MCS 1043. Significance level used - 95%. 2. MCS 1016 and MCS 1242 showed some evidence of biodegradation, but neither was significant at 95%. 3. Aroclor 1254 showed no evidence of biodegradation. 4. The compounds studied ranked from the most to the least degradable were : 4-6, MCS 1043, MCS 1242, MCS 1016, Aroclor 1254. (See Table I) ----------------------------------------------------------------------- 5. In order to detect biodegradability at the 95% level of significance in 20 observations, approximately one-half of the amount of test material added must be degraded. For MCS 1016 this would require about one week; for Aroclor 1242, about 2 days. 6. The day to day variation in amount of test compound found both before and after the 21 hour degradation period is shown in Figures 1 thru 6. DISCUSSION This was a preliminary analysis of existing data. This will be followed by a designed experiment to investigate several variations in experimental technique. The designed experiment .is intended to lead to experimental conditions which will provide more consistent results. . db Monsanto Company Organic Chemicals Division Applied Sciences Section St. Louis, Missouri 11/70 - J. D. Hinchen, R. E. Keller DSW 182864 STLCOPCB4050961 TABLE I COMPARISON OF TEST MATERIALS DIFFERENCE BETWEEN FEED SAMPLE + 21 HOUR SAMPLE Material Ave. Diff.(mg) rUrD HB-4-0MCS 1016 Aroclor 1242 Aroclor 1254 MCS 1043 O>3* 0 1 --StT -7--.fr 0.11 0.30 0.55 . Std. Dev. of Diff. S/D 3rr?5 8v80ca 1.0 1.07 ca 1.0 0.95 fr.22 4.26 10 3.56 25.0 1.73 Significance It NS NS NS * * = 95% Significance -***--= 99. fr%--Significance NOTE 10 mg of MIPB and HB-T were added every 24 ho For the other compound , the addition rate was 1 mg every 4 hours. s. DSW 182865 STLCOPCB4050962 i I i i '-ywnjCL| y*r>c$tit4^ '`dh'ty/ oSVM 182866 STLCOPCB4050963 F l& lA G 2, STLCOPCB4050964 3 3 1 3 3 1^0 is j* I < i j jj i ;.i ] * V> ^ fO * -- DSW 182868 STLCOPCB4050965 *Kw STLCOPCB4050966 STLCOPCB4050967 f^naj QM'WtoQQ $ L [ [ L r L [ r v ! t, f L [ [ DSW 182871 f J i i STLCOPCB4050968 Statistics Special Study 71-2 Job No. 85050-16 STUDY OF OPERATING VARIABLES IN THE ACTIVATED SLUDGE BI ODEGRADEABI LI T Y TEST INTRODUCTION In order to provide more meaningful information on the biodegradation of Monsanto products, several of the operating variables in the activated sludge test procedure were studied. These variables were time cycle, suspended solids, amount of test material fed, method of feeding, aeration rate, method of sampling and concentration of test material in the solvent. The effects of these variables on the percent disappearance of the test material during the time period involved were sought. Also of interest was the vari ability of the calculated percent disappearance. Three test materials (biphenyl, 2, 4'-polychlorinated biphenyl and 2, 2', 4, 4', 6, 6'-polychlorinated biphenyl) were used. CONCLUSIONS 1. Higher percent disappearance was obtained at longer time cycles, lower suspended solids, higher amount of test material fed, surface feed ing and sidearm sampling (biphenyl and 2,4'-PCB). 2. On the hexachlorinated material deep injection feeding and low concentration in solvent gave higher results. 3. At the levels studied, aeration rate has no effect. 4. The longer time cycle and lower suspended solids gave more re producible result. DATA AND OBSERVATIONS The experimental design used was a Plackett and Burman screening design which permits study of seven main effects in eight experimental runs. The factors and levels were as follows: Factor Low Level High Level Time Cycle Suspended Solids Amt. of Material Fed Method of Feed Aeration Rate Method of Sampling Concentration in solvent 24 hrs. 1250 g 1 mg Surface 1/2 cu ft/min Sidearm 1/2 max. 48 hrs. 2500 g 5 mg Deep Injection 1 cu ft/min Surface max. DSW 182872 STLCOPCB4050969 Statistics Special Study 71-2 Page 2._______ The experimental design is shown below: Run No. Time Suspended Amt. of Method Cycle Solids Material of Feed Aeration Method Concentratior Rate of Sampling in Solvent 1 24 1250 2 24 1250 3 24 2500 4 24 2500 5 48 1250 6 48 1250 7 48 2500 8 48 2500 1 mg 5 mg 1 mg 5 mg 1 mg 5 mg 1 mg 5 mg Surface Surface Injection Injection Injection Injection Surface Surface 1 cu ft/min \n " i" 1/2 " 1/2 " 1" 1/2 " 1" Sidearm Surface Surface Sidearm Sidearm Surface Surface Sidearm 1/2 max rnax nax 1/2 max max 1/2 max 1/2 max max The differences between factor levels were calculated for each test material by subtracting the results of the runs at the low level of each factor from the results at the high level of each factor. The response was % disappearance of the test material during the time period under study. Runs 1 through 4 had four results each and runs 5 through 8 had two results each. The dif ferences given below are based on the average percent disappearance for each run. Differences Between Factor Levels Biphenyl 2,4' -PCB ^2,2', 4, 4', 6, 6'-PCB Total Time Cycle 5. 6 Suspended Solids -11.4** Amt. Material 10. 2* Method of Feed 3. 6 Aeration Rate - 3.5 Method of Sampling - 8.4* Cone, in Solvent - 1.6 39. 5*** -34.5*** - 2.2 33.1*** 1.2 - 3.4 - 3. 3 3. 3 -2.3 ' 8. 1 -12.1** -1.7 6. 1 -15.6*** 1 COo Ul 48. 4 * * * -43. 6*** 16. 1 24. 6 - 4.0 - 5. 7 ^ ^ (One wild value in run 8 was replaced by a more reasonable result. ) For each material an analysis of variance was run on the factors giving the three highest differences as shown above. The asterisks show statistical significance at 90% (one asterisk) and 95% (two asterisks). Reproducibility of the data, expressed as estimated standard deviation, was calculated for each run as shown below. 0S\N 182873 STLCOPCB4050970 Run No. 1 2 3 4 5 6 7 8 Statistics Special Study 71-2 Page 3. Biphenyl 4. 0 0. 3 5. 3 6. 7 0. 0 0. 0 10. 6 1.2 2,4' -PCB 0. 8 8. 7 10. 7 8. 1 0. 1 0. 0 3. 3 1.2 2, 2',4,4', 6, 6'-PCB Ave 17. 3 9. 3 23. 9 16. 1 0. 2 6. 6 6.4 11.2 9. 8 6. 1 13. 3 10. 3 0. 1 2.2 6.8 . 4. 5 Ave. Std. Deviation Factor Time Suspended Solids Sample Size Low Level 9.9 4. 3 7. 5 High Level 3.4 8. 7 5.8 RECOMMENDATIONS The following conditions were recommended for adoption in routine degradation testing: 48 hr cycle, low suspended solids, surface feed, 5 mg sample, sidearm sampling and low concentration in solvent. MONSANTO COMPANY Organic Chemicals Division Applied Sciences Section St. Louis, Mo. 3-71 J. D. Hinchen SW1828?4 STLCOPCB4050971