Biodegradability and Acute Aquatic 

Toxicity of Alcohol Propoxy Sulfates or 

Sasol’s ALFOTERRA® Surfactants

Sasol Report Number R&D ENV 10-005 01-10

Sasol North America

May 2010

Work conducted by Geoff Russell and Nomihla Valashiya-Mdleleni

Report edited by Marta Bourke
12/14/2011 Page 2 of 66

Interoffice Correspondence 

To Ricky Stackhouse

From Geoff Russell, Marta Bourke, Victoria Stolarski

Date May 30, 2010

Subject: PROJECT SUMMARY: Sasol Report R&D ENV 10-005 01-10 

Biodegradability and Acute Aquatic Toxicity of Alcohol Propoxy Sulfate or 

Sasol’s ALFOTERRA® Surfactants. 

Key Words: ALFOTERRA®, Alcohol propoxylates, Alcohol propoxy sulfates, Biodegradation, 

Aquatic Toxicity, Oil recovery, Emulsification

Conclusions: This report documents Sasol North America Inc.’s additional biodegradation and 

aquatic toxicity studies on the new products, ALFOTERRA® alcohol propoxy sulfate surfactants.

This work confirms that the ALFOTERRA® alcohol propoxy sulfates, based on either C12-C13, 

C14-C15, or C16-C17 branched / linear primary alcohol, pose little risk to the environment. The 

ALFOTERRA surfactants are not intended or expected to be released into the environment 

when used as a surfactant for the enhanced recovery of petroleum oil. 

Using ALFOTERRA® 123-4S (ALFO 23 -4S) surfactant as the model compound for the studies 

contained herein, the following conclusions can be made:

1) The rate and extent of the biodegradation of alcohol propoxy sulfates, specifically 

branched C12-13 alcohol 4-mole propoxy sulfate (ALFO 23-4S) is notably less than that 

of the standard detergent surfactant alcohol ethoxy sulfate, or specifically - linear 

dodecyl 4-mole ethoxy sulfate (POLYSTEP B-430S from Stepan Company) as shown by 

the remaining dissolved organic carbon in the ALFO 23-4S effluent from Porous Pot, 

Die-Away, and Electrolytic Respirometer testing.

2) Acute aquatic toxicities to fish of the neat material and the effluents from Porous Pot and 

Die-Away testing were determined using standard US EPA methods. The results 

showed that both ALFO 23-4S and the control POLYSTEP B-430S had the same LC 50 

values for the neat and effluent samples despite the significant remaining dissolved 

organic carbon in the effluent of the ALFO 23-4S sample. 

3) Methylene Blue Active Substances (MBAS) analysis of both the neat and effluent 

samples proved that ALFO 23-4S undergoes primary degradation and elimination of 

surfactancy despite the significant remaining dissolved organic carbon in the effluent of 

the ALFO 23-4S sample after porous pot or 28-day die-away testing.

4) The acute aquatic toxicity to fish of the neat ALFO 23-4S is eliminated once the ALFO 

23-4S passes through the porous pot or 28 die-away test. The results of 96-hour static 

P. promelas (EPA-821-R-02-013) showed for example that the LC50 of 14% dilution for 

the ALFO 23-4S feed to the porous pot decreased to LC50 >100% dilution for the 

effluent.

5) Using ALFOTERRA® surfactants for enhanced hydrocarbon recovery (EHR) will not 

result in the discharge of any material into surface or ground water. Any rinse aid 
12/14/2011 Page 3 of 66

containing ALFOTERRA® surfactants, after processing through standard domestic or 

industrial waste water treatment, will pose no threat to the environment.

Results Summary: Sasol designed and assembled a porous pot system in accordance with 

the “Fate, Transport and Transformation Test Guidelines OPPTS 835.3240 Simulation Test—

Aerobic Sewage Treatment: Activated Sludge Units Guideline” during the spring of 2009 to 

further evaluate the biodegradation of branched alcohol propoxy sulfate surfactants. The neat 

surfactant, and the influent and effluent to the Porous Pot System and additionally to the OECD 

301-A 28 Day Die-away Biodegradation Study was further evaluated by 96-hour static P. 

promelas (EPA-821-R-02-013) and Methylene Blue Active Substance. All sample details, 

experimental information, and discussion of the results are given in the following detailed report

and appendices. A summary of the most relevant results is given here.

Table I - Porous Pot Data: ALFO 23-4S compared to Benzoate Control

DOC of Benzoate or ALFO 23-4S in Feed (mg/L)

11.47 10.39

Steady State established after foaming and zooglea issues

Blank Benzoate ALFO 23-4S Blank Blank Blank Benzoate Benzoate Benzoate Benz/Blk ALFO 23-4S ALFO 23-4S ALFO 23-4S Alfo/Blk

Feed Feed Feed Effluent Effluent Mean Effluent Effluent Mean Mean Δ Effluent Effluent Mean Mean Δ

9-Jul 298.728 311.296 298.594

10-Jul 317.728 325.822 328.198 10.925 10.027 10.476 10.282 10.059 10.170 (0.306) 16.795 17.069 16.932 6.456

11-Jul 328.520 344.872 340.120 12.123 10.784 11.453 10.257 11.208 10.733 (0.721) 23.031 21.277 22.154 10.700

12-Jul 320.754 329.708 327.200 10.177 9.901 10.039 9.635 11.296 10.465 0.426 24.781 21.062 22.921 12.882

13-Jul 308.686 326.120 319.074 9.601 12.228 10.914 7.604 9.700 8.652 (2.262) 21.760 20.971 21.365 10.451

14-Jul 339.732 340.814 327.676 10.750 14.165 12.457 8.695 11.253 9.974 (2.483) 19.603 21.720 20.662 8.204

15-Jul 341.820 338.308 337.690 11.782 13.150 12.466 9.664 12.519 11.091 (1.375) 20.341 20.116 20.228 7.762

17-Jul 232.758 233.788 240.148 12.684 12.014 12.349 10.059 11.731 10.895 (1.454) 20.030 19.936 19.983 7.634

18-Jul 232.886 231.650 211.742 11.512 9.657 10.585 9.846 10.122 9.984 (0.601) 18.593 18.202 18.397 7.813

19-Jul 214.990 208.772 239.962 10.142 8.769 9.455 9.585 8.777 9.181 (0.274) 17.149 16.910 17.029 7.574

20-Jul 211.682 217.640 225.588 10.135 9.646 9.890 10.248 9.934 10.091 0.200 17.453 18.026 17.739 7.849

21-Jul 207.632 228.256 192.536 9.948 10.538 10.243 9.276 9.871 9.573 (0.670) 18.599 19.865 19.232 8.989

22-Jul 170.546 162.330 187.782 10.278 10.342 10.310 8.374 10.016 9.195 (1.115) 18.656 19.230 18.943 8.633

23-Jul 291.736 297.840 295.308 11.071 12.114 11.592 10.128 12.021 11.074 (0.518) 18.546 17.728 18.137 6.545

24-Jul 281.972 282.374 263.638 12.209 13.765 12.987 11.293 12.585 11.939 (1.048) 19.924 19.956 19.940 6.953

25-Jul 271.570 248.690 278.154 12.292 12.273 12.282 10.240 10.609 10.424 (1.858) 19.288 19.466 19.377 7.095

26-Jul 284.028 284.438 284.248 11.954 9.425 10.689 8.959 9.459 9.209 (1.481) 17.847 17.849 17.848 7.159

27-Jul 299.512 290.840 272.186 10.939 8.797 9.868 7.975 7.508 7.741 (2.127) 18.242 18.240 18.241 8.373

28-Jul 300.796 293.262 295.778 9.882 9.287 9.584 8.158 7.811 7.984 (1.600) 17.745 18.582 18.163 8.579

29-Jul 280.696 283.508 283.382 9.548 9.386 9.467 8.711 7.597 8.154 (1.313) 18.532 19.619 19.075 9.609

Average 10.945 10.856 10.900 9.420 10.214 9.817 19.311 19.254 19.282

Std Dev 1.00 1.69 0.97 1.55 2.04 1.44

Mean Difference (Test-Blank Control) -1.08 8.38

Std Deviation (Test-Blank Control) 0.81 1.61

Number of Paired Observations 19 19

Calculated Student's T Value 5.84 22.76

Critical Student T Value 1.73 1.73

Percent Removal 109.44% 19.33%

The resulting porous pot data was analyzed as described in the OPPTS 835.3240 guidelines. 

Each sample, Blank, Benzoate, and ALFO 23-4S, was run in duplicate. The benzoate control 

showed on average 109% removal verses the blank feed. The ALFO 23-4S showed only 19% 

removal verses the blank feed. Though the ALFO 23-4S does degrade somewhat when run 

through the porous pot, a significant portion of the original dissolved organic carbon can be 

seen in the effluent.
12/14/2011 Page 4 of 66

Table II - Porous Pot Data: ALFO 123-4S compared to Linear C12 alcohol ethoxy sulfate 

(AES), a standard commercial detergent surfactant

DOC of AES or ALFO 23-4S in Feed (mg/L)

16.91 16.04

Steady state established after AES introduced as comparative sample

AES ALFO 23-4S Blank Blank Blank Blank AES AES AES AES ALFO 23-4S ALFO 23-4S ALFO 23-4S ALFO 23-4S

Feed Feed Feed Effluent Effluent Mean Effluent Effluent Mean Mean Δ Effluent Effluent Mean Mean Δ

15-Aug 247.940 239.436 243.984

16-Aug 227.632 242.160 235.068 9.695 8.290 8.992 10.478 10.862 10.670 1.678 21.293 19.181 20.237 11.245

17-Aug 281.066 271.800 270.500 11.568 8.570 10.069 11.509 14.866 13.187 3.119 17.302 17.302 17.302 7.233

18-Aug 285.140 270.510 269.560 12.018 9.307 10.662 11.688 15.705 13.696 3.034 18.969 22.359 20.664 10.002

19-Aug 275.450 271.450 269.650 13.800 10.053 11.926 11.848 15.251 13.549 1.623 19.597 22.014 20.805 8.879

20-Aug 289.350 278.560 281.450 14.611 11.347 12.979 17.829 21.568 19.698 6.720 15.622 17.571 16.597 3.618

21-Aug 279.750 278.800 277.350 16.138 11.615 13.877 14.716 19.332 17.024 3.147 17.644 18.968 18.306 4.430

22-Aug 280.540 278.340 281.650 17.584 12.708 15.146 12.209 17.113 14.661 (0.485) 23.570 21.705 22.637 7.492

23-Aug 298.340 287.560 285.350 16.477 13.405 14.941 11.595 16.266 13.931 (1.010) 25.438 21.192 23.315 8.374

24-Aug 286.440 279.340 281.680 14.895 13.977 14.436 12.185 15.317 13.751 (0.685) 26.305 19.852 23.078 8.642

25-Aug 295.740 289.440 290.680 15.076 14.925 15.000 11.531 14.278 12.904 (2.096) 26.019 19.862 22.941 7.940

26-Aug 298.420 295.560 300.120 14.906 13.342 14.124 10.975 12.606 11.791 (2.333) 24.315 18.374 21.345 7.221

27-Aug 330.952 316.158 318.600 15.615 13.905 14.760 10.818 12.286 11.552 (3.208) 23.274 18.981 21.127 6.367

28-Aug 307.162 290.204 300.828 12.299 14.862 13.580 10.349 11.129 10.739 (2.841) 23.970 18.040 21.005 7.425

29-Aug 304.460 301.334 292.984 13.277 16.367 14.822 10.875 11.683 11.279 (3.543) 24.091 18.669 21.380 6.559

30-Aug 325.070 320.074 320.618 12.634 16.180 14.407 11.779 12.318 12.048 (2.358) 25.882 22.884 24.383 9.976

31-Aug 291.448 298.708 293.032 12.591 15.182 13.886 20.714 20.531 20.623 6.736 27.283 24.304 25.794 11.907

1-Sep 294.140 285.684 288.552 13.315 13.724 13.519 15.978 15.113 15.545 2.026 26.113 22.066 24.090 10.570

2-Sep 312.742 309.938 313.838 13.188 13.010 13.099 11.366 10.581 10.973 (2.126) 25.023 21.006 23.014 9.915

3-Sep 290.976 289.786 290.064 14.452 12.917 13.684 10.427 10.439 10.433 (3.252) 25.200 21.760 23.480 9.795

4-Sep 288.360 296.040 290.426 13.879 12.045 12.962 10.066 9.636 9.851 (3.112) 24.601 21.613 23.107 10.145

5-Sep 309.616 312.466 321.020 12.706 11.765 12.235 9.901 9.223 9.562 (2.674) 23.713 21.243 22.478 10.242

6-Sep 287.982 284.368 285.684 12.154 10.926 11.540 10.149 9.594 9.871 (1.669) 23.138 21.749 22.443 10.904

7-Sep 284.012 281.480 275.438 12.281 11.856 12.068 10.503 10.411 10.457 (1.611) 22.527 22.286 22.406 10.338

8-Sep 316.224 310.068 314.572 13.398 13.973 13.686 12.498 11.436 11.967 (1.719) 23.158 23.409 23.283 9.598

9-Sep 298.202 283.170 284.760 14.156 16.722 15.439 12.835 11.529 12.182 (3.257) 23.764 24.306 24.035 8.596

10-Sep 289.590 291.248 281.028 13.958 20.085 17.021 12.443 11.069 11.756 (5.265) 22.332 23.412 22.872 5.851

11-Sep 313.108 314.666 321.496 13.364 17.689 15.527 12.641 10.469 11.555 (3.972) 22.748 22.751 22.749 7.223

12-Sep 298.286 292.386 285.142 12.454 13.183 12.819 11.709 10.070 10.889 (1.929) 20.968 22.204 21.586 8.767

13-Sep 295.626 286.828 284.828 11.568 11.801 11.685 11.608 9.767 10.688 (0.997) 21.284 22.282 21.783 10.098

14-Sep 319.138 315.114 319.714 10.784 11.753 11.268 11.620 9.710 10.665 (0.604) 24.308 22.193 23.250 11.982

15-Sep 319.138 315.114 319.714 11.152 11.740 11.446 16.719 10.765 13.742 2.296 24.853 22.927 23.890 12.445

16-Sep 291.394 281.264 291.620 10.475 11.996 11.235 17.648 10.704 14.176 2.941 22.944 22.809 22.876 11.641

17-Sep 309.616 315.482 315.364 10.270 13.251 11.761 17.593 10.528 14.060 2.300 23.992 23.299 23.645 11.885

18-Sep 300.366 295.708 302.800 10.234 12.364 11.299 16.903 10.486 13.694 2.396 25.509 23.497 24.503 13.204

19-Sep 299.902 294.464 288.270 10.007 11.515 10.761 14.706 10.180 12.443 1.682 23.861 22.900 23.380 12.619

20-Sep 305.896 315.156 317.666 9.932 10.770 10.351 13.268 10.482 11.875 1.524 23.232 22.389 22.810 12.460

21-Sep 295.528 286.798 288.486 11.445 11.675 11.560 13.377 11.486 12.431 0.871 23.479 23.480 23.479 11.920

22-Sep 294.174 292.998 287.644 11.901 12.035 11.968 16.809 15.046 15.927 3.960 23.499 24.853 24.176 12.208

23-Sep 324.054 296.356 322.126 13.533 15.133 14.333 12.245 13.439 12.842 (1.491) 23.338 23.302 23.320 8.987

24-Sep 301.344 300.094 296.898 11.314 13.262 12.288 12.698 15.840 14.269 1.981 23.617 23.046 23.332 11.044

Average 12.877 12.980 12.929 12.920 12.728 12.824 23.194 21.651 22.422

Std Dev 1.81 2.23 1.67 2.69 3.57 2.99 3.20 1.89 2.20

Mean Difference (Test-Blank Control) (0.105) 9.494

Std Deviation (Test-Blank Control) 2.90 2.32

Number of Paired Observations 40 40

Calculated Student's T Value (0.23) 25.87

Critical Student T Value 1.67 1.67

Percent Removal 100.62% 40.81%

During the above time period the ALFO 23-4S showed 41% removal versus the blank feed. 

However, the AES showed 100% removal versus the blank feed. Though similar surfactants, 

the branching of the alcohol and alkoxylate portions of the ALFO 23-4S significantly inhibits 

complete biodegradation within the evaluation period, unlike the all-linear AES which quickly 

and completely degrades.
12/14/2011 Page 5 of 66

Graph I - OECD 301-A Die-Away Data: ALFO 123-4S compared to Linear C12 alcohol 

ethoxy sulfate (AES), a standard commercial detergent surfactant

OECD Die-Away DOC Loss

0

20

40

60

80

100

0 5 10 15 20 25 30

Days

%

D

O

C Loss

AES ALFO Benz

A 28-day die-away biodegradation study was run on the neat ALFO 23-4S and AES surfactants

shown above. Additionally, the die-away test was run on the effluents on the porous pot to 

show that the biodegradation of the ALFO 23-4S, though slow, will continue (data in full report). 

Again the neat ALFO 23-4S shows only partial degradation in the 28 day period.

Graph II - Methyl Blue Active Substance (MBAS) Data: ALFO 123-4S compared to Linear 

C12 alcohol ethoxy sulfate (AES), a standard commercial detergent surfactant

Porous Pot: MBAS Primary Biodegradation 

0

5

10

15

20

25

30

35

40

45

Blank AES ALFO 23-4S

MBAS (mg/L)

Influent

Effluent

Influent

Effluent
12/14/2011 Page 6 of 66

MBAS was run on the blank, AES, and ALFO 23-4S influent and effluent samples of the porous 

pot study (PPS). The results show that significant primary biodegradation occurred for both 

surfactants. The MBAS percent reduction in activity for the AES and ALFO 23-4S during the 

PPS were 99% and 94% respectively.

Surfactantcy (3mg/L) seen in the blank influent is due to the commercial laundry soap included 

in the blank recipe.

Table III - 96-hour static P. promelas (EPA-821-R-02-013) of neat, influent and effluent 

samples of ALFO 23-4S and AES

Sample 

Description

Environmental 

Enterprise USA

Sample ID

Survival 

NOEC %

Survival 

LOEC %

LC50 %

(95% Confidence 

Interval)

PPS Blank Influent 

Feed

Q-642-09-1336-38-1 12% 25% 14.7 % 

(12.4% - 17.5%)

PPS AES Influent 

Feed

Q-643-09-1336-38-2 3.0% 6.0% 5.47%

(4.91% - 6.10%)

PPS ALFO 23-4S 

Influent Feed

Q-644-09-1336-38-3 6.0% 12.0% 9.48% 

(8.28% - 10.9%)

AES Neat

(40 mg/L active)

Q-645-09-1336-38-4 6.0%

(2.4 mg/L)

12.0%

(4.8 mg/L)

13.8% (5.52 mg/L)

(12.3% - 15.5%)

ALFO 23-4S Neat

(40 mg/L active)

Q-646-09-1336-38-5 3.0% 

(1.2 mg/L) 

6.0%

(2.4 mg/L)

7.76% (3.10 mg/L)

(6.64% - 9.08%)

PPS Blank 

Effluent

Q-647-09-1336-38-6 100% >100% >100%

(NA)

PPS AES Effluent Q-648-09-1336-38-7 100% >100% >100%

(NA)

PPS ALFO 23-4S 

Effluent

Q-649-09-1336-38-8 100% >100% >100% 

(NA)

ALFO 23-4S 

Die-Away Influent

(50 mg/L) 0-day

Q-715-09-1336-58-2 6%

(3.0 mg/L)

12%

(6.0 mg/L)

8.8% (4.4 mg/L)

(8.0% - 9.7%)

ALFO 23-4S 

Die-Away Effluent

(50mg/L) 28-day

Q-716-09-1336-58-1 100% >100% >100%

NA

Data reported is the weight-percent of the original test solution in the final dilution. For example, line 1 shows that in 

order to achieve 50% survival rate, the PPS Blank Influent solution had to be diluted to 14.7% PPS Blank Influent and 

85.3% water. The higher the value in the table the less effect the original solution had on the organisms. Values of 

>100% indicate that all of the organisms survived in the solution as supplied to the test. No dilution was required.

Acute aquatic toxicity testing on fat-head minnows showed that the toxicity of the neat and 

influent ALFO 23-4S is completely removed by processing through the porous pot system or 28-

day die-away, despite the incomplete degradation seen. In fact, the effluent toxicity of the ALFO 

23-4S is equal to that of the AES which shows complete degradation in the laboratory tests.

It can be concluded from this data that ALFO 23-4S and similar branched alcohol propoxy 

sulfates will not undergo complete biodegradation but will exhibit primary degradation which 

renders the original molecule and resulting metabolites inert to aquatic life.
12/14/2011 Page 7 of 66

Biodegradability and Acute Aquatic Toxicity 

of Alcohol Propoxy Sulfates or Sasol’s 

ALFOTERRA® Surfactants 

Sasol Report Number R&D ENV 10-005 01-10

Sasol North America

May 2010

Work conducted by Geoff Russell and Nomihla Valashiya-Mdleleni

Report edited by Marta Bourke
12/14/2011 Page 8 of 66

Table of Contents

TABLES..................................................................................................................................... 9

TERMS ...................................................................................................................................... 9

INTRODUCTION ......................................................................................................................10

OBJECTIVES ...........................................................................................................................10

CONCLUSIONS........................................................................................................................11

RESULTS OF ...........................................................................................................................11

EXPERIMENTS ........................................................................................................................11

TEST MATERIALS................................................................................................................11

BIODEGRADATION STUDY.................................................................................................13

Porous Pot System (PPS) Biodegradation Test Method....................................................13

Porous Pot System Daily Operations.................................................................................15

Porous Pot System Experimental Results .........................................................................18

PPS Results Summary:.....................................................................................................22

OECD 301 A: DOC Die-Away Experimental Results .............................................................22

MBAS Analysis..................................................................................................................24

Aquatic Acute 96hr Fathead Minnow Results ....................................................................26

OECD 301F Biodegradability Electrolytic Respirometer Study ..............................................29

OECD 301 F Electrolytic Respirometer Results:................................................................32

OECD 301 F Electrolytic Respirometer Summary: ............................................................34

Discussion ................................................................................................................................35

Environmental Fate and Effects of ALFOTERRA® Surfactants..............................................35

Biodegradation Pathway of Anionic Surfactants Studied .......................................................36

REFERENCES .........................................................................................................................39

APPENDICES...........................................................................................................................40

Table of Figures

Figure 1: Porous Pot System (PPS) Assembly Test..................................................................14

Figure 2: Hydraulic Retention Time (HRT) Through Porous Pot System ..................................15

Figure 3: Mixed Liquor Total Suspended Solids (MLTSS) in Porous Pot Candle......................16

Figure 4: Porous Pot Effluent TSS ...........................................................................................17

Figure 5: Porous Pot Effluent pH Values..................................................................................17

Figure 6: Porous Pot DOC Feeds and Effluents.......................................................................18

Figure 7: Porous Pot Effluents DOC.........................................................................................19

Figure 8: Porous Pot Effluents - Un-acclimated and acclimated Response ..............................20

Figure 9: Return to Steady State (DOC)...................................................................................20

Figure 10: Porous Pot Effluent DOC: A Comparison of AES and ALFO 23-4S........................21

Figure 11: Porous Pot Effluent: Increased HRT effect on DOC ...............................................22

Figure 12: Die-Away Porous Pot Effluent .................................................................................23

Figure 13: OECD 301A Die-Away DOC Loss of neat ALFO 23-4S, AES, and Benzoate..........24

Figure 14: Porous Pot MBAS Primary Biodegradation .............................................................25

Figure 15: ALFO 23-4S Comparison MBAS Porous Pot and Die Away....................................25

Figure 16: AES and ALFO 23-4S Neat Material Acute Toxicity Before Any Biodegradation .....27

Figure 17: Porous Pot Feed and Effluent: Aquatic Toxicity AES and ALFO 23-4S ...................28

Figure 18: Acute Toxicity Test, Die-Away Test ALFO 23-4S ....................................................29

Figure 19: Biodegradation Study - Alcohols (OECD 301F).......................................................32

Figure 20: Biodegradation Study - Alcohol 4 Mole Propoxylates (OECD 301F).........................33

Figure 21: Biodegradation Study - Alcohol 4 Propoxy Sulfate (OECD 301F)...........................34

Figure 22: Biodegradation profile of ALFO 23-4S Derivatives ..................................................35

Figure 23: Alcohol Ether Sulfate Structure: AES and ALFO 23-4S..........................................36
12/14/2011 Page 9 of 66

TABLES

Table 1: Properties of the four mole propoxy alkoxylate derivatives (nonionic) of the six alcohols 

tested........................................................................................................................................12

Table 2: Properties of the anionic Surfactants based on the above propoxylates which are 

comparable to the ALFOTERRA® Surfactant ............................................................................13

Table 3: Results of fathead minnow 96hr acute toxicity test (see Appendix 23)........................27

TERMS

TERM DEFINITION

(xxx-xx-x) Reference to internal laboratory sample numbers

AES Alcohol ethoxysulfate

ALFOTERRA® Trade name given to the sodium salts of branched-alcohol propoxy 

sulfate surfactants manufactured by Sasol Italy and imported by Sasol 

North America

ALFO 23-4S ALFOTERRA® 123-4S Surfactant

DOC Demand Oxygen (mg/L)

HRT Hydraulic retention time (hours)

MBAS activity Methylene Blue Active Substances (mg/L)

MLTSS Mixed Liquor Total suspended solids (mg/L)

PEG Polyethylene glycol

PPG Polypropylene glycol

PPS Porous Pot System test

TSS Total suspended solids (mg/L)

SDS linear alcohol sulfate dodecyl sulfate

SRT Sludge retention time (days)
12/14/2011 Page 10 of 66

INTRODUCTION

Results presented in Research Report R&D ENV-07-024 01-07 “Biodegradability and Acute 

Aquatic Toxicity of Alcohol Propoxy Sulfates” prompted Sasol to further evaluate the fate and 

effects of alcohol propoxy sulfate surfactants or Sasol’s ALFOTERRA®surfactants. In this 

second study, the ALFOTERRA® Surfactants are evaluated following the “Fate, Transport and 

Transformation Test Guidelines OPPTS 835.3240 Simulation Test—Aerobic Sewage 

Treatment: Activated Sludge Units Guideline”. Specifically, the simulated waste treatment 

system referred to as the Porous Pot System (PPS) was requested. 

Other studies were conducted to supplement the porous pot results and are also included in this 

report. These additional studies are “OECD 301A: DOC Die-Away” on the Porous Pot effluents 

and the neat ALFOTERRA® Surfactant. Additionally, acute aquatic toxicity test by 96-hour static 

P. promelas (EPA-821-R-02-013) was conducted on the effluents of the Porous Pot and DieAway tests. 

Finally, to further understand the impact of the molecular structure on the biodegradation 

response of the ALFOTERRA® Surfactant molecule, “OECD 301F: Electrolytic Respirometer 

testing” (OECD, 1993) was conducted on alcohols, alcohol propoxylates, and alcohol propoxy 

sulfates with varying carbon chain structures. 

. 

OBJECTIVES

1. Set-up and operate a PPS evaluating a representative ALFOTERRA® Surfactant; 

compare the ALFOTERRA® performance in the PPS against standards based on DOC 

removal and MBAS analysis. Change PPS operating conditions throughout the study to 

further enhance the understanding of PPS and fate of the ALFOTERRA®studied.

2. After reaching specific steady state in the PPS, determine the acute aquatic toxicity on 

fathead minnow of PPS feeds, effluents and neat ALFOTERRA®.

3. Conduct an OECD 301A; DOC Die-away on the PPS effluents and on the neat 

ALFOTERRA® with two standards. 

4. At the end of the 28-day die-away, conduct an acute aquatic minnow toxicity test on the 

ALFOTERRA®die-away water.

5. Conduct OECD 301F: Electrolytic Respirometer Test on a) starting feed alcohols, and b) 

alcohol propoxylates, and c) the sodium salt of alcohol propoxy sulfates. 
12/14/2011 Page 11 of 66

CONCLUSIONS

1. The rate and extent of the biodegradation of alcohol propoxy sulfates, specifically 

branched C12-13 alcohol 4-mole propoxy sulfate (ALFO 23-4S) is notably less than that 

of the standard detergent surfactant alcohol ethoxy sulfate or specifically linear dodecyl 

4-mole ethoxy sulfate (POLYSTEP B-430S from Stepan Company) as shown by the 

remaining dissolved organic carbon in the ALFO 23-4S effluent from Porous Pot, DieAway, and Electrolytic Respirometer testing.

2. Acute aquatic toxicities to fish of the neat material and the effluents from Porous Pot and 

Die-Away testing were determined using standard US EPA methods. The results 

showed that both ALFO 23-4S and the control POLYSTEP B-430S had the same LC 50 

values for the neat and effluent samples despite the significant remaining dissolved 

organic carbon in the effluent of the ALFO 23-4S sample. 

3. Methylene Blue Active Substances (MBAS) analysis of both the neat and effluent 

samples proved that ALFO 23-4S undergoes primary degradation and elimination of 

surfactancy despite the significant remaining dissolved organic carbon in the effluent of 

the ALFO 23-4S sample after porous pot or 28-day die-away testing.

4. The acute aquatic toxicity to fish of the neat ALFO 23-4S is eliminated once the ALFO 

23-4S passes through the porous pot or 28 die-away test. The results of 96-hour static 

P. promelas (EPA-821-R-02-013) showed for example that the LC50 of 14% dilution for 

the ALFO 23-4S feed to the porous pot decreased to LC50 >100% dilution for the 

effluent.

5. Using ALFOTERRA® surfactants for enhanced hydrocarbon recovery (EHR) will not 

result in the discharge of any material into surface or ground water. Any rinse aid 

containing ALFOTERRA® surfactants, after processing through standard domestic or 

industrial waste water treatment, will pose no threat to the environment.

RESULTS OF EXPERIMENTS

TEST MATERIALS

 ALFOTERRA®123-4S (referred to as ALFO 23-4S) - four mole propoxy sulfate based 

on a C12-13 branched primary alcohol ISALCHEM®123. This is the product utilized in 

all of the environmental fate and effect testing. It was chosen as the representative 

structure as it is most likely the product with the highest level of concern of all 

ALFOTERRA® materials listed in the PMN - i.e. high C12 content (possible toxicity 

issues) and highest branching content (possible biodegradation issues).

 POLYSTEP B-430S (referred to as AES, Appendix 1) manufactured by Stepan - four 

mole ethoxy sulfate based on a C12-14 linear primary alcohol. This AES is a common 

anionic surfactant found in a variety of household detergent products; it is utilized as the 

comparative anionic surfactant evaluated against ALFO 23-4S. 
12/14/2011 Page 12 of 66

 ALFOL®12, LIAL®123, ALCHEM®123, ISALCHEM®123, SAFOL®23 and TDAalcohols which contain 12-13 carbons but vary in degrees of branching. These alcohols, 

their propoxylate derivatives, and the anionic sulfate (sodium salt forms) are used in a 

systematic biodegradation exercise. This biodegradation test was conducted for the 

purpose of shedding light on what aspect of the molecule’s structure promotes a

negative impact on the biodegradation profile.

General Molecular Structure: Alcohol (branched and/or linear) 4-Propoxylate Sulfate 

Surfactant

 

CH3-(CH2)m - CH-(CH2)n – CH3

|

CH2 – O - (CH2 – CH – O)y – SO3

-

Na+

 |

 CH3

Table 1 lists the trade names and descriptions of the six alcohols and propoxy derivatives 

(nonionic) evaluated in this study. Carbon chain length, degree of branching, molecular 

weights, percent propylene oxide by weight, and moles of propylene oxide are given for each of 

the six products. The red columns correspond to the alcohol and the red portions of the 

compound above while the blue refers to the propoxylate portion.

Table 1: Properties of the four mole propoxy alkoxylate derivatives (nonionic) of the six alcohols 

tested

Base Alcohol

(lab sample 

number)

alcohol

carbon 

chain

alcohol 

percent

branching

MW

propoxylate

Percent by 

weight

PO (avg.)

moles PO 

(avg.)

ALFOL®12

1314-159

C12 0% 414 54.8 3.91

ALCHEM®123

1314-48-1

C12-C13 5% 414 52.96 3.78

LIAL®123

1314-46-1

C12-C13 53% 421 53.60 3.89

SAFOL®23

1314-49-1

C12-C13 50% 430 55.0 4.08

ISALCHEM®123

1314-47-1

C12-C13 94% 425 54.12 3.96

TDA 

1314-18-1

C13 100% 435 53.5 4.01

Table 2 lists the anionic sodium salt derivatives of the six alcohol 4-mole propoxylates described 

in Table 1 as the ALFOTERRA® Surfactants. The ALFOTERRA® Surfactant based on 

ISALCHEM 123 alcohol (ALFO 23-4S) listed in bold is the primary product evaluated throughout 

this body of work. The table includes the molecular weights, percent active of the anionic 

sodium salt, percent active of the un-sulfated propoxylate and percent sodium sulfate. Water 

makes up the remaining ingredient of the ALFOTERRA® Surfactant commercial product.

alcohol (branched or linear)

sulfate

propylene oxide - avg. 4 moles 
12/14/2011 Page 13 of 66

Table 2: Properties of the anionic Surfactants based on the above propoxylates which are 

comparable to the ALFOTERRA®Surfactant 

Anionic surfactant

Alcohol Base (and lab 

numbers)

MW 

propoxylate 

sulfate

percent

anionic

active

percent unsulfated

propoxylate

percent

Na2SO4

ALFOL®12

1311-197-1

516 29.09 0.47 0.13

ALCHEM®123

1311-74-4

516 27.20 0.48 0.10

LIAL

®

123

1311-74-2

523 27.67 0.8 0.09

SAFOL®23

1311-74-5

532 27.42 0.47 0.11

ISALCHEM®123

1311-74-3

527 27.29 0.8 0.10

TDA 

1311-74-1

537 28.46 1.93 0.09

Note: Row highlighted bold - alcohol propoxy sulfate is employed in PPS, Die-Away 

studies and acute toxicity testing.

BIODEGRADATION STUDY

Porous Pot System (PPS) Biodegradation Test Method

The PPS (Figure 1) used in this study utilized the methods described in the US EPA OPPTS 

Test Guideline 835.3220 “Porous Pot Test” and ASTM test E 1798-96 “Standard Test Method 

for Assessing Treatability or Biodegradability, or Both, of Organic Chemicals in Porous Pots” 

with a series of modifications designed to reduce the amount of feed utilized. The system 

consisted of a series of six cylindrical glass containers approximately 17” x 2.75”. A constant 

temperature of 25°C was maintained inside the glass cylinders by wrapping each cylinder with 

tygon tubing held in place with duct-tape. Porous high density polyethylene (HDPE) candles 

were located on the interior of the glass containers. The candles held an average of 250 mL of 

mixed liquor activated sludge. A glass aeration line extended from the top of the glass container 

to the bottom of the candle. House air was passed through AFP Flow Meters at 100-150 

cc/min. Synthetic sewage feed was pumped into the units from three-1 gallon plastic jugs 

placed in an ice chest. For replication, two feed lines/jug delivered feed to two separate PPS by 

means of six #QG metering pumps manufactured by F.M.I., Inc. (Oyster Bay, NY). Each PPS 

was connected to a separate 1 L side-arm Erlenmeyer vacuum flask which captured effluent. 

The source of the sludge for the candles was derived from the recycled sludge from the City of 

Lake Charles, Louisiana Domestic Waste Water Plant A and the SBR aeration basin from City 

of Sulphur Regional Waste Water Treatment Plant.
12/14/2011 Page 14 of 66

 

Figure 1: Porous Pot System (PPS) Assembly Test

Synthetic sewage stock solution was made following the EPA Guideline 835.3220, except at 1⁄2 

the concentration of each component. The recipe used for the sewage stock was 65 grams 

glucose, 65 grams beef extract, 65 grams nutrient broth, 65 grams di-potassium hydrogen 

phosphate and 25 grams of ammonia nitrate. The dry ingredients were added to 2 liters of DI 

water and dissolved by stirring at 70°C. The synthetic sewage stock was refrigerated to 

maintain quality.

For the various feeds delivered to the PPS, the stock was diluted in DI water for a DOC 

concentration of 250-300 mg/L on average. The basic recipe for synthetic feed was 15 grams 

synthetic sewage concentrate, 2 grams of a 1% stock solution of Tide Cold Powder (V1255-49-

1), 10 grams of 1% stock solution of sodium sulfate. These concentrated ingredients (referred 

to as the blank) were diluted to 3000 grams with tap water. The same recipe as the blank was 

followed for the PPS test controls (sodium benzoate and AES) and study test material (ALFO 

23-4S) except for the addition of 10-12 grams of 1% solution of benzoate, AES or ALFO 23-4S 

respectively, before dilution to 3000 grams. Synthetic sewage feeds were made daily and 

refrigerated for use the following day. Sewage feeds in the ice chest were replaced daily with 

the cooled, refrigerated feed and crushed ice packed around the one gallon feed jugs to 

maintain the temperature ~ 4°C. 

Tide Cold Powder was used because the simulated sewage recipe was devoid of any 

surfactants and the ALFO 23-4S test material is an anionic surfactant somewhat similar to the 

surfactant contained in the Tide Cold Powder. As the sludge inoculate used in the PPS were 

exposed/conditioned to commercial household surfactants, it was deemed prudent to 

consistently expose them to a common household detergent product in the synthetic sewage 

recipe background.

During the PPS study it was determined that an antifoaming agent was necessary to control 

foaming and lifting of the sludge from inside the porous candles being fed by either surfactant. 

The antifoam used was J.T. Baker’s Antifoam B Silicone Emulsion; 0.3-0.6 mL was added daily 

to all feeds including blank and benzoate.
12/14/2011 Page 15 of 66

Porous Pot System Daily Operations 

The six PPS were operated from April through early October 2009. Parameters that were 

measured almost daily were hydraulic retention time (HRT), MLTSS concentrations in the PPS 

candles, TSS concentrations in the effluents, pH of effluents and DOC concentration in the feed 

and effluents. Hydraulic retention time (HRT) was maintained at 6 hours. The sludge retention 

time (SRT) was maintained between 5-10 days by wasting at least 1/10th to 1/5th of the mixed 

liquor volume each day. The numbers in the following figures refer to the 6 pots run in parallel

with three feed materials (duplicate of each) at any time.

In Figure 2, the out-of -range points are times when the feed line clogged and flow was reduced. 

During the first part of July, the fritted glass aerators were changed to a larger size as poor 

mixing was observed inside the candle permitting DOC breakthrough. It was realized that the 

larger glass aerators had displaced the working volume inside the candles from 250 mL to a 

new volume of 215 mL. The flows were readjusted on the FMI pumps to adjust HRT back to 6 

hrs. To gather additional information on the impact of HRT on DOC concentration of the 

effluents, the HRT was increased to 12 hrs for the last 2-weeks of the study.

In the following four tables, the legend numbers refer to each of the six candles of the PPS. The 

specific composition within the candles will be addressed later. The following four tables are 

meant to show the consistency of retention time (HRT), initial mixed liquor total suspended 

solids (MLTSS) and total suspended solids (TSS) of the PSS effluents throughout the duration 

of the study despite the candle contents.

Hydraulic Retention Time (HRT) Through Porous Pots 

0.0

2.0

4.0

6.0

8.0

10.0

12.0

14.0

16.0

6-Apr 26-Apr 16-May 5-Jun 25-Jun 15-Jul 4-Aug 24-Aug 13-Sep 3-Oct

Date

H

R

T (hours)

1 2 3

4 5 6

Figure 2: Hydraulic Retention Time (HRT) Through Porous Pot System 

Figure 2 shows the hydraulic retention time throughout the porous pot study. With the exception 

of the Zoogleal issue in early July and a secondary study in October, the HRT was maintained 

consistently at 6 hrs.
12/14/2011 Page 16 of 66

Mixed Liquor Total Suspended Solids (MLTSS) in Porous Pot Candle

Time period from April through October 2009

0

1,000

2,000

3,000

4,000

5,000

6,000

7,000

8,000

9,000

10,000

6-Apr 16-

Apr

26-

Apr

6-

May

16-

May

26-

May

5-Jun 15-

Jun

25-

Jun

5-Jul 15-

Jul

25-

Jul

4-Aug 14-

Aug

24-

Aug

3-Sep 13-

Sep

23-

Sep

3-Oct 13-

Oct

Date

mg/L

1 2 3

4 5 6

Figure 3: Mixed Liquor Total Suspended Solids (MLTSS) in Porous Pot Candle

Figure 3 depicts the variation in the concentration of the MLTSS during the study period. During 

June, the MLTSS composition became enriched with bacteria species called Zooglea. This 

species occurs in waste treatment plants and becomes a nuisance when its numbers bloom; the 

bacteria produce a heavy slime layer on its surface resulting in a jelly like consistency in the 

sludge. This jelly like consistency occurred in the sludge for all the candles and increased the 

viscosity of the mixture. MLTSS concentrations were also increasing during this period. The 

aerators in the candles were not sufficiently large to accommodate this increased viscosity and 

aeration/mixing was impaired. On July 4th, half of the volume of MLTSS was wasted to try and 

control the problem. Increased wasting continued (i.e. shortening SRT) to try and control the 

Zooglea. 
12/14/2011 Page 17 of 66

Porous Pot Effluent TSS

-10

10

30

50

70

90

1-Jun 11-Jun 21-Jun 1-Jul 11-Jul 21-Jul 31-Jul 10-Aug 20-Aug 30-Aug 9-Sep 19-Sep

Date

TSS (mg/L)

1 2 3

4 5 6

0

Figure 4: Porous Pot Effluent TSS

Figure 4 depicts the TSS in the effluents during the trial; effluent was disposed daily. The period 

in June reflects what occurred in the candles during the Zoogleal bloom - increased viscosity 

and poor aeration/mixing. Breakthrough of DOC into the effluent vessels allowed continued 

biodegradation and increased microbial growth, thus, TSS in effluents was going through an 

atypical increase. The porous pot has the capability to block MLTSS from escaping into the 

effluent. After correcting the problem in the candles, TSS in the effluents returned expected and 

stabilized values for the duration of the test (July - Sept). TSS values occurring below 0 mg/L 

TSS are common on the occasions that fibers on the filter pads break off; it is then lost during 

the filtering process. 

Figure 5. Porous Pot Effluent pH Values

5.00

5.50

6.00

6.50

7.00

7.50

8.00

8.50

9.00

1-Jun 21-Jun 11-Jul 31-Jul 20-Aug 9-Sep 29-Sep

pH

1 2 3

4 5 6

Figure 5: Porous Pot Effluent pH Values
12/14/2011 Page 18 of 66

Figure 5 depicts the pH values in the effluents during the trial; the starting pH of the feeds 

averaged 7.5. The drop in the effluent pH for units 3 and 6 were those units treating the ALFO 

23-4S test material. This decrease began occurring after the concentration of the test material 

was increased. The drop in pH is indicative of partial/incomplete breakdown of the parent 

material into fatty acids. To control/buffer the pH, 0.5 grams of sodium bicarbonate was added 

each day to all the feeds. The addition of the buffer raised the pH slightly to 7.6. This buffer 

addition controlled the effluent pH and was continued through the rest of the study period. 

 

Figure 6. Porous Pot DOC Feed and Effluents

0

50

100

150

200

250

300

350

400

5/7/2009 6/6/2009 7/6/2009 8/5/2009 9/4/2009 10/4/2009

D

O

C

mg/L

1 Effluent

2 Effluent

3 Effluent

4 Effluent

5 Effluent

6 Effluent

Feed 

Feed

Feed 

Figure 6: Porous Pot DOC Feeds and Effluents

Figure 6 depicts the PPS DOC values for the three feeds (blank, benzoate, and sample) in 

blues and the six effluents during the period of mid-May through early October when the study 

ended. The effluent DOC values reflect a loss of steady state from the end of May through early 

June - the period of the Zoogleal bloom. Effluent DOCs were returning to a steady state in late 

June but began to elevate when the HRT decreased due to volume displacement from the new 

larger aerator/diffusers. Effluent values returned to steady state by early August when the feed 

pumps were adjusted to correct the HRT back to 6 hrs. In general, during periods of steady 

state (mid July through October), DOC removal rates for all units was above 90%. This 

compares to the typical removal efficiency seen in domestic waste treatment plants.

Porous Pot System Experimental Results

The primary objective of the PPS was to assess the fate of ALFO 23-4S after processing 

through a simulated waste water treatment process. Additionally, there was interest in 

developing a deeper understanding of the degradation mechanism of the alcohol propoxy 

sulfate molecules and how this degradation compares to that of standard alcohol ethoxy 

sulfates. As a result, the control materials, HRT rate, and bacteria conditions were 

systematically varied throughout the study. In all situations, samples were run in duplicate.

 
12/14/2011 Page 19 of 66

The next series of figures demonstrates the response of the effluent DOC values, especially 

those for the ALFO 23-4S, at each specific test condition evaluated.

Porous Pot Effluents DOC Pre Unacclimated ALFO 23-4S

0

25

50

75

100

125

150

7-May 17-May 27-May 6-Jun 16-Jun 26-Jun 6-Jul 16-Jul 26-Jul

Date

mg/L D

O

C

Effluent 1 Effluent 2 Effluent 3

Effluent 4 Effluent 5 Effluent 6

feed increased to 15 

mg/L active

feed increased to 30 mg/L 

active in #2 and #5

Figure 7: Porous Pot Effluents DOC

Figure 7 shows the DOC values for the first three months of the PPS study. The effluent from 

the ALFO 23-4S candles are depicted in brown (effluent 3 and 6), the effluent from the benzoate 

candles are depicted in green (effluent 2 and 5), and the effluent from the blank candles are in 

blue (effluent 1 and 4). 

The study was started in mid May with the addition of benzoate and ALFO 23-4S to blank 

candle that had been acclimating since mid-April. Shortly after this addition, the DOC values of 

the ALFO 23-4S started to climb relative to the blank and benzoate samples. There was then a 

period of upsets (foaming, Zooglia, aeration changes) that impacted the effluent quality as seen 

by the erratic DOC values throughout June and early July. The system was brought under 

control around July 5 and a steady state was reestablished by July 8. On July 9th the 

concentrations of benzoate and ALFO 23-4S were increased to 15 mg/L as active. Shortly 

after, an increase in the DOC of the ALFO 23-4S effluent was again observed suggesting a 

difference between the degradation rate of the ALFO23-4S and the benzoate. The benzoate 

DOC curve (green) is identical to the blank (blue) under steady state conditions. 

To validate the increased DOC of the ALFO 23-4S, the benzoate was replaced with ALFO 23-

4S in the feed of candles 2 and 5 (green on graph) on July 30. Additionally, the ALFO 23-4S 

concentration was raised in candles 2 and 5 to 30 mg/L as active or approximately 13 mg/L as 

DOC. Immediately the DOC in the effluent of candles 2 and 5 increased to values equal to the 

original ALFO 23-4S candles (3 and 6). 
12/14/2011 Page 20 of 66

Figure 8. Porous Pot DOC: ALFOTERRA

Un-acclimated and Acclimated Response

0

5

10

15

20

25

30

28-Jul 30-Jul 1-Aug 3-Aug 5-Aug 7-Aug 9-Aug 11-Aug

Date

mg/L D

O

C

Blnk U-Alfo A-Alfo

Blnk U-Alfo A-Alfo

Figure 8: Porous Pot Effluents - Un-acclimated and acclimated Response 

Figure 8 shows in more detail the change in effluent DOC values when ALFO 23-4S is added to 

what was originally the benzoate candles (now labeled U-ALFO for un-acclimated). Most 

interesting is that the original ALFO candles and the U-ALFO candles have exactly the same 

effluent DOC values, thus confirming the validity of the response. 

Figure 9. Return to Steady State: Post Unacclimated Alfoterra Pre AES 

0

5

10

15

20

25

30

7-Aug 9-Aug 11-Aug 13-Aug 15-Aug 17-Aug

DOC mg/L

Benz Alfo Blnk

Benz Alfo Blnk

Figure 9: Return to Steady State (DOC)
12/14/2011 Page 21 of 66

As a final confirmation of the DOC response to all samples (benzoate, blank, ALFO) the feed 

conditions were changed such that the candles that were fed ALFO 23-4S from the beginning of 

the study were now fed blank feed only (yellow and red above), The candles that were fed blank 

feed from the beginning of the study were now fed benzoate (light blue, dark blue), and the 

candles that were originally benzoate but had been switched to ALFO 23-4S feed was kept with 

the ALFO 23-4S (pink, purple). Both the benzoate and the ALFO 23-4S feed had 30 mg/L as 

active added during this period. Figure 9 illustrates that the effluent DOC levels for the new 

benzoate and new blank (formerly acclimated ALFO 23-4S) rapidly merged together within a 

few days confirming that the PPS reacts rapidly to the feed changes and the response is real.

Porous Pot DOC: A Comparison of 1214E4S and ALFO 23-4S 

0

5

10

15

20

25

30

13-Aug 23-Aug 2-Sep 12-Sep 22-Sep

Date

D

O

C

mg/L

AES Alfo Blnk

AES Alfo Blnk

Feed changed from 30 

to 40 mg.L as active

Figure 10: Porous Pot Effluent DOC: A Comparison of AES and ALFO 23-4S

Now that the PPS response for ALFO 23-4S had been established and verified, there was 

interest in knowing how this response compared with the common detergent surfactant alcohol 

ethoxy sulfate (AES). POLYSTEP B-430S (linear C12-14 alcohol based 4-mole ethoxy sulfate 

from Stepan Co.) was added as the AES control replacing benzoate. The concentration for both 

the AES and ALFO 23-4S was 40 mg/L as active or influent DOC values of 17 mg/L and 16 

mg/L respectively. At the end of this period (Sept. 24th), the effluent DOC values for AES (dark

blue, light blue) were approximately equal to the blank (brown, yellow) while the ALFO 23-4S 

(pink, purple) maintained its slightly higher effluent DOC response. 

This difference in the effluent response of the ALFO 23-4S and the AES was not unexpected. 

There are multiple literature references, discussed in later sections of this report, that discuss 

the how branching impedes the biodegradation response of molecules.
12/14/2011 Page 22 of 66

Porous Pot: Increased Hydraulic Rentenion Time

(6hr-12hr)

0

5

10

15

20

25

30

19-Sep 24-Sep 29-Sep 4-Oct 9-Oct

Date

D

O

C

mg/L

AES Alfo Blnk

AES Alfo Blnk

HRT from 6hr to 12hr

Figure 11: Porous Pot Effluent: Increased HRT effect on DOC

Finally, there was interest in seeing if the biodegradation of the ALFO 23-4S would be increased 

if the HRT was increased. Figure 11 illustrates the effluent DOC response when the treatment 

time (HRT) was doubled from 6 to 12 hrs. The FMI pumps were turned down (reducing feed 

rate) gradually each day starting on September 25th and completed on September 30th. The 

data shows that the effluent DOC for ALFO 23-4S (pink, purple) moves towards the 10 mg/L 

baseline established for the DOC effluents of the AES (light blue, dark blue) and the blank 

(brown, yellow) confirming that degradation of ALFO 23-4S continues with increased time. The 

PPS study ended on October 10th.

PPS Results Summary:

The results of the PPS study showed that even after processing ALFO 23-4S through a 

simulated waste-water treatment, a predominate amount of the original DOC remains in the 

effluent. This led to two questions; 1) will the ALFO 23-4S continue to degrade after leaving a 

treatment facility and reaching receiving waters and, if so, what will the extent and timing of the 

degradation be? And 2) is the ALFO 23-4S sufficiently broken down through waste water 

treatment so that it no longer has the aquatic toxicity profile as the parent product? The 

questions are addressed in the next two sections of this report.

OECD 301 A: DOC Die-Away Experimental Results

To determine the biodegradation rate of ALFO 23-4S DOC effluent, the effluent from the PPS 

was subjected to a DOC Die-away test. This test was used to simulate degradation that could 

be expected if the PPS effluent reached receiving waters. 

Using OECD 301A: DOC Die-Away, experiments were conducted on the effluents from the PPS 

system during the period of AES and ALFO 23-4S comparison. In conjunction, a DOC Die-
12/14/2011 Page 23 of 66

Away test was conducted on benzoate and the neat or parent compounds for the AES and 

ALFO 23-4S (before PPS treatment). 

The effluents were collected from the PPS for two consecutive days on September 10th and 11th. 

The effluent duplicates were combined and refrigerated on September 10th and then added to

the combined duplicates from September 11th. The pooled effluents for the blank, AES and 

ALFO 23-4S were each added to 2-liter Erlenmeyer flasks and placed on a bench top shaker for 

mixing.

To make the neat samples, on the same day (September 11th), mineral water solution was 

added to six 2-liter Erlenmeyer flasks. Using 1% stock solutions for benzoate, AES and ALFO 

23-4S, additions were made to each flask for a concentration of 50 mg/L active for each test

compound; each test material had a duplicate flask. The flasks were placed on the bench-top 

shaker and mixed at 125 rpm.

Day -0 sampling for DOC occurred before the inoculate concentrate was added to each vessel. 

The inoculate source was equal parts of aeration sludge from City of Sulphur Regional Waste 

Water Treatment Plant and soil slurry supernatant. The inoculate concentrate was added to 

each flask to a diluted target concentration of 25 mg/L TSS.

Die-Away Porous Pot Effluent

0

5

10

15

20

25

30

0 5 10 15 20 25 30

Days

D

O

C (mg/L)

PP Blank PP AES PP ALFO

Figure 12: Die-Away Porous Pot Effluent

Results for the PPS effluent Die-Away test can be seen in Figure 12. For all samples, the DOC

continued to decline over the 28-day period. However, the degradation of the ALFO 23-4S

(green) never reaches that of the blank (blue) or the AES (pink) in the time allotted. 
12/14/2011 Page 24 of 66

OECD Die-Away DOC Loss

0

20

40

60

80

100

0 5 10 15 20 25 30

Days

%

D

O

C Loss

AES ALFO Benz

Figure 13: OECD 301A Die-Away DOC Loss of neat ALFO 23-4S, AES, and Benzoate.

The die-away test was conducted on the neat ALFO 23-4S, AES, and benzoate (not subjected 

to porous pot). In this case, the difference in the degradation rate of the ALFO 23-4S and the 

AES is obvious. Benzoate and AES showed a rapid loss in DOC reaching 100% DOC loss by 

day 17 whereas the ALFO 23-4S had a modest DOC decline with 20% DOC loss at day 28.

MBAS Analysis

In order to determine if primary degradation is occurring on the ALFO 23-4S during the porous 

pot test and the die-away test, Methylene Blue Active Substances (MBAS) analysis was run on 

the influents and effluents. Samples were preserved with formaldehyde for a final concentration 

of 1% preservative. 
12/14/2011 Page 25 of 66

Porous Pot: MBAS Primary Biodegradation 

0

5

10

15

20

25

30

35

40

45

Blank AES ALFO 23-4S

MBAS (mg/L)

Influent

Effluent

Figure 14: Porous Pot MBAS Primary Biodegradation

Figure 14 shows the MBAS results for the influent and effluent for the porous pot study. The 

small MBAS activity in the blank is attributed to the surfactant in the representative Tide Cold 

Water powder that was added to all three PPS feed, as discussed earlier. A significant 

decrease in surfactancy can be seen for both the AES and ALFO 23-4S after treatment through 

the PPS. The green bar represents the PPS influent feed (40 mg/L as active for AES and ALFO 

23-4S) while the blue bar represents the respective effluent All samples were collected from 

the PPS on September 11th. The MBAS percent reduction in activity for the AES and ALFO 23-

4S during the PPS were 99% and 94% respectively.

Figure 15. Comparison Porous Pot and Die-away 

0

10

20

30

40

50

Porous Pot Die-away

MBAS mg/L

Influent Effluent

Figure 15: ALFO 23-4S Comparison MBAS Porous Pot and Die Away

Figure 15 compares the influent and effluent MBAS results for ALFO 23-4S through both porous 

pot and die-away degradation methods. In both cases, the influent contained approximately 40 

mg/L as active ALFO 23-4S. The Porous pot effluent was generated after a 6hr HRT while the 
12/14/2011 Page 26 of 66

die-away effluent was taken after 28 days. The MBAS percent reduction in activity for ALFO 23-

4S in the PPS was 94% and 90% in the 28-day Die-Away. 

MBAS Results Summary:

MBAS analysis of both the PPS and Die-Away test methods confirm that the ALFO 23-4S 

molecule undergoes significant primary biodegradation though most of the original DOC 

remains. The question that that remained unanswered was - is the residual ALFO 23-4S 

material still aquatically toxic?

Aquatic Acute 96hr Fathead Minnow Results

A 96-hour static P. promelas (EPA-821-R-02-013) test was conducted by Environmental 

Enterprises USA, Inc. (EE USA) during the PPS and Die-Away study period. Water samples of 

the PPS synthetic sewage feeds (blank, AES + blank, and ALFO 23-4S + blank), neat AES, 

neat ALFO 23-4S (40 mg/L as active), and PPS effluents (blank, AES and ALFO 23-4S) were 

sent for toxicity testing on September 9. Replicate effluents were combined and represented 

two days (combined) to achieve the required volume needed for 5 replicates /concentration in 

the fathead minnow test. The toxicity endpoints for water samples represented in the PPS 

study period is reported as the percent dilution of the test solution, while AES and ALFO 23-4S 

neat were compared at a “as is” concentration of 40 mg/L with toxicity endpoints reported mg/L. 

On October 9th, at the end of the 28-day DOC Die-Away for ALFO 23-4S, the remaining solution 

was put on ice and sent for 96-hour static minnow test. At this same date, a fresh sample of 

ALFO 23-4S at 50 mg/L was made and shipped on ice to EE USA to use as a control. The 

results from the testing are given below in Table 3.

 

Sample 

Description

Environmental 

Enterprise USA

Sample ID

Survival 

NOEC 

%

Survival 

LOEC %

LC50 %

(95% Confidence Interval)

PPS Blank Influent 

Feed

Q-642-09-1336-

38-1

12% 25% 14.7 %

(12.4% - 17.5%)

PPS AES Influent 

Feed

Q-643-09-1336-

38-2

3.0% 6.0% 5.47%

(4.91% - 6.10%)

PPS ALFO 23-4S 

Influent Feed

Q-644-09-1336-

38-3

6.0% 12.0% 9.48% 

(8.28% - 10.9%)

AES Neat

(40 mg/L active)

Q-645-09-1336-

38-4

6.0%

(2.4

mg/L)

12.0%

(4.8 mg/L)

13.8% (5.52 mg/L)

(12.3% - 15.5%)

ALFO 23-4S 

(40 mg/L active)

Q-646-09-1336-

38-5

3.0%

(1.2

mg/L) 

6.0%

(2.4 mg/L)

7.76% (3.10 mg/L)

(6.64% - 9.08%)

PPS Blank Effluent Q-647-09-1336-

38-6

100% >100% >100%

(NA)

PPS AES Effluent Q-648-09-1336-

38-7

100% >100% >100%

(NA)

PPS ALFO 23-4S 

Effluent

Q-649-09-1336-

38-8

100% >100% >100% 

(NA)

ALFO 23-4S 

Die-Away Influent

(50 mg/L) 0-day

Q-715-09-1336-

58-2

6%

(3.0

mg/L)

12%

(6.0 mg/L)

8.8% (4.4 mg/L)

(8.0% - 9.7%)

ALFO 23-4S 

Die-Away Effluent

(50mg/L) 28-day

Q-716-09-1336-

58-1

100% >100% >100%

NA
12/14/2011 Page 27 of 66

Table 3: Results of fathead minnow 96hr acute toxicity test (see Appendix 23)

Note: numbers highlighted in bold are Sasol NA notebook number. Concentrations (mg/L) in red 

were calculated as mg/L in the endpoint dilution value.

Aquatic toxicity results from Table 3 are depicted in the following Figures -16, 17, 18.

AES and ALFO 23-4S

Acute Toxicity 96 hr Fathead Minnow

0.0

2.0

4.0

6.0

8.0

10.0

LC50 LOEC NOEC

Toxicity Endpoint

mg/L

AES ALFO 23-4S

 

Figure 16: AES and ALFO 23-4S Neat Material Acute Toxicity Before Any Biodegradation

Figure 16 is a comparison of aquatic toxicity for neat AES and ALFO 23-4S surfactants before 

any biological treatment. The toxicities for each endpoint are typical of anionic surfactants. 

ALFO 23-4S has slightly higher end points than the AES suggestion that the neat AFLO 23-4S 

is slightly more aquatically toxic than the neat AES. The LC50 for the neat, non-biologically 

treated ALFO23-4S was 5.7 mg/L.
12/14/2011 Page 28 of 66

Figure 17. Porous Pot: Aquatic Toxicity Test 96hr Minnow

40mg/L AES and ALFO active

0

20

40

60

80

100

120

Blank Feed Blank Effluent AES+Feed AES Effluent ALFO+Feed ALFO Effluent

Source

Percent S

ource

LC50

LOEC

NOEC

Figure 17: Porous Pot Feed and Effluent: Aquatic Toxicity AES and ALFO 23-4S

Figure 17 compares the toxicity results before and after the PPS. It shows that, in all cases, 

including the ALFO 23-4S, the aquatic toxicity is eliminated after a 6 hr retention time through 

the porous pot. All three PPS effluents show no acute toxicity for the toxicity endpoints of LC50, 

NOEC and LOEC - all values being greater than 100% effluent.

At the end of the 28-day Die-Away test on October 9th, the water remaining in the 2-liter 

Erlenmeyer shake flask for ALFO 23-4S was sent for toxicity testing. In addition, a sample of

neat ALFO 23-4S (50 mg/L as active), reflecting the Die-Away concentration before bacterial 

activity day-0, was sent for testing.
12/14/2011 Page 29 of 66

Figure 18. OECD Die-Away Test ALFOTERRA (50mg/active or 22mg/L DOC)

0

20

40

60

80

100

120

LC50 LOEC NOEC

Toxicity Endpoint

Percent S

ource

ALFO Day 0 ALFO Day 28

calculated LC50 

= 4.4 mg/L

Figure 18: Acute Toxicity Test, Die-Away Test ALFO 23-4S

Acute toxicity of ALFO 23-4S before and after the 28-day Die-Away test was determined. 

Figure 18 compares the toxicity from 96-hr acute minnow of Die-Away water at the end of the 28 

days (blue) with that at the beginning of the test (green). Two sets of water samples, Day 0 

(made identical to the ALFO 23-4S test water at the start containing 50 mg/L ALFO 23-4S) and 

the day 28, were evaluated. The results are given as the calculated dilution percent of the two 

water samples. The calculated concentration for the LC50 endpoint for the 0-day sample was 

12% dilution or 4.4 mg/L - similar to the toxicity observed from the neat in the PPS toxicity tests. 

No toxicity remained in the ALFO 23-4S Die-Away 28-day water and toxicity endpoints for LC50, 

LOEC and NOEC exceeded 100% effluent solution as provided. 

Fat Head Minnow Results Summary:

Three test methods (Porous Pot, Die-Away, Electrolytic Respirometer (from previous report))

have shown that ALFO 23-4S only loses ~30% of its original DOC values at the conclusion of 

the tests suggesting alcohol propoxy sulfates do not biodegrade well. However, MBAS confirms 

that at least primary degradation is occurring. More importantly, Acute Aquatic Testing on Fat 

Head Minnows proves that the ALFO 23-4S residual DOC in the effluent after standard 

biodegradation treatment has lost the aquatic toxicity of the parent molecule.

OECD 301F Biodegradability Electrolytic Respirometer Study 

Though the above data suggests that with standard water treatment methods, ALFO 23-4S and 

similar molecules pose little-to-no threat the aquatic life, there is interest in understanding why

the residual DOC in ALFO 23-4S effluent after standard biological degradation testing is so 

much higher than for AES. To better understand the role of chemical structure on the 
12/14/2011 Page 30 of 66

biodegradation mechanism of alcohol propoxy sulfates, Electrolytic Respirometer (OECD 301F)

biodegradation studies were conducted on alcohol propoxy sulfate raw materials with a known 

structure. As detailed in Table 1 & 2, six alcohols in the C12-C13 carbon range with varying 

degrees of branching were studied. These six alcohols were then propxylated to an average of 

four moles of propylene oxide using KOH catalyst and studied again. Finally, the six alcohol 

propoxylates were sulfated using a thin-film sulfation method to yield the sodium salt derivatives

and studied again. Sodium benzoate was used as the standard in each test.

The biodegradation tests were performed quadruplicate in using two 20 channel Coordinated 

Environmental Services (CES) Ltd (Cornwell, UK) aerobic electrolytic respirometer units 

following OECD 301F guidelines with slight modifications. The CES electrolytic respirometer 

unit consists of 20 one-liter vessels fitted with heads containing O2 and CO2 sensors. The 

system measures the amount of CO2 generated by means of a cuvette filled with 1% sodium 

hydroxide and a conductivity probe that determines the change in conductivity as CO2 is 

absorbed from the headspace gases of the test vessel. A differential pressure sensor head is 

designed to measure the amount of O2 used by respiring microorganisms. It achieves this by 

detecting the difference in pressure during the uptake of oxygen and generation of CO2 and 

then supplying an equal amount of O2 back to the headspace using an electrolytic cell.

The electrolytic cell contains a chamber filled with copper sulfate solution. Oxygen is generated 

in this chamber by passing an electrical charge (initiated by the test vessel head pressure 

change sensor) between electrodes. It is connected to the headspace of the sample vessel by 

a vent. Valves are used to balance the pressure and return generated O2 to the headspace of 

the test vessel. The amount of CO2 (mg) absorbed and O2 (mg) generated is recorded by a 

computer.

The quadruplicate samples were split between the two respirometer units; two replicates in Unit 

1 and two replicates in Unit 2. 

As stated above, slight deviations were made to some of the OECD 301F guidelines8to better 

accommodate the poor water solubility of the alcohols and alcohol propoxylates evaluated in 

this study. The inoculum for these tests was derived from the recycled sludge from the City of 

Sulphur, LA Regional Waste Water Treatment Plant; sludge was SBR aeration basin. The 

sludge was allowed to settle and supernatant decanted and refilled with mineral water to dilute 

any background organic. This was repeated three times. The final decant sludge concentrate 

was blended using a Waring Blender for two minutes on low. Total suspended solids (TSS) 

were measured on the concentrate and the volume determined to add to the large pot of 45 

liters of mineral water. The TSS in the medium is targeted to a final concentration of 25 mg/L 

TSS to meet the OECD guideline specification of < 30 mg/L TSS. Sodium benzoate (source: 

J.T. Baker – 99% purity) was used as the positive reference test material. From the 45 liter pot 

of inoculum containing mineral water is 1kg is dispended into the 1-liter test vessels. 

The following describes the technique used for dispersing water insoluble alcohols and alcohol 

propoxylates for biodegradation testing. Approximately 500 mg mound of silica gel (Aldrich, 

Merck grade 9385) with a mesh size of 230-400 Ǻ was weighed onto a creased piece of 

weighing paper. A spatula was then used to tap the top of the mound and flatten it so that it 

became a thick “plateau” to prevent drops of insoluble test material from rolling off of the silica 

gel. Then, a 1 mL pipette was held vertically and individual drops of insoluble test material were 

dispensed onto the silica gel until a weight close to 50-60 mg was reached. Special care was

taken to ensure that drops were not placed on top of each other so that none of the insoluble 

solution soaked down to the weighing paper. The weight of the insoluble test material was then 
12/14/2011 Page 31 of 66

recorded. The sodium benzoate powder (50-60mg) was added to the 500 mg silica gel in the 

same manner. The weighing paper was then rolled into a cone and the silica gel/test material 

was poured into a 125 mm x 15 mm test tube. The silica gel/test material was mixed using a 

VWR Scientific Products, Mini Multiple Vortexer MV1, set at highest speed (2500 rpm) for fifteen 

minutes. 

The 1-liter test vessels containing 1kg diluted inoculate in the mineral medium were placed on a 

magnetic stirrer and stirred prior to receiving the contents from the test tubes. The silica gel/test 

material samples were then poured into the test vessels. A uniform dispersion of the test 

material was immediately observed in all units and remained as such throughout the entire 28-

day test. The 1-liter vessels were then placed in the water bath (25 ± 1°C) of the electrolytic 

respirometer units. A higher temperature (25 instead of 22°C) was used for consistency with 

the PPS temperature of 25°C; also, two of the alcohols (ALFOL®12 and ALCHEM®) are solids at 

the guideline 22°C. The respirometer underwent a 60 minute initialization/equalization before 

data collection began at every six hour interval. Both CO2 generation and O2 uptake data were 

recorded every six hours during the 28 day test. Each flask contained a stir bar positioned over 

a magnetic stirrer to provide vigorous mixing. Blanks were also included in the study for validity 

purposes. These blanks were prepared identically to the samples (i.e. test medium, inoculum, 

and silica gel) but without the test compounds. 

The procedure for the six anionic ALFOTERRA

® Surfactants is achieved by adding 10 mL from 

a 1% stock solution for each anionic surfactant into the 1-liter vessel containing 990g of 

inoculated mineral water. Final concentration targeted is 100 mg/L as active. Since these are 

anionic surfactants, the silica gel dispersion technique did not apply.

The results for theoretical oxygen demand (ThO2) is obtained by the COD wet chemistry HACH 

method utilizing low range 0-150 mg/L COD vials and a DR/2000 spectrophotometer. The 

results reported as ThO2 is determined by generating COD values on specific weight from each 

test material, including benzoate. The mg O2 / mg sample factor used was calculated by 

dividing the COD mg/L value by the mg/L material used in the COD analysis. The THO2 value 

used for each material is the recorded weight in mg of sample multiplied by the mg O2 / mg

added to each 1-liter test vessel.
12/14/2011 Page 32 of 66

OECD 301 F Electrolytic Respirometer Results:

ALFOTERRA Hydrophobe Biodegradation Study

OECD 301F Theoretical Oxygen demand

0

20

40

60

80

100

120

0 5 10 15 20 25 30

Days

% THO

D

Isalchem 23 Alchem 23

TDA Safol 23

Lial 23 Alfol 12

Benzoate

Figure 19: Biodegradation Study - Alcohols (OECD 301F)

The 28-day biodegradation results for the six alcohols are shown in Figure 19. As expected, the

degradation rate decreases as the amount of branching in the alcohol increases. This trend is 

especially true for the first 10 days where branching content appears to be the primary driver.

The 100% linear ALFOL®12 (dark green) gives the fastest and most complete biodegradation 

while the 100% branched TDA Alcohol (red) gives the slowest. The alcohols that are a mixture 

of both linear and branched molecules fall in between. The only anomaly is the ALCHEM®

alcohol (brown) which is 100% linear and performs equal to the 100% linear ALFOL®12 at the 

beginning of the study but does not reach the same 100% THOD after the 28 days. This can 

be explained by the C13 content in the Alchem which must degrade slower than the 100% C12 

content of the ALFOL®12. 

Also given in Figure 19 is a 3-D model of the ALFOTERRA® Surfactant molecule. The circled 

portion denotes the alcohol hydrophobe of the surfactant, which in the given case is 

ISALCHEM®123 alcohol. From the above data it can be concluded that the ISALCHEM®123 

alcohol portion of the ALFOTERRA® molecule could impact the biodegradation in slightly 

negative way due to its branching effect, especially when compared to the 100% linear C1214

alcohol derived AES that was used in the earlier discussed studies as the control. However, 

since all alcohols easily reached the 60% ThO2 threshold by day 28, the alcohol structure does 

not completely explain the poorer biodegradation performance of the ALFO 23-4S. 
12/14/2011 Page 33 of 66

ALFOTERRA Hydrophobe 4PO Biodegradation Study

OECD 301F Theoretical Oxygen demand

0

20

40

60

80

100

120

0 5 10 15 20 25 30

Days

% THO

D

Isalchem 23 Alchem 23

TDA Safol 23

Lial 23 Alfol 12

Benzoate

Figure 20: Biodegradation Study - Alcohol 4 Mole Propoxylates (OECD 301F)

In order to determine the effect of the propoxylate portion on the biodegradation of the 

ALFOTERRA® molecule, the six alcohols were propoxylated using KOH catalyst to contain an 

average 4 moles propylene oxide. Figure 20 shows the resulting degradation curve after 28 

days in the electrolytic respirometer. The most notable difference between the alcohol (Figure

19) and alcohol propoxylate (Figure 20) response curves is that the degradation of the 

propoxylates within the first 10-15 days is generally slower than that of the alcohol. Secondly, 

the branching in the alcohol back-bone has less of an impact on the results. In fact, if not for the 

slower start of the TDA alcohol and the lower final THOD of the ALFOL®12 alcohol, all samples 

would have performed statistically the same. Most surprising is that all samples biodegrade to 

>80% in 28 days. Again, this leads one to conclude that the alcohol and propoxylate have only 

a slight impact on the poor degradation of the ALFO 23-4S.

The ALFOTERRA® Surfactant molecule image in the figure now shows the alcohol propoxylate 

portion of the molecule circled. The portion circled is a nonionic surfactant that has both a 

hydrophobic (alcohol chain) tail and hydrophilic head group of a polypropylene glycol (PPG) 

chain. The PPG head group extends the hydrophobic property than imparts surfactancy. Thus, 

the alcohol propoxylate has poor water solubility. 
12/14/2011 Page 34 of 66

ALFOTERRA Hydrophobe 4PO Sulfate Biodegradation Study

OECD 301F Theoretical Oxygen demand

0

20

40

60

80

100

120

0 5 10 15 20 25 30

Days

% THO

D

Isalchem 23 Alchem 23

TDA Lial 23

Safol 23 Alfol 12

Benzoate

Figure 21: Biodegradation Study - Alcohol 4 Propoxy Sulfate (OECD 301F)

Finally, the sulfate (sodium salt) derivatives of the six alcohol propoxylates (4 moles) were 

evaluated through electrolytic respirometer. The results are given in Figure 21. Now, the poor 

biodegradation associated with the alcohol propoxy sulfate is seen; the addition of the sulfate 

head-group or the interaction of the head-group and the branched alcohol propoxylate

significantly decreases the biodegradability of the molecule. This is somewhat surprising as the 

sulfate molecules are extremely water soluble. In general, one would expect that water soluble 

molecules are more bio-available and would be much easier for the microbes to degrade than 

insoluble molecules such as the alcohol propoxylate. However, this is not the case with this 

molecule. 

Again, there is some impact of branching on degradation. The molecule based on 100% 

branched TDA alcohol (red) is the slowest to degrade while the 100% ALFOL®12 alcohol (dark

green) based product is the fastest. However, the branching affect is not so pronounced that 

the biodegradation of the ALFO 23-4S could be much improved by going to a 100% linear 

molecule. 

OECD 301 F Electrolytic Respirometer Summary:

Though branching on the back-bone of the alcohol hydrophobe and the addition of propylene 

oxide does impact the biodegradation of alcohol propoxy sulfates when compared to the 

industry standard linear alcohol ethoxy sulfates, it is the interaction of the sulfate head group

with the branched alcohol propoxylate that is responsible for the decrease in biodegradation. 

To easily see the impact of each structural component for the model compound ALFO 23-4S, 

the biodegradation curves for the ISALCHEM 123 alcohol, ISALCHEM 123-4PO and 

ISALCHEM 123-4PO-S (commercial name is ALFOTERRA®123-4S) are given in Figure 22. 
12/14/2011 Page 35 of 66

Biodegradation Profile of Isalchem 123 Derivatives

OECD 301F Theoretical Oxygen Demand 

0

20

40

60

80

100

0 5 10 15 20 25 30

Days

%

T

H

O

D

Isalchem123-4PO

Isalchem 123-4POS

Isalchem 23 OH

Alcohol

4PO

4POS

Figure 22: Biodegradation profile of ALFO 23-4S Derivatives

Discussion

Environmental Fate and Effects of ALFOTERRA® Surfactants

The results of the PPS studies reveal that DOC removal of ALFO 23-4S through the simulated 

waste treatment plant is inadequate. At best there was a DOC loss of 60%. It was also shown 

that the level of treatment reached a steady state and leveled off within a few days. The 

standards (benzoate and the model anionic AES surfactant) showed excellent to complete 

removal as compared to the blank. The poor performance in biodegradation of ALFO 23-4S is 

also supported by the DOC Die-Away and the electrolytic respirometer results. 

In the PPS and DOC Die-Away studies, more than 90% of MBAS activity of the ALFO 23-4S is 

lost. This demonstration of high degree of primary degradation for the surfactant molecule is 

also shown to have occurred in the model AES surfactant. The degree of primary 

biodegradation for ALFO 23-4S and AES surfactants is similar. 

Further evidence that primary degradation or breakdown of the parent molecule is occurring 

comes from the aquatic toxicity testing. The 96-hour minnow acute toxicity LC50 of < 5 mg/L for 

both the neat ALFO 23-4S and AES are typical concentrations for anionic surfactants. 

However, the significantly improved acute aquatic toxicity in the effluents from the PPS and DieAway at 28-days indicates that the parent molecule for ALFO 23-4S has undergone significant 

primary degradation so as to no longer be toxic. If the DOC that remains in both the PPS 

effluent and Die-Away of the ALFO 23-4S was intact parent compound, the resulting effluent 

solution would have been toxic for at least the 100% solution where the residual DOC is about 

15 mg/L greater than the control samples - clearly over the LC50 point. However, the 

remaining DOC is incapable of short term toxicity which is demonstrated by the endpoints of 

LC50, NOEC and LOEC of greater than 100% effluents. 
12/14/2011 Page 36 of 66

Biodegradation Pathway of Anionic Surfactants Studied

C14 

br-AES 3EO

C4 branched # 2 carbon

ALFOTERRA® 4PO

C4 branched # 2 carbon

SO4

PEG

SO4 SO4

PEG PPG

br-C14 br-C14

AES 3EO

C14 linear alcohol

Figure 23: Alcohol Ether Sulfate Structure: AES and ALFO 23-4S

Figure 23 shows a 3-D model for anionic surfactant molecules of a linear and branched 

ethoxylated sulfate (AES) and a branched alcohol propoxy sulfate (ALFOTERRA® Surfactant); 

each propoxylated with 4 moles of PO. Note the reduced length and multiple appendages of 

the ALFOTERRA® molecule. The structure of the molecule when a branched alcohol chain and 

multiple propoxylate groups are present could act as a barrier to bacterial attack. Additionally, 

note how the PPG branch and resulting sulfate position protect the fourth sulfate oxygen (not 

seen directly in this view) from direct exposure. In fact, almost all oxygen molecules in the 

ALFOTERRA molecule are protected by the bulky alcohol and/or propylene oxide carbon chain. 

The experimental results confirm that the rate of biodegradation for ALFO 23-4S is impacted by 

the reduced linearity of the molecule’s structure from branching and PPG.

Surfactant molecules are highly complex and different degradation pathways are possible for 

any given surfactant (Sharvelle, 2007). Complete removal may be difficult due to formation of 

by-products that are more slowly degraded compared to the parent compound. The recognized 

and described metabolic pathways for biodegradation of alcohol ethoxylates and alcohol ethoxy

sulfates (AES) surfactant molecules is based on specific points of bacterial attack. 

Three routes have been identified whereby bacteria can initiate biodegradation of linear alcohol 

ethoxylates (LAE) (the un-sulfated form of AES). (White, 1993) The routes are (Figure 24): 

 w/β-oxidation of the carbon chain of the alcohol hydrophobe in the intact surfactant

 exo-cleavage of the ethylene glycol C2 units from the PEG moiety and/or

 cleavage at the ether bond which joins the PEG chain to the hydrophobic alkyl group 

(central scission). 

The attack routes for AES are:

 omega oxidation on the carbon chain of the alcohol hydrophobe

 central fission separating the alcohol from the ethylene glycol (EG) chain

 ether cleavage along the EG chain and 

 sulfate hydrolysis. 
12/14/2011 Page 37 of 66

In previously published work Hales and White discuss the degradation of an alcohol ethoxy

sulfate - linear dodecyl 3 mole ethoxy sulfate (similar to linear dodecyl 4 mole ethoxy sulfate 

used in this study). In their work, all four AES routes were identified in studies employing four 

bacteria species (Hales, 1986; White and Russell, 1998). In fact, they were able to identify 

mono-, di- and triethyleneglycol sulfates as the metabolites of the ether cleavage of the three 

ether bonds in the dodecyl triethoxysulfate.

In 1982, Yoshimura (Yoshimura, 1982) showed the impact of branching in the alcohol 

hydrophobe portion of AES. Yoshimura’s results showed that AES with a linear hydrophobe is 

ready biodegradable, whereas a branched hydrophobe is much more resistant. The 

degradation difficulty of the branched AES surfactant was attributed to steric hindrance 

preventing enzymic attack. 

In this same study, Yoshimura also shows the impact of alkyoxylate structure on biodegradation 

by comparing linear alcohol ethoxy sulfate (AES) linear alcohol propoxyl sulfate (APS). The 

results showed that the APS surfactant is more resistant to biodegradation than the AES. It was 

suggested that this is likely due to the inclusion of a secondary carbon atom in the structure of 

the propylene oxide group which shields the oxygen molecule from attack.

In general, it can be assumed that the AES is biodegraded through either of the following 

pathways:

(a) Enzymic hydrolysis of the sulfate group yielding sulfate and alcohol ethoxylate. The latter is 

further degraded through the biodegradation pathway of a primary alcohol ethoxylate by 

way of central scission. Or

(b) Enzymes attack the terminal methyl group of the linear alcohol reducing the carbon chain 

length as it moves forward. This is followed by β-oxidation.

If both the AES and ALFO 23-4S used in this study follow route (a) then metabolites yielded can 

be alcohol, PEG and/or PPG chains respectively. It would then be expected that the AES and 

ALFO 23-4S degradation would be more similar or that the ALFO 23-4S degradation would be 

more similar to the alcohol propoxy sulfate from which it was derived. This was not the case.

In an extensive study comparing the biodegradation of low molecular weight polyethylene glycol 

(PEG 300) with polypropylene glycol (PPG 425) Zgola (Zgola-Grześkowiak, 2005) showed that 

the biodegradation pathways of PEG and PPG are very different. PEG is biodegraded through 

clipping of the PEG chain resulting in metabolites that were identified as shortened PEG

ethoxymers. That is, the molecular weight of all PEG molecules is reduced consistently over 

time throughout the degradation process. In contract, the biodegradation of PPG does not 

involve chain scission at all. In fact, Zgola showed that while the homolog distribution pattern of 

PPG propoxymers remains unchanged (average molecular weight do not decrease) during the 

biodegradation test, the total quantity of molecules reduces over time.; i.e. no metabolites that 

are shortened version of the parent molecule are formed. In other words, the concentrations of 

the individual homologs of PPG decrease instead of shifting to shorter or lower molecular weight 

homologs. The result is that the bacteria appear to be attacking the whole PPG molecule at one

time. Zgola does not address how this mechanism is impacted with the addition of a sulfate 

head group.

Finally, in a 2002 paper, Ellis identified enzymes that work to degrade branched alcohol sulfates 

but not linear alcohol sulfates, and vice verse. In this paper a novel alkylsulfatase enzyme was

isolated that is specific to a branched primary alkyl sulfate surfactant 2-butyloctyl sulfate (Ellis, 

2002). This identified novel sulfatase enzyme is produced from a specific strain of 
12/14/2011 Page 38 of 66

Pseudomonas AE-D bacteria grown on 2-butyloctyl sulfate; this enzyme is not active on the 

linear alcohol sulfate dodecyl sulfate (SDS). In contrast, Pseudomonas sp. Strain C12B, well 

known for its ability to degrade linear SDS, was unable to grow on 2-butyloctyl sulfate. 

Additionally, its alkylsulfatases which is responsible for initiating the degradation of SDS by 

releasing the parent alcohol, exhibited no hydrolytic activity towards 2-butyloctyl sulfate.

Clearly small structural differences in otherwise similar molecules can drastically impact the 

biodegradation rate, pathway and active bacterial species. 

ω-/βOxidation

Central 

Scission

Ether 

Cleavage Sulfate 

Hydrolysis

AES

APS

No Cleavage

Restricted 

Sulfate 

Hydrolysis

ω-/βOxidation

Central 

Scission

Ether 

Cleavage Sulfate 

Hydrolysis

AES

APS

No Cleavage

Restricted 

Sulfate 

Hydrolysis

Figure 24: Anionic Linear-AES and branched APS (ALFO 23-4S) Molecule Biodegradation 

Pathway - Points of Attack

Figure 24 contains the 3-D models for AES and ALFO 23-4S anionic surfactant molecules. The 

arrows compare points of attack as discussed in the literature references above. For AES, all 

points of attack can and do occur rapidly; however for ALFO 23-4S, the central scission of the 

alcohol and PPG chain, steric hindrance preventing ether cleavage, and restricted sulfate 

hydrolysis all compromise the rate at which the molecule degrades. 

Yet, in spite of the degradation barriers, the work detailed in this report proves that primary 

biodegradation of alcohol propoxy sulfate surfactants, and specifically ALFO 23-4S, does occur. 

Further, and more importantly, primary degradation results in the elimination of the aquatic 

toxicity that is associated with the anionic surfactant parent. With responsible handling, and 

treatment minimally through domestic or industrial waste water treatment facilities of any rinse 

waters containing the alcohol propoxy sulfates, Sasol’s ALFOTERRA®surfactants should be 

harmless to the environment.
12/14/2011 Page 39 of 66

REFERENCES

1. OECD.1993.OECD Guidelines for the Testing of Chemicals – OECD 301A: DOC DieAway and 301F: Manometric Respirometry Test. Organization of Economic Cooperation 

and Development. Paris.

2. Sharvelle, S., Lattyak, R., Banks, M. K. (2007). Evaluation of biodegradability and 

biodegradation kinetics for anionic, nonionic, and amphoteric surfactants. Water Air Soil 

Pollut. 183, 177-186. 

3. White G. F., (1993) Bacterial biodegradation of ethoxylated surfactants. Pestic Sci 37, 

159-166.

4. Hales, S. G., Watson, G. K., Dodgson, K. S., 

5. White G. F. (1986). A comparative study of the biodegradation of the surfactants sodium 

dodecyltriethoxy sulfate by four detergent-degrading bacteria. J. Gen. Microbiol 132, 

953-961.

6. White, G. F., and Russell, N. J. (1988). Mechanisms of bacterial biodegradation of alkyl 

sulfate and alkylpolyethoxy sulfate surfactant. Biodeterioration 7, 325-332.

7. Yoshimura, K., Masuda F., (1982). Biodegradation of sodium alkyl poly(oxyalkylene) 

sulfates. JOACS 59, 328-332.

8. Zgola-Grześkowiak, A., Grzeskowiak, T., Zembrzuska, J., Lukaszewski, Z., (2005). 

Comparison of biodegradation of poly(ethylene glycol)s and poly(propylene glycol)s. 

Chemosphere 64, 803-809.

9. Ellis, A. J., Hales, S. G., Ur-Rehman, Naheed G. A., White, G. F., (2002). Novel 

alkylsulfatases required for biodegradation of the branched primary alkyl sulfate 

surfactant 2-butyloctyl sulfate. Appli. Environ. Microbiol. 68, 1, 31-36.

10. US EPA OPPTS 835.3220 “Porous Pot Test” Fate, Transport and Transformation Test 

Guidelines (April 1992).
12/14/2011 Page 40 of 66

APPENDICES 

Appendix 1: Stepan AES...........................................................................................................41

Appendix 2: Figure 2 - HRT......................................................................................................42

Appendix 3: Figure 3 - MLTSS.................................................................................................45

Appendix 4: Figure 4 - Porous Pot Effluent TSS.......................................................................48

Appendix 5: Figure 5 - Porous Pot Effluent pH Values .............................................................49

Appendix 6: Figure 6 - Porous Pot DOC Feed and Effluents....................................................51

Appendix 7: Figure 7 - PPS Effluent DOC Un-acclimated ALFO 23-4S.....................................54

Appendix 8: Figure 8 - PPS for Fragment Analysis Effluent......................................................58

Appendix 9: Figure 9 - Return to Steady State: Post Un-acclimated ALFO 23-4S....................59

Appendix 10: Figure 10 - Porous Pot DOC: Comparison of 1214E4S and ALFO 23-4S...........60

Appendix 11: Figure 11 - PPS: Increased HRT ........................................................................61

Appendix 12: Figure 12 - Die-Away Porous Pot Effluent ..........................................................61

Appendix 13: Figure 13 - OECD Die-Away DOC Loss % .........................................................61

Appendix 14: Figure 14 - Porous Pot: MBAS Primary Biodegradation......................................62

Appendix 15: ALFO 23-4S Comparison Porous Pot and Die Away ..........................................62

Appendix 16: Figure 16 - AES and ALFO 23-4S Acute Toxicity ...............................................62

Appendix 17: Figure 17 - Porous Pot Aquatic Toxicity 96 Hour Minnow ...................................62

Appendix 18: Figure 18- OECD Die-Away Test ALFO 23-4S ...................................................62

Appendix 19: Figure 19: ALFOTERRA Hydrophobe Biodegradation Study - OECD Demand .63

Appendix 20: Figure 20 - ALFOTERRA Hydrophobe 4PO Biodegradation Study - OECD .......64

Appendix 21: Figure 21- ALFOTERRA Hydrophobe 4PO Sulfate Biodegradation Study - OECD

.................................................................................................................................................65

Appendix 22: ALFOTERRA Hydrophobe Biodegradation Study, Comparison of Alcohol, 

Nonionic, and Anionic ...............................................................................................................66

Appendix 23: Aquatic Toxicity Reports from Environmental Enterprise USA Laboratory TSR No. 

R&D ENV 10-004 01-10............................................................................................................66
12/14/2011 Page 41 of 66

Appendix 1: Stepan AES

POLYSTEP® B-430S

Actives, % 28

Cloud Point 5oC (41oF)

Viscosity 53 cps @ 25oC (77oF)

Pour Point -2oC (28oF)

FDA Clearances 21 CFR 176.170, 176.180

Chemical Description SODIUM LAURYL ETHER SULFATE, 4 EO

Form at 25°C Liquid
12/14/2011 Page 42 of 66

Appendix 2: Figure 2 - HRT

1 2 3 4 5 6

6-Apr 5.98 6.32 6.08 5.92 6.16 6.19

7-Apr 5.89 5.93 5.91 5.98 5.88 5.96

8-Apr 5.90 5.95 5.96 5.94 5.93 5.94

9-Apr 5.97 6.04 6.02 5.97 6.03 5.94

10-Apr 5.94 6.00 6.00 5.94 5.95 6.01

11-Apr 5.91 5.94 5.92 5.85 6.05 5.93

12-Apr 5.83 6.00 5.98 5.92 5.92 5.90

13-Apr 5.90 6.02 6.00 5.98 5.92 6.05

14-Apr 5.91 5.98 5.96 5.95 5.94 5.99

15-Apr 6.22 5.96 5.95 5.90 5.93 5.98

16-Apr 5.90 6.01 5.97 5.90 5.96 5.97

17-Apr 5.90 5.96 6.14 6.07 6.13 6.14

18-Apr 5.90 5.98 6.09 5.92 6.00 5.99

19-Apr 5.97 5.98 5.90 5.92 6.01 6.09

20-Apr 5.93 6.00 6.01 6.01 6.07 6.03

21-Apr 5.95 6.02 6.04 5.90 6.03 6.05

22-Apr 5.95 6.03 6.02 6.15 6.06 6.06

23-Apr 5.99 6.05 6.00 5.93 6.14 6.04

24-Apr 6.01 5.99 6.01 5.89 5.80 6.07

25-Apr 6.01 5.99 5.98 5.95 5.99 5.96

26-Apr 6.00 5.95 5.96 5.96 6.10 6.04

27-Apr 5.98 5.95 6.00 5.94 6.06 6.11

28-Apr 5.89 6.03 6.07 5.98 5.99 6.02

29-Apr 5.83 5.99 6.02 5.95 5.92 6.07

30-Apr 5.88 5.97 6.11 6.00 5.92 6.11

1-May 5.94 5.97 5.94 5.68 5.95 5.83

2-May 5.90 6.00 5.99 5.89 5.93 6.01

3-May 5.91 5.88 5.91 5.84 5.89 6.00

4-May 5.87 5.94 5.94 5.91 5.96 6.05

5-May 5.91 5.94 5.94 5.92 6.01 6.01

6-May 5.90 5.95 5.95 5.90 5.99 5.99

7-May 5.91 5.96 5.94 5.93 5.95 6.00

8-May 5.91 5.93 5.92 5.93 5.94 5.95

9-May 5.91 6.00 5.98 5.89 5.92 5.98

10-May 5.90 6.03 5.94 5.88 5.93 6.03

11-May 5.91 6.04 6.00 5.92 5.95 5.98

12-May 5.94 5.99 5.97 5.94 5.94 6.04

13-May 5.91 6.06 5.98 6.10 5.87 6.11

14-May 5.90 6.00 5.93 5.87 5.94 5.96

15-May 5.90 6.00 5.93 5.87 5.94 5.96

16-May 5.86 5.89 5.86 5.77 5.81 6.46

17-May 5.91 5.97 5.94 5.88 5.96 5.98

18-May 5.90 5.97 5.94 5.91 5.93 5.95

19-May 5.91 6.02 6.03 5.90 5.97 5.98

20-May 5.90 6.01 5.96 5.86 5.96 6.02

21-May 5.93 6.04 5.94 5.86 5.88 5.93

22-May 5.95 6.03 6.02 6.15 6.06 6.06

23-May 6.07 6.14 6.06 6.00 6.06 5.96

24-May 6.31 6.15 5.95 5.89 5.96 5.88

25-May 6.01 5.90 5.97 5.90 5.97 5.86

26-May 5.89 5.93 5.99 5.89 5.94 5.96

27-May 5.87 5.93 5.97 5.86 5.95 5.84

28-May 5.90 5.91 5.85 5.85 5.85 5.79

29-May 5.63 6.04 5.97 5.99 6.12 5.99

30-May 5.63 6.04 5.97 5.99 6.12 5.99

31-May 5.81 5.90 5.84 5.86 5.94 5.94

1-Jun 5.98 5.89 5.89 5.86 5.96 5.88

2-Jun 6.01 5.92 5.92 5.89 5.99 5.91

3-Jun 5.99 5.93 5.87 5.90 5.94 5.91

4-Jun 5.97 5.71 5.95 5.96 6.06 5.96

Figure 2. Hydraulic Retention Time (HRT) Through Porous Pots (hrs)
12/14/2011 Page 43 of 66

1 2 3 4 5 6

5-Jun 6.05 6.02 5.96 5.88 5.95 5.89

6-Jun 6.57 6.03 5.92 5.94 5.98 5.96

7-Jun 5.75 6.06 5.93 5.94 5.94 6.01

8-Jun 5.69 6.06 6.07 6.18 5.98 5.96

9-Jun 5.98 6.48 6.12 6.10 6.18 6.10

10-Jun 5.76 5.78 5.90 5.91 5.94 5.88

11-Jun 5.83 6.03 5.92 5.94 5.95 5.92

12-Jun 5.79 5.89 5.86 5.88 5.95 5.94

13-Jun 5.87 5.93 5.87 5.88 6.08 5.95

14-Jun 5.96 6.04 5.89 5.99 6.02 6.07

15-Jun 5.92 5.98 5.99 5.90 5.99 6.04

16-Jun 5.79 5.92 5.87 5.94 5.92 5.91

17-Jun 5.89 5.95 5.89 5.94 5.98 6.03

18-Jun 5.84 5.93 5.88 5.97 5.96 6.06

19-Jun 5.85 5.94 5.89 5.96 5.78 5.87

20-Jun 5.87 5.96 5.88 6.02 6.06 5.89

21-Jun 5.85 5.97 5.89 6.08 5.87 5.94

22-Jun 5.99 6.06 6.04 5.85 6.02 6.01

23-Jun 6.01 6.06 6.04 5.90 5.94 6.02

24-Jun 5.97 6.11 6.03 5.89 6.03 6.02

25-Jun 5.84 5.97 5.91 5.77 5.93 5.97

26-Jun 6.15 5.89 5.85 5.83 5.96 5.99

27-Jun 6.08 5.82 5.84 5.86 5.91 5.88

28-Jun 5.86 5.72 5.89 5.87 6.10 5.94

29-Jun 6.00 6.00 5.90 6.08 5.72 5.93

30-Jun 5.83 6.13 5.90 6.04 5.76 5.96

1-Jul 5.29 4.99 5.12 5.26 4.99 5.14

2-Jul 5.14 5.20 5.21 5.74 5.08 5.23

3-Jul 5.14 5.97 5.40 5.05 5.05 5.26

4-Jul 5.20 5.15 5.42 5.30 5.21 5.69

5-Jul 4.88 4.83 5.06 4.80 4.89 5.02

6-Jul 4.94 4.90 4.85 5.09 4.90 5.09

7-Jul 4.92 4.88 4.83 4.84 4.89 4.91

8-Jul 5.01 4.89 4.86 4.88 4.95 4.93

9-Jul 5.10 4.89 4.82 4.80 4.91 4.94

10-Jul 4.97 4.89 4.86 4.88 4.93 4.89

11-Jul 4.90 4.85 4.77 4.76 4.82 4.73

12-Jul 5.01 8.29 12.59 4.93 5.20 4.96

13-Jul 5.00 4.92 4.85 4.95 4.89 4.79

14-Jul 4.97 4.96 4.84 4.89 4.92 4.79

15-Jul 5.07 5.06 4.99 17.30 5.11 5.06

16-Jul 5.13 5.11 5.26 5.01 5.16 5.05

17-Jul 5.14 5.07 5.25 5.44 5.10 5.27

18-Jul 5.14 4.97 5.22 5.28 5.22 5.27

19-Jul 5.13 5.09 5.20 5.27 5.14 5.24

20-Jul 5.12 5.07 5.24 5.41 7.07 5.25

21-Jul 5.11 5.01 5.23 5.01 5.11 5.23

22-Jul 5.56 5.43 5.75 5.49 5.62 5.88

23-Jul 5.90 5.93 5.91 5.94 6.17 5.95

24-Jul 5.86 5.89 5.92 5.93 5.96 5.88

25-Jul 5.96 6.00 6.04 6.04 6.29 6.00

26-Jul 6.15 6.29 6.32 6.24 6.55 6.31

27-Jul 5.99 5.99 5.99 5.98 5.95 5.97

28-Jul 5.92 5.95 6.11 6.08 5.91 6.16

29-Jul 5.92 5.99 6.01 6.03 5.94 5.97

30-Jul 6.09 6.09 6.03 6.09 6.01 6.12

31-Jul 5.96 6.10 5.98 5.99 5.93 6.02

1-Aug 5.93 5.95 5.93 5.95 5.88 5.96

2-Aug 6.25 5.90 5.98 5.97 5.91 5.94

3-Aug 6.22 5.91 5.96 5.95 5.89 5.96

4-Aug 5.81 5.87 5.94 5.97 5.90 5.95

Figure 2. Hydraulic Retention Time (HRT) Through Porous Pots (hrs)
12/14/2011 Page 44 of 66

1 2 3 4 5 6

5-Aug 5.88 5.87 5.89 6.27 5.89 5.98

6-Aug 5.88 5.88 5.94 5.97 5.96 5.97

7-Aug 5.88 5.88 5.94 5.97 5.96 5.97

8-Aug 5.90 5.87 5.96 5.94 5.90 5.94

9-Aug 5.88 5.93 6.99 6.01 5.90 5.97

10-Aug 5.84 5.90 6.02 6.03 5.88 5.94

11-Aug 5.85 5.92 5.89 6.03 5.88 5.91

12-Aug 5.85 5.88 5.95 6.03 5.90 5.95

13-Aug 5.86 5.92 5.94 5.94 5.89 5.89

14-Aug 5.85 5.85 5.96 5.93 5.87 5.94

15-Aug 5.85 5.89 5.90 6.06 5.85 5.97

16-Aug 5.89 5.98 6.00 5.94 5.95 5.95

17-Aug 5.85 9.92 5.90 5.90 5.88 5.90

18-Aug 5.87 5.92 6.03 5.93 5.87 5.92

19-Aug 5.90 5.92 5.88 6.53 5.92 5.90

20-Aug 5.90 5.94 6.42 5.92 5.91 5.90

21-Aug 5.91 6.13 5.81 5.93 6.00 5.89

22-Aug 5.88 5.91 5.80 5.89 5.90 5.90

23-Aug 5.99 6.02 6.09 6.09 6.15 6.12

24-Aug 5.91 5.87 6.05 5.88 5.92 5.92

25-Aug 5.92 5.90 5.94 5.90 5.87 5.91

26-Aug 5.89 5.87 5.93 5.88 5.86 5.89

27-Aug 5.92 5.91 5.96 5.89 6.01 5.93

28-Aug 5.98 5.95 6.00 5.96 5.91 5.96

29-Aug 5.91 5.98 5.96 5.97 5.73 5.92

30-Aug 6.03 6.03 6.08 6.03 6.09 6.08

31-Aug 5.96 5.91 5.98 5.97 5.92 6.05

1-Sep 5.90 5.89 5.95 5.87 5.90 5.91

2-Sep 5.96 5.88 5.95 5.91 6.00 5.84

3-Sep 5.94 5.85 5.96 5.95 5.98 5.88

4-Sep 5.89 5.86 5.83 5.98 5.88 5.83

5-Sep 5.93 5.81 5.95 5.93 5.85 5.83

6-Sep 5.91 5.87 5.95 5.90 5.89 5.93

7-Sep 5.95 5.89 5.97 5.92 5.86 5.86

8-Sep 6.01 5.93 5.98 5.96 5.92 5.86

9-Sep 5.91 5.94 6.01 5.94 5.90 5.92

10-Sep 5.93 5.90 6.30 5.92 5.86 5.89

11-Sep 5.98 5.98 5.90 5.87 5.81 5.85

12-Sep 5.92 5.88 5.94 5.92 5.83 5.93

13-Sep 5.98 5.88 5.92 5.94 5.88 5.89

14-Sep 6.18 5.91 5.95 5.94 5.89 5.89

15-Sep 5.89 5.89 5.98 5.93 5.92 5.82

16-Sep 5.98 5.86 5.80 5.98 5.82 5.86

17-Sep 5.94 5.87 5.82 5.96 5.88 5.83

18-Sep 5.94 5.93 5.88 5.97 5.93 5.86

19-Sep 5.95 5.92 5.94 6.00 5.93 5.92

20-Sep 5.81 5.83 5.79 6.06 5.80 5.77

21-Sep 6.00 6.01 6.01 6.12 5.96 6.06

22-Sep 6.12 6.12 6.11 6.21 6.05 6.13

23-Sep 6.15 6.15 6.14 6.34 6.11 6.15

24-Sep 6.15 6.15 6.14 6.34 6.11 6.15

25-Sep 5.92 5.92 5.95 10.79 5.89 5.89

26-Sep 6.70 6.50 6.62 6.00 6.18 6.95

27-Sep 7.99 7.58 7.85 7.35 7.69 7.91

28-Sep 9.12 9.04 8.98 8.88 8.89 9.48

29-Sep 11.15 10.68 10.43 10.88 10.48 10.19

30-Sep 11.68 12.03 11.55 11.80 11.64 11.17

1-Oct 11.78 11.94 11.51 11.93 11.60 12.54

2-Oct 11.59 12.06 11.53 11.90 12.17 11.94

3-Oct

4-Oct 11.66 12.15 11.58 12.09 11.61 11.77

5-Oct 11.60 11.94 11.59 11.67 11.84 12.09

6-Oct 11.46 11.98 11.43 12.01 11.57 11.29

7-Oct 11.61 11.75 11.35 11.73 11.62 11.39

Figure 2. Hydraulic Retention Time (HRT) Through Porous Pots (hrs)
12/14/2011 Page 45 of 66

Appendix 3: Figure 3 - MLTSS

Pot 1 2 3 4 5 6

6-Apr 3,720 3,870 3,410 3,750 3,320 3,440

7-Apr 4,460 3,850 3,740 3,920 3,670 3,690

8-Apr 3,880 4,020 3,630 3,890 3,510 3,760

9-Apr 3,770 4,130 3,640 3,790 3,390 3,870

10-Apr 3,990 4,230 3,660 3,910 3,560 3,990

11-Apr 3,690 4,240 3,960 4,070 3,590 3,940

12-Apr 3,800 4,280 3,770 4,030 3,640 4,150

13-Apr 3,760 4,230 3,910 4,030 3,600 3,820

14-Apr 3,580 4,280 3,710 3,870 3,630 3,660

15-Apr 3,670 4,360 3,670 3,930 3,470 3,550

16-Apr 3,620 4,540 3,880 4,210 3,750 3,750

17-Apr 3,690 4,580 4,060 4,220 3,850 4,030

18-Apr 3,700 4,140 3,510 3,600 3,830 3,620

19-Apr 3,720 3,980 3,700 3,420 3,630 4,100

20-Apr 3,680 3,790 3,730 3,170 3,460 3,860

21-Apr 3,460 3,430 3,700 3,360 3,670 3,730

22-Apr 3,400 3,520 3,510 3,260 3,350 3,500

23-Apr 3,550 3,680 3,290 3,300 3,440 3,680

24-Apr 3,530 3,620 3,250 3,220 3,260 3,420

25-Apr 3,400 4,040 3,440 3,360 3,230 3,900

26-Apr 3,370 3,950 3,460 3,190 2,890 3,510

27-Apr 3,280 3,960 3,390 3,180 2,750 3,180

28-Apr 3,380 3,890 3,380 3,120 2,750 3,150

29-Apr 3,410 3,890 3,320 2,990 2,710 2,650

30-Apr 3,530 4,020 3,310 2,820 2,720 2,680

1-May 3,000 3,290 2,520 2,890 2,230 2,240

2-May 3,130 3,480 2,930 3,060 2,850 2,390

3-May 3,820 4,030 3,650 3,650 3,040 2,840

4-May 4,280 4,080 3,680 3,810 2,980 3,150

5-May 4,110 4,070 3,860 3,830 2,880 3,430

6-May 4,270 4,200 4,030 3,980 2,720 3,690

7-May 3,700 3,560 3,210 3,320 2,490 3,290

8-May 3,790 3,960 3,330 3,380 2,690 3,420

9-May 4,480 4,090 3,730 3,940 2,970 3,970

10-May 4,180 4,190 3,610 3,950 3,300 4,070

11-May 4,180 4,520 3,760 3,990 3,220 4,130

12-May 4,050 4,460 3,780 3,830 3,190 4,240

13-May 4,140 3,800 4,000 4,230 4,000 3,970

14-May 4,290 3,980 4,220 4,330 4,160 4,140

15-May 4,280 4,240 4,020 4,500 4,230 4,020

16-May 4,490 4,418 4,100 4,720 4,550 4,130

17-May 4,610 4,690 4,250 4,790 4,650 4,130

18-May 4,580 4,540 4,180 4,620 4,590 3,960

19-May 4,250 4,270 4,030 4,460 4,480 3,800

20-May 4,190 4,270 4,020 4,730 4,570 4,050

21-May 4,430 4,270 4,160 4,650 4,540 3,970

22-May 4,420 4,310 4,080 4,630 4,670 3,780

23-May 4,530 4,180 4,170 4,750 4,710 3,920

24-May 4,570 4,380 4,140 4,980 4,950 4,030

25-May 4,300 4,370 3,680 4,890 4,720 3,870

26-May 4,200 4,250 3,740 4,920 4,720 3,950

27-May 3,840 3,920 3,660 4,540 4,330 3,760

28-May 3,230 3,980 3,560 4,350 4,210 4,990

29-May 3,610 3,390 3,370 3,950 3,930 3,430

30-May 3,540 3,210 3,370 3,810 3,790 3,460

31-May 3,610 3,510 3,300 3,920 3,770 3,480

1-Jun 3,300 3,270 2,930 3,800 3,430 3,070

2-Jun 3,530 3,450 3,100 3,930 3,550 3,010

Figure 3. Mixed Liquor Total Suspended Solids (MLTSS)
12/14/2011 Page 46 of 66

Pot 1 2 3 4 5 6

3-Jun 3,850 3,390 3,210 4,240 3,810 3,260

4-Jun 4,140 3,910 3,240 4,470 4,050 3,520

5-Jun 4,220 4,220 3,610 4,670 4,460 3,930

6-Jun 4,320 4,350 3,920 4,500 4,220 3,700

7-Jun 4,420 4,390 4,330 4,550 4,380 3,060

8-Jun 4,370 4,430 4,240 4,300 4,430 3,280

9-Jun 4,760 4,270 4,490 4,370 4,580 3,800

10-Jun 4,880 4,970 4,750 4,360 4,740 4,140

11-Jun 4,870 4,950 4,710 4,410 5,030 4,370

12-Jun 4,890 5,190 4,890 4,640 4,990 5,330

13-Jun 4,620 4,880 5,010 4,390 4,880 4,970

14-Jun 4,390 4,720 4,950 4,200 4,670 4,650

15-Jun 3,900 4,320 4,950 4,160 4,640 4,500

16-Jun 4,290 4,270 4,870 4,000 4,380 4,230

17-Jun 4,180 4,140 4,850 4,010 4,260 4,330

18-Jun 3,830 3,700 4,470 3,780 3,830 4,160

19-Jun 3,630 3,580 4,100 3,730 3,770 4,050

20-Jun 3,580 4,130 4,130 3,390 3,770 3,900

21-Jun 3,570 3,000 4,100 3,270 3,890 3,890

22-Jun 3,040 2,770 3,960 2,960 2,900 3,680

23-Jun 2,700 2,550 3,560 2,790 4,350 3,660

24-Jun 2,640 2,520 3,230 2,270 3,630 3,800

25-Jun 2,370 2,480 3,310 2,400 3,810 3,270

26-Jun 2,500 1,668 3,350 2,410 3,660 3,510

27-Jun 2,400 2,410 3,210 2,620 3,790 3,390

28-Jun 2,440 2,940 3,480 3,030 3,660 3,530

29-Jun 2,540 3,360 3,270 3,290 3,360 3,150

30-Jun 3,190 3,610 3,520 3,490 3,020 3,440

1-Jul 3,540 4,340 4,010 3,490 3,200 3,630

2-Jul 3,780 4,670 3,480 3,490 2,990 5,780

3-Jul 4,320 4,630 3,240 3,680 2,920 3,890

4-Jul 6,370 5,050 4,060 4,880 3,360 4,160

5-Jul 5,560 5,450 4,770 5,100 4,640 4,770

6-Jul 5,030 5,300 5,020 5,060 3,870 3,880

7-Jul 3,510 5,240 6,320 4,840 3,750 4,400

8-Jul 4,710 4,870 5,060 4,340 4,120 5,050

9-Jul 5,200 4,930 4,760 5,350 4,680 5,160

10-Jul 5,240 4,960 4,790 4,620 5,170 5,340

11-Jul 6,120 6,250 5,380 7,020 5,660 5,850

12-Jul 5,650 4,840 4,280 6,020 5,840 5,470

13-Jul 5,800 4,370 4,800 6,630 6,110 5,690

14-Jul 5,040 5,720 5,520 5,400 6,800 5,750

15-Jul 6,660 6,790 6,300 4,410 6,420 6,310

16-Jul 6,380 6,820 6,170 5,280 6,780 6,010

17-Jul 5,970 6,400 6,080 6,030 6,750 6,000

18-Jul 5,510 5,730 5,740 5,520 5,830 5,530

19-Jul 5,330 5,590 5,240 5,310 5,530 5,090

20-Jul 5,310 5,560 5,010 5,150 5,100 4,950

21-Jul 5,160 4,380 4,960 5,390 5,000 4,780

22-Jul 5,320 5,240 4,800 5,070 4,890 4,620

23-Jul 4,680 4,610 4,350 3,640 4,510 4,470

24-Jul 4,610 4,700 4,520 4,290 4,700 4,280

25-Jul 3,460 4,260 4,760 4,250 4,060 4,060

26-Jul 3,460 4,260 4,760 4,250 4,060 3,960

27-Jul 4,670 4,170 4,510 4,920 4,650 4,270

28-Jul 4,600 4,740 4,810 4,860 4,840 4,400

29-Jul 4,530 4,440 4,400 4,540 4,700 4,140

30-Jul 4,210 4,220 4,120 4,200 4,270 3,840

31-Jul 4,260 3,880 3,890 4,100 4,120 3,710

1-Aug 4,320 3,890 3,980 4,210 4,370 3,880

2-Aug 4,380 4,200 4,180 4,100 4,230 3,930

3-Aug 4,230 4,250 4,150 4,060 4,220 3,990

4-Aug 4,680 5,020 4,820 4,790 4,910 4,620

5-Aug 4,430 4,630 4,630 4,280 4,640 4,290

Figure 3. Mixed Liquor Total Suspended Solids (MLTSS)
12/14/2011 Page 47 of 66

Pot 1 2 3 4 5 6

5-Aug 4,430 4,630 4,630 4,280 4,640 4,290

6-Aug 4,300 4,740 4,460 4,350 4,360 4,580

7-Aug 4,120 4,480 4,230 4,170 4,140 4,350

8-Aug 4,060 4,340 3,230 4,010 3,970 4,240

9-Aug 3,620 3,660 3,410 3,530 3,620 3,730

10-Aug 4,310 4,140 3,710 4,090 3,900 4,270

11-Aug 4,100 3,770 3,560 3,760 3,500 3,950

12-Aug 3,720 4,040 3,520 3,890 3,470 3,730

13-Aug 4,230 4,240 3,710 4,110 3,930 4,040

14-Aug 4,360 4,350 3,960 4,120 4,090 4,020

15-Aug 4,790 4,960 4,580 6,040 4,800 3,360

16-Aug 4,590 4,160 4,780 4,650 4,650 4,750

17-Aug 4,530 4,150 4,600 4,760 4,680 4,040

18-Aug 4,050 4,350 4,340 4,210 4,010 4,270

19-Aug 3,860 4,090 4,210 3,970 3,780 3,800

20-Aug 3,750 3,890 4,100 3,860 3,690 3,580

21-Aug 3,700 3,550 3,960 3,680 3,410 3,080

22-Aug 3,820 3,300 3,920 3,560 3,420 3,100

23-Aug 4,090 3,290 3,560 3,710 3,640 3,510

24-Aug 4,120 3,070 3,280 3,310 3,710 3,340

25-Aug 5,230 4,420 4,590 4,580 5,350 4,550

26-Aug 4,780 4,340 4,500 4,580 4,910 4,080

27-Aug 4,870 4,400 4,470 4,590 5,090 4,140

28-Aug 4,460 3,090 5,300 4,120 4,650 3,860

29-Aug 4,240 3,940 4,250 4,140 5,120 4,170

30-Aug 4,090 3,510 4,130 4,010 4,240 3,880

31-Aug 3,770 3,690 4,120 3,920 4,420 4,040

1-Sep 3,770 3,420 3,890 4,060 4,590 3,630

2-Sep 3,900 3,230 3,970 4,220 4,230 3,470

3-Sep 4,000 4,710 3,970 4,280 4,200 3,580

4-Sep 3,760 3,250 3,770 3,910 4,220 3,220

5-Sep 4,830 4,670 4,960 5,240 4,980 4,380

6-Sep 4,940 4,700 5,130 4,980 4,840 4,560

7-Sep 4,630 4,440 4,950 4,880 5,120 4,370

8-Sep 4,400 4,500 4,830 5,030 5,330 4,530

9-Sep 4,360 4,780 4,840 5,090 5,160 3,400

10-Sep 4,300 4,740 4,450 4,860 4,760 4,680

11-Sep 4,000 4,560 4,130 4,670 4,590 4,200

12-Sep 5,770 6,210 5,990 6,650 6,050 5,860

13-Sep 4,610 5,300 5,230 5,260 4,810 4,900

14-Sep 4,680 5,210 5,320 5,390 5,160 5,550

15-Sep 4,840 5,270 5,260 5,330 5,110 5,070

16-Sep 4,460 5,100 5,060 5,050 4,670 4,650

17-Sep 4,120 4,650 4,690 4,400 4,170 4,670

18-Sep 4,440 4,730 4,780 4,850 4,440 4,900

19-Sep 5,510 5,780 6,050 5,910 5,640 6,060

20-Sep 5,420 5,320 5,720 5,770 4,900 5,830

21-Sep 4,910 5,160 5,520 5,280 5,180 6,470

22-Sep 5,880 5,540 5,920 5,280 5,190 5,850

23-Sep 4,610 4,620 4,740 4,350 4,430 5,150

24-Sep 4,600 4,460 4,130 4,120 4,060 4,530

25-Sep 4,560 4,300 4,320 3,410 4,640 4,470

26-Sep 5,760 5,850 5,430 4,950 5,560 5,850

27-Sep 5,250 5,550 5,200 4,260 5,150 5,180

28-Sep 4,600 4,850 4,460 3,930 4,780 4,580

29-Sep 4,000 4,130 3,830 3,110 3,880 3,740

30-Sep 3,800 3,880 3,860 3,030 3,850 3,740

1-Oct 3,550 3,960 3,790 3,490 4,070 3,910

2-Oct 3,600 3,930 3,810 3,120 3,800 3,640

3-Oct

4-Oct 4,290 4,700 4,540 3,760 4,660 4,070

5-Oct 3,730 4,480 4,590 3,780 4,530 3,990

6-Oct 3,550 4,640 4,350 2,810 5,330 3,630

7-Oct 3,650 4,190 4,210 3,810 4,070 3,520

Figure 3. Mixed Liquor Total Suspended Solids (MLTSS)
12/14/2011 Page 48 of 66

Appendix 4: Figure 4 - Porous Pot Effluent TSS

POT 1 2 3 4 5 6 POT 1 2 3 4 5 6

2-Jun 24 22 32 18 18 38 26-Jul 1 0 -2 1 -1 -2

3-Jun 13 20 47 18 11 41 27-Jul 5 3 2 5 2 2

4-Jun 11 7 44 7 0 28 28-Jul 1 -3 -1 0 -2 -3

5-Jun 7 13 33 13 6 21 29-Jul 6 4 6 7 3 3

6-Jun 14 17 11 14 13 7 30-Jul 1 1 1 3 2 2

7-Jun 10 10 14 14 12 21 31-Jul 5 3 3 5 5 1

8-Jun 5 2 9 11 4 12 1-Aug -2 2 -4 0 2 0

9-Jun 9 2 10 18 9 12 2-Aug 0 4 0 3 2 1

10-Jun 6 7 7 19 11 9 3-Aug 2 6 2 2 5 2

11-Jun 9 5 5 11 0 9 4-Aug 0 4 3 3 3 -1

12-Jun 9 9 11 12 1 14 5-Aug -2 -10 -3 -1 -2 -6

13-Jun 8 13 9 17 3 15 6-Aug 4 6 5 4 5 0

14-Jun 13 22 17 33 9 14 7-Aug 1 3 1 3 3 -2

15-Jun 2 25 9 16 9 11 8-Aug -9 5 7 6 0 1

16-Jun 11 28 7 12 8 5 9-Aug 0 4 1 5 2 -1

17-Jun 19 31 6 14 16 6 10-Aug 2 3 2 7 2 0

18-Jun 7 31 5 30 18 7 11-Aug -1 0 -3 0 0 -5

19-Jun 21 30 7 17 25 9 12-Aug 3 1 2 2 2 1

20-Jun 29 36 14 32 27 18 13-Aug -2 -3 0 -3 -3 -2

21-Jun 39 42 26 46 38 25 14-Aug -1 -2 -5 1 -3 -3

22-Jun 34 14 11 19 19 9 15-Aug 2 1 3 4 2 5

23-Jun 37 20 13 19 12 9 16-Aug 4 5 9 4 4 2

24-Jun 48 23 22 34 19 15 17-Aug 5 -1 7 6 3 2

25-Jun 40 19 23 30 19 16 18-Aug 0 -2 5 2 1 0

26-Jun 33 25 24 36 38 18 19-Aug 1 -2 3 3 0 1

27-Jun 29 13 13 21 29 11 20-Aug 7 4 9 7 8 7

28-Jun 22 24 13 15 18 18 21-Aug -1 -3 0 0 -4 1

29-Jun 26 14 17 23 21 29 22-Aug 8 9 10 6 8 7

30-Jun 16 10 5 15 22 19 23-Aug

1-Jul 21 20 16 23 41 23 24-Aug 1 0 3 -1 -1 -1

2-Jul 16 10 13 12 34 16 25-Aug -2 -3 2 -2 -4 -2

3-Jul 16 13 13 11 31 24 26-Aug -1 -2 3 0 -4 -3

4-Jul 9 11 10 8 22 44 27-Aug -2 0 5 -3 -2 1

5-Jul 4 4 6 7 10 7 28-Aug -4 0 6 1 -3 0

6-Jul 1 1 0 0 10 0 29-Aug 7 4 15 0 3 4

7-Jul 0 -1 -2 3 3 5 30-Aug 1 4 12 8 2 2

8-Jul 4 4 1 3 6 1 31-Aug 2 1 4 2 -1 2

9-Jul 2 0 2 1 0 -4 1-Sep -1 0 9 1 1 3

10-Jul 0 1 3 2 3 2 2-Sep -3 -1 5 -2 -3 0

11-Jul 8 11 19 10 10 16 3-Sep 1 0 5 0 -2 -2

12-Jul -4 1 28 -2 -4 0 4-Sep -6 -5 -1 -1 -3 -13

13-Jul 10 11 18 12 8 12 5-Sep 3 0 5 4 0 0

14-Jul -2 -4 -2 -2 -4 -2 6-Sep -4 -8 -6 -4 -4 -2

15-Jul 5 2 2 9 1 1 7-Sep

16-Jul 2 2 5 4 3 4 8-Sep

17-Jul 2 1 4 2 3 4 9-Sep

18-Jul 1 1 4 -1 0 0 10-Sep 9 5 10 9 0 2

19-Jul 1 0 3 -1 -1 -2 11-Sep 2 -1 6 2 -2 -1

20-Jul 1 0 2 -2 0 -3 12-Sep -1 -4 6 -9 1 5

21-Jul 3 2 3 0 1 -1 13-Sep

22-Jul 1 -3 -3 -2 -2 -3 14-Sep -2 -3 2 -2 -4 -8

23-Jul 4 3 1 1 1 -3 15-Sep 6 2 7 4 0 1

24-Jul 2 -1 -6 1 -1 2 16-Sep

25-Jul 1 0 -2 1 -1 -2 17-Sep 2 -1 2 -1 -3 -6

26-Jul 1 0 -2 1 -1 -2 18-Sep -1 1 1 4 0 -2

Figure 4 - Porous Pot Effluent TSS 
12/14/2011 Page 49 of 66

Appendix 5: Figure 5 - Porous Pot Effluent pH Values

POT 1 2 3 4 5 6

2-Jun 7.40 7.30 7.00 7.50 7.30 7.10

3-Jun 7.40 7.30 7.00 7.50 7.30 7.10

4-Jun 7.75 7.43 7.44 7.73 7.50 7.58

5-Jun 7.48 7.49 7.37 7.65 7.55 7.71

6-Jun 7.71 7.45 7.33 7.61 7.50 7.68

7-Jun 7.74 7.66 7.58 7.60 7.46 7.75

8-Jun 7.76 7.71 7.60 7.56 7.67 7.70

9-Jun 7.68 7.71 7.34 7.50 7.58 7.59

10-Jun 7.59 7.69 7.38 7.44 7.34 7.57

11-Jun 7.53 7.67 7.54 7.48 7.39 7.62

12-Jun 7.60 7.69 7.35 7.47 7.39 7.64

13-Jun 7.59 7.64 7.45 7.40 7.38 7.60

14-Jun 7.56 7.54 7.43 7.42 7.29 7.59

15-Jun 7.65 7.65 7.26 7.56 7.32 7.66

16-Jun 7.70 7.70 7.30 7.53 7.52 7.68

17-Jun 7.68 7.64 7.31 7.52 7.53 7.63

18-Jun 7.58 7.59 7.29 7.51 7.52 7.62

19-Jun 7.52 7.55 7.27 7.50 7.40 7.60

20-Jun 7.53 7.62 7.29 7.48 7.25 7.61

21-Jun 7.48 7.60 7.27 7.47 7.21 7.60

22-Jun 7.49 7.55 7.29 7.47 7.48 7.61

23-Jun 7.56 7.55 7.41 7.55 7.37 7.64

24-Jun 7.64 7.53 7.48 7.60 7.42 7.68

25-Jun 7.56 7.41 7.44 7.55 7.38 7.58

26-Jun 7.57 7.32 7.43 7.52 7.47 7.46

27-Jun 7.59 7.29 7.45 7.39 7.45 7.44

28-Jun 7.55 7.30 7.48 7.53 7.37 7.48

29-Jun 7.43 7.22 7.31 7.46 7.16 7.25

30-Jun 7.45 7.22 7.33 7.60 7.27 7.53

1-Jul 7.28 7.48 7.45 7.31 7.21 7.67

2-Jul 7.43 7.66 7.62 7.48 7.35 7.66

3-Jul 7.39 7.56 7.54 7.44 7.20 7.67

4-Jul 7.40 7.71 7.57 7.50 7.27 7.78

5-Jul 7.39 7.45 7.07 7.50 7.30 7.02

6-Jul 7.35 7.08 6.68 7.35 7.37 6.64

7-Jul 7.25 7.05 6.60 7.29 7.29 6.70

8-Jul 7.42 7.23 6.56 7.55 7.32 6.85

9-Jul 7.21 7.25 6.60 7.49 7.26 6.85

10-Jul 7.19 7.12 6.62 7.41 7.21 6.71

11-Jul 7.25 7.42 6.47 7.50 7.24 6.60

12-Jul 7.27 7.35 6.18 7.31 7.22 6.46

13-Jul 7.27 6.85 6.12 7.31 7.20 6.42

14-Jul 7.32 7.23 6.01 7.26 7.23 6.13

15-Jul 7.43 7.55 6.66 7.56 7.35 6.69

16-Jul 7.54 7.70 7.05 7.21 7.38 7.12

17-Jul 7.60 7.91 7.11 7.37 7.36 7.12

18-Jul 7.48 7.80 6.57 7.24 7.15 6.58

19-Jul 7.39 7.77 6.30 7.10 7.09 6.43

20-Jul 7.52 7.88 7.08 7.34 7.24 7.09

21-Jul 7.50 7.88 7.26 7.54 7.31 7.38

22-Jul

23-Jul 7.52 7.85 7.29 7.57 7.49 7.46

24-Jul

25-Jul 7.62 7.70 7.51 7.39 7.28 7.59

26-Jul 7.54 7.59 7.55 7.42 7.32 7.56

27-Jul 7.54 7.60 7.63 7.46 7.41 7.60

28-Jul 7.54 7.62 7.63 7.46 7.47 7.55

29-Jul 7.58 7.57 7.64 7.43 7.40 7.54

Figure 5. Porous Pot Effluent pH Values
12/14/2011 Page 50 of 66

POT123456

30-Jul 7.61 7.59 7.66 7.46 7.45 7.53

31-Jul 7.59 7.55 7.58 7.46 7.41 7.44

1-Aug 7.55 7.52 7.63 7.51 7.42 7.49

2-Aug 7.50 7.43 7.59 7.47 7.38 7.47

3-Aug 7.61 7.51 7.63 7.55 7.41 7.51

4-Aug 7.56 7.43 7.56 7.50 7.36 7.46

5-Aug 7.50 7.37 7.50 7.53 7.39 7.39

6-Aug 7.43 7.39 7.40 7.50 7.43 7.31

7-Aug 7.40 7.48 7.43 7.57 7.45 7.33

8-Aug 7.32 7.52 7.39 7.50 7.42 7.32

9-Aug 7.27 7.42 7.38 7.39 7.37 7.33

10-Aug 7.27 7.37 7.45 7.46 7.32 7.35

11-Aug 7.32 7.40 7.48 7.54 7.36 7.43

12-Aug 7.33 7.43 7.42 7.54 7.41 7.40

13-Aug 7.37 7.44 7.45 7.47 7.45 7.34

14-Aug 7.37 7.38 7.44 7.54 7.46 7.35

15-Aug 7.41 7.42 7.36 7.56 7.43 7.39

16-Aug 7.50 7.37 7.35 7.55 7.37 7.37

17-Aug 7.53 7.47 7.32 7.48 7.40 7.35

18-Aug 7.55 7.51 7.35 7.44 7.43 7.40

19-Aug 7.41 7.32 7.29 7.29 7.38 7.28

20-Aug 7.37 7.33 7.36 7.30 7.43 7.23

21-Aug 7.39 7.34 7.44 7.42 7.57 7.36

22-Aug 7.51 7.31 7.45 7.39 7.62 7.40

23-Aug

24-Aug 7.61 7.54 7.54 7.61 7.67 7.57

25-Aug 7.57 7.55 7.50 7.61 7.63 7.50

26-Aug 7.70 7.61 7.42 7.70 7.51 7.53

28-Aug 7.52 7.43 7.35 7.69 7.51 7.52

29-Aug 7.46 7.43 7.23 7.64 7.40 7.41

30-Aug 7.51 7.49 7.21 7.70 7.34 7.32

31-Aug 7.45 7.49 7.35 7.65 7.34 7.35

1-Sep 7.62 7.55 7.48 7.65 7.44 7.37

2-Sep 7.69 7.49 7.52 7.69 7.46 7.33

3-Sep 7.74 7.52 7.69 7.74 7.53 7.52

4-Sep 7.75 7.58 7.76 7.79 7.59 7.60

5-Sep 7.72 7.60 7.78 7.79 7.54 7.64

6-Sep 7.72 7.60 7.78 7.79 7.54 7.64

7-Sep 7.69 7.45 7.44 7.71 7.41 7.74

8-Sep 7.60 7.46 7.32 7.74 7.37 7.60

9-Sep 7.62 7.57 7.18 7.73 7.32 7.51

10-Sep 7.67 7.49 7.10 7.75 7.30 7.49

11-Sep 7.70 7.61 7.21 7.74 7.34 7.56

12-Sep 7.72 7.60 7.35 7.78 7.35 7.69

13-Sep 7.63 7.52 7.39 7.74 7.33 7.78

14-Sep 7.57 7.51 7.34 7.70 7.36 7.83

15-Sep 7.55 7.53 7.24 7.65 7.34 7.82

16-Sep 7.68 7.50 7.26 7.71 7.32 7.77

17-Sep 7.70 7.51 7.29 7.71 7.33 7.76

18-Sep 7.84 7.39 7.25 7.71 7.35 7.77

19-Sep 7.62 7.35 7.27 7.68 7.29 7.69

20-Sep 7.58 7.44 7.34 7.67 7.26 7.62

21-Sep 7.88 7.46 7.34 7.65 7.35 7.69

22-Sep 7.67 7.56 7.41 7.45 7.34 7.65

23-Sep 7.76 7.58 7.37 7.40 7.33 7.59

24-Sep

25-Sep 7.66 7.44 7.29 7.26 7.26 7.35

26-Sep

27-Sep 7.51 7.42 7.34 7.38 7.44 7.23

28-Sep 7.61 7.48 7.36 7.51 7.55 7.26

29-Sep 7.65 7.39 7.35 7.54 7.55 7.20

30-Sep 7.64 7.24 7.33 7.56 7.54 7.30

1-Oct 7.64 7.26 7.33 7.64 7.51 7.37

2-Oct 7.63 7.39 7.32 7.67 7.49 7.30

3-Oct

4-Oct

5-Oct 7.60 7.41 7.36 7.75 7.49 7.31

6-Oct

7-Oct 7.60 7.42 7.36 7.78 7.47 7.39

Figure 5. Porous Pot Effluent pH Values
12/14/2011 Page 51 of 66

Appendix 6: Figure 6 - Porous Pot DOC Feed and Effluents

Feed A Feed B Feed C 1 2 3 4 5 6

12-May 310.398 316.130 316.772 17.195 11.863 12.431 12.887 14.817 10.413

13-May 323.046 321.604 321.728 13.227 15.342 13.220 12.990 16.077 12.142

14-May 313.892 314.746 321.610 14.684 17.409 14.468 13.690 17.959 13.849

15-May 305.876 327.414 332.126 14.495 14.161 18.376 14.024 15.209 17.925

16-May 318.106 329.830 331.640 13.711 12.700 17.998 13.076 13.527 17.853

17-May 311.960 327.690 331.468 12.881 13.083 18.845 12.278 12.477 19.944

18-May 318.772 329.720 330.044 12.906 12.740 18.765 13.556 13.730 20.410

19-May 313.296 323.344 329.574 14.350 15.506 20.804 14.350 14.181 19.876

20-May 272.580 286.502 284.856 14.861 14.060 18.592 14.383 14.180 19.391

21-May 312.214 320.656 327.432 16.580 14.957 21.984 15.965 17.324 21.712

22-May 321.518 316.058 310.722 18.519 16.866 22.555 16.454 18.761 24.559

23-May 323.230 323.756 317.790 20.229 19.077 23.349 16.571 17.046 24.231

24-May 308.490 319.932 315.658 21.052 20.525 41.704 17.337 19.330 32.580

25-May 297.152 309.944 311.882 23.396 21.497 75.121 17.636 19.986 34.103

26-May 290.896 300.350 305.654 24.677 25.577 74.135 18.171 26.590 47.658

27-May 303.948 320.906 321.942 25.563 37.302 48.765 20.633 31.911 39.951

28-May 310.456 325.792 324.302 27.795 38.224 33.350 25.853 32.850 44.821

29-May 308.198 329.032 318.428 32.055 40.734 53.497 23.635 39.629 51.591

30-May 296.924 276.622 268.186 38.556 36.171 78.989 21.435 37.213 66.243

31-May 216.942 296.162 297.132 40.411 46.503 109.984 20.625 52.842 86.845

1-Jun 296.924 276.622 268.186 110.847 39.752 143.003 21.819 54.170 95.653

2-Jun 309.826 312.510 310.448 36.631 35.625 122.379 24.765 31.562 60.394

3-Jun 314.672 315.498 303.930 35.408 34.163 64.634 23.197 29.790 38.381

4-Jun 313.684 306.896 296.604 25.778 30.849 45.249 20.403 25.652 34.204

5-Jun 314.274 300.292 299.738 24.414 26.534 39.658 18.906 21.720 43.952

6-Jun 261.456 257.154 235.154 21.898 23.696 43.764 18.753 23.738 40.287

7-Jun 204.636 301.458 278.546 21.512 21.598 39.313 18.922 22.008 62.674

8-Jun 322.322 335.992 319.156 20.129 20.363 36.067 19.547 19.282 37.601

9-Jun 336.056 337.702 330.460 18.257 16.911 33.803 18.752 17.586 29.144

10-Jun 330.406 334.096 331.746 16.352 13.618 26.687 18.165 16.279 30.173

11-Jun 310.176 335.696 313.558 19.105 14.642 26.226 17.517 15.001 32.621

12-Jun 311.394 322.772 318.046 17.836 15.879 27.491 17.122 13.982 31.409

13-Jun 289.226 302.326 289.092 16.745 17.467 26.470 16.379 14.133 31.842

14-Jun 277.004 304.380 288.482 16.921 16.429 24.133 17.134 13.417 30.072

15-Jun 323.524 316.720 311.228 16.830 16.997 22.378 16.218 12.160 27.404

16-Jun 331.302 328.136 323.884 17.694 18.709 24.452 19.130 14.697 27.252

17-Jun 333.544 332.582 320.976 19.066 19.989 23.082 18.820 16.230 25.117

18-Jun 332.956 335.962 333.660 23.874 24.348 24.102 19.231 23.853 24.861

19-Jun 329.614 334.470 331.158 26.433 29.704 25.941 19.841 32.051 27.063

20-Jun 278.082 291.078 278.800 25.624 29.934 24.671 20.212 27.498 23.934

21-Jun 286.218 284.252 280.466 29.820 31.296 27.129 28.732 27.801 24.856

22-Jun 273.466 276.470 275.190 33.364 33.462 30.625 37.706 35.463 27.315

23-Jun 276.566 294.430 279.050 34.683 32.957 34.865 40.621 47.134 28.881

24-Jun 306.758 311.944 304.682 34.006 31.203 36.116 39.870 48.725 28.024

25-Jun 293.562 306.332 297.578 35.440 30.835 33.907 41.929 45.603 28.067

26-Jun 305.384 306.356 298.104 35.583 38.343 32.526 46.820 38.563 27.105

Figure 6. Porous Pot DOC Feed and Effluents (mg/l DOC)
12/14/2011 Page 52 of 66

Feed A Feed B Feed C 1 2 3 4 5 6

27-Jun 277.322 300.150 282.706 36.638 45.194 30.356 41.004 36.676 31.391

28-Jun 272.306 293.016 273.298 40.541 45.753 29.512 36.005 38.039 36.945

29-Jun 272.306 293.016 273.298 54.947 40.619 82.448 34.307 58.378 82.663

30-Jun 296.130 297.244 293.114 58.806 40.602 89.648 40.328 53.557 59.334

1-Jul 307.998 307.116 310.264 39.933 37.372 46.079 46.377 37.911 38.859

2-Jul 308.760 316.152 311.286 31.779 31.256 42.059 42.875 53.276 33.021

3-Jul 283.986 299.444 277.836 28.970 30.295 41.395 41.661 65.009 32.403

4-Jul 273.722 284.732 306.266 23.856 24.307 29.441 32.022 38.571 30.051

5-Jul 272.864 275.804 271.366 14.309 11.763 17.811 14.068 19.159 16.284

6-Jul 235.468 239.252 231.832 11.747 8.449 15.297 11.373 17.490 13.665

7-Jul 312.272 311.134 306.592 11.830 8.623 15.178 13.382 16.489 16.560

8-Jul 276.180 312.632 311.290 11.570 8.991 15.406 13.794 16.002 15.762

9-Jul 298.728 311.296 298.594 9.496 8.664 15.919 11.637 13.499 15.759

10-Jul 317.728 325.822 328.198 10.925 10.282 16.795 10.027 10.059 17.069

11-Jul 328.520 344.872 340.120 12.123 10.257 23.031 10.784 11.208 21.277

12-Jul 320.754 329.708 327.200 10.177 9.635 24.781 9.901 11.296 21.062

13-Jul 308.686 326.120 319.074 9.601 7.604 21.760 12.228 9.700 20.971

14-Jul 339.732 340.814 327.676 10.750 8.695 19.603 14.165 11.253 21.720

15-Jul 341.820 338.308 337.690 11.782 9.664 20.341 13.150 12.519 20.116

16-Jul

17-Jul 232.758 233.788 240.148 12.684 10.059 20.030 12.014 11.731 19.936

18-Jul 232.886 231.650 211.742 11.512 9.846 18.593 9.657 10.122 18.202

19-Jul 214.990 208.772 239.962 10.142 9.585 17.149 8.769 8.777 16.910

20-Jul 211.682 217.640 225.588 10.135 10.248 17.453 9.646 9.934 18.026

21-Jul 207.632 228.256 192.536 9.948 9.276 18.599 10.538 9.871 19.865

22-Jul 170.546 162.330 187.782 10.278 8.374 18.656 10.342 10.016 19.230

23-Jul 291.736 297.840 295.308 11.071 10.128 18.546 12.114 12.021 17.728

24-Jul 281.972 282.374 263.638 12.209 11.293 19.924 13.765 12.585 19.956

25-Jul 271.570 248.690 278.154 12.292 10.240 19.288 12.273 10.609 19.466

26-Jul 284.028 284.438 284.248 11.954 8.959 17.847 9.425 9.459 17.849

27-Jul 299.512 290.840 272.186 10.939 7.975 18.242 8.797 7.508 18.240

28-Jul 300.796 293.262 295.778 9.882 8.158 17.745 9.287 7.811 18.582

29-Jul 280.696 283.508 283.382 9.548 8.711 18.532 9.386 7.597 19.619

30-Jul 284.598 279.092 271.936 9.377 8.547 18.041 9.578 7.721 18.567

31-Jul 301.422 304.646 308.092 10.393 9.521 19.124 9.134 7.809 18.238

1-Aug 231.226 263.306 256.808 9.078 16.723 18.275 9.476 13.278 17.869

2-Aug 273.140 249.208 242.722 10.443 21.702 19.466 9.916 19.507 17.994

3-Aug 251.392 247.012 248.248 10.186 19.422 19.289 9.594 19.641 19.032

4-Aug 275.774 281.930 285.424 9.462 19.051 19.051 9.397 19.272 18.114

5-Aug 276.810 274.422 265.708 8.544 18.627 19.138 8.548 18.773 17.710

6-Aug 270.762 271.014 262.204 8.349 17.032 17.478 7.592 17.735 16.699

7-Aug 274.794 276.876 274.850 8.612 16.222 17.900 8.730 18.433 17.053

8-Aug 265.740 277.300 223.008 8.506 17.371 21.936 9.676 18.421 17.222

9-Aug 270.040 269.878 264.678 8.633 18.979 23.501 10.896 18.210 17.241

10-Aug 232.574 247.624 229.542 9.122 19.029 22.332 11.119 21.136 18.452

11-Aug 276.030 289.588 263.990 9.647 18.593 15.954 11.359 23.890 12.555

12-Aug 284.890 258.046 273.788 9.763 18.736 11.577 11.251 22.779 9.150

13-Aug 269.506 279.070 262.104 10.869 18.046 10.947 12.736 22.367 9.964

14-Aug 277.934 276.676 246.624 12.368 17.517 10.210 11.895 21.079 10.961

15-Aug 247.940 239.436 243.984 11.797 16.757 8.796 11.953 20.191 10.419

16-Aug 227.632 242.160 235.068 10.478 21.293 8.290 10.862 19.181 9.695

17-Aug 281.066 271.800 270.500 11.509 17.302 8.570 14.866 17.302 11.568

18-Aug 285.140 270.510 269.560 11.688 18.969 9.307 15.705 22.359 12.018

Figure 6. Porous Pot DOC Feed and Effluents (mg/l DOC)
12/14/2011 Page 53 of 66

Feed A Feed B Feed C 1 2 3 4 5 6

19-Aug 275.450 271.450 269.650 11.848 19.597 10.053 15.251 22.014 13.800

20-Aug 289.350 278.560 281.450 17.829 15.622 11.347 21.568 17.571 14.611

21-Aug 279.750 278.800 277.350 14.716 17.644 11.615 19.332 18.968 16.138

22-Aug 280.540 278.340 281.650 12.209 23.570 12.708 17.113 21.705 17.584

23-Aug 298.340 287.560 285.350 11.595 25.438 13.405 16.266 21.192 16.477

24-Aug 286.440 279.340 281.680 12.185 26.305 13.977 15.317 19.852 14.895

25-Aug 295.740 289.440 290.680 11.531 26.019 14.925 14.278 19.862 15.076

26-Aug 298.420 295.560 300.120 10.975 24.315 13.342 12.606 18.374 14.906

27-Aug 330.952 316.158 318.600 10.818 23.274 13.905 12.286 18.981 15.615

28-Aug 307.162 290.204 300.828 10.349 23.970 14.862 11.129 18.040 12.299

29-Aug 304.460 301.334 292.984 10.875 24.091 16.367 11.683 18.669 13.277

30-Aug 325.070 320.074 320.618 11.779 25.882 16.180 12.318 22.884 12.634

31-Aug 291.448 298.708 293.032 20.714 27.283 15.182 20.531 24.304 12.591

1-Sep 294.140 285.684 288.552 15.978 26.113 13.724 15.113 22.066 13.315

2-Sep 312.742 309.938 313.838 11.366 25.023 13.010 10.581 21.006 13.188

3-Sep 290.976 289.786 290.064 10.427 25.200 12.917 10.439 21.760 14.452

4-Sep 288.360 296.040 290.426 10.066 24.601 12.045 9.636 21.613 13.879

5-Sep 309.616 312.466 321.020 9.901 23.713 11.765 9.223 21.243 12.706

6-Sep 287.982 284.368 285.684 10.149 23.138 10.926 9.594 21.749 12.154

7-Sep 284.012 281.480 275.438 10.503 22.527 11.856 10.411 22.286 12.281

8-Sep 316.224 310.068 314.572 12.498 23.158 13.973 11.436 23.409 13.398

9-Sep 298.202 283.170 284.760 12.835 23.764 16.722 11.529 24.306 14.156

10-Sep 289.590 291.248 281.028 12.443 22.332 20.085 11.069 23.412 13.958

11-Sep 313.108 314.666 321.496 12.641 22.748 17.689 10.469 22.751 13.364

12-Sep 298.286 292.386 285.142 11.709 20.968 13.183 10.070 22.204 12.454

13-Sep 295.626 286.828 284.828 11.608 21.284 11.801 9.767 22.282 11.568

14-Sep 319.138 315.114 319.714 11.620 24.308 11.753 9.710 22.193 10.784

15-Sep 319.138 315.114 319.714 16.719 24.853 11.740 10.765 22.927 11.152

16-Sep 291.394 281.264 291.620 17.648 22.944 11.996 10.704 22.809 10.475

17-Sep 309.616 315.482 315.364 17.593 23.992 13.251 10.528 23.299 10.270

18-Sep 300.366 295.708 302.800 16.903 25.509 12.364 10.486 23.497 10.234

19-Sep 299.902 294.464 288.270 14.706 23.861 11.515 10.180 22.900 10.007

20-Sep 305.896 315.156 317.666 13.268 23.232 10.770 10.482 22.389 9.932

21-Sep 295.528 286.798 288.486 13.377 23.479 11.675 11.486 23.480 11.445

22-Sep 294.174 292.998 287.644 16.809 23.499 12.035 15.046 24.853 11.901

23-Sep 324.054 296.356 322.126 12.245 23.338 15.133 13.439 23.302 13.533

24-Sep 301.344 300.094 296.898 12.698 23.617 13.262 15.840 23.046 11.314

25-Sep 300.364 292.060 297.084 11.130 22.490 11.974 15.955 22.102 11.065

26-Sep 317.504 321.496 321.742 10.436 22.485 11.243 16.001 21.810 10.419

27-Sep 290.010 302.172 291.054 10.373 22.744 10.329 15.872 20.982 11.089

28-Sep 306.476 296.318 288.270 10.404 23.165 10.408 14.406 21.075 11.006

29-Sep 281.454 265.706 259.256 10.932 23.759 10.758 13.608 20.633 10.953

30-Sep 309.146 315.066 326.780 10.891 22.285 10.801 13.099 20.355 11.242

1-Oct 281.634 299.206 305.734 10.495 22.540 10.844 14.001 19.488 10.390

2-Oct 309.044 304.350 305.666 11.365 26.022 11.399 12.684 20.468 10.976

3-Oct 299.736 282.914 293.654 12.626 20.104 10.331 11.082 19.193 11.403

4-Oct 281.762 263.418 266.922 12.244 20.534 10.092 11.092 19.931 12.310

5-Oct 300.662 297.672 298.714 12.810 19.083 8.635 10.097 19.076 12.698

6-Oct 294.066 283.294 279.308 13.882 19.841 9.215 10.321 18.704 13.530

7-Oct 280.762 283.152 278.744 12.498 20.916 8.630 10.060 19.672 13.849

Figure 6. Porous Pot DOC Feed and Effluents (mg/l DOC)
12/14/2011 Page 54 of 66

Appendix 7: Figure 7 - PPS Effluent DOC Un-acclimated ALFO 23-4S

POT 1 2 3 4 5 6

7-May 13.056 14.259 17.506 15.989 24.859 14.375

8-May 11.390 12.079 14.995 13.005 14.419 10.447

9-May 11.166 8.076 13.943 10.410 16.768 7.723

10-May 13.851 9.932 12.975 13.780 16.504 8.973

11-May 14.648 8.654 12.079 15.043 17.640 9.791

12-May 17.195 11.863 12.431 12.887 14.817 10.413

13-May 13.227 15.342 13.220 12.990 16.077 12.142

14-May 14.684 17.409 14.468 13.690 17.959 13.849

15-May 14.495 14.161 18.376 14.024 15.209 17.925

16-May 13.711 12.700 17.998 13.076 13.527 17.853

17-May 12.881 13.083 18.845 12.278 12.477 19.944

18-May 12.906 12.740 18.765 13.556 13.730 20.410

19-May 14.350 15.506 20.804 14.350 14.181 19.876

20-May 14.861 14.060 18.592 14.383 14.180 19.391

21-May 16.580 14.957 21.984 15.965 17.324 21.712

22-May 18.519 16.866 22.555 16.454 18.761 24.559

23-May 20.229 19.077 23.349 16.571 17.046 24.231

24-May 21.052 20.525 41.704 17.337 19.330 32.580

25-May 23.396 21.497 75.121 17.636 19.986 34.103

26-May 24.677 25.577 74.135 18.171 26.590 47.658

27-May 25.563 37.302 48.765 20.633 31.911 39.951

28-May 27.795 38.224 33.350 25.853 32.850 44.821

29-May 32.055 40.734 53.497 23.635 39.629 51.591

30-May 38.556 36.171 78.989 21.435 37.213 66.243

31-May 40.411 46.503 109.984 20.625 52.842 86.845

1-Jun 110.847 39.752 143.003 21.819 54.170 95.653

2-Jun 36.631 35.625 122.379 24.765 31.562 60.394

3-Jun 35.408 34.163 64.634 23.197 29.790 38.381

4-Jun 25.778 30.849 45.249 20.403 25.652 34.204

5-Jun 24.414 26.534 39.658 18.906 21.720 43.952

6-Jun 21.898 23.696 43.764 18.753 23.738 40.287

7-Jun 21.512 21.598 39.313 18.922 22.008 62.674

8-Jun 20.129 20.363 36.067 19.547 19.282 37.601

9-Jun 18.257 16.911 33.803 18.752 17.586 29.144

10-Jun 16.352 13.618 26.687 18.165 16.279 30.173

11-Jun 19.105 14.642 26.226 17.517 15.001 32.621

12-Jun 17.836 15.879 27.491 17.122 13.982 31.409

13-Jun 16.745 17.467 26.470 16.379 14.133 31.842

14-Jun 16.921 16.429 24.133 17.134 13.417 30.072

15-Jun 16.830 16.997 22.378 16.218 12.160 27.404

16-Jun 17.694 18.709 24.452 19.130 14.697 27.252

17-Jun 19.066 19.989 23.082 18.820 16.230 25.117

18-Jun 23.874 24.348 24.102 19.231 23.853 24.861

19-Jun 26.433 29.704 25.941 19.841 32.051 27.063

20-Jun 25.624 29.934 24.671 20.212 27.498 23.934

21-Jun 29.820 31.296 27.129 28.732 27.801 24.856

22-Jun 33.364 33.462 30.625 37.706 35.463 27.315

23-Jun 34.683 32.957 34.865 40.621 47.134 28.881

24-Jun 34.006 31.203 36.116 39.870 48.725 28.024

25-Jun 35.440 30.835 33.907 41.929 45.603 28.067

26-Jun 35.583 38.343 32.526 46.820 38.563 27.105

27-Jun 36.638 45.194 30.356 41.004 36.676 31.391

28-Jun 40.541 45.753 29.512 36.005 38.039 36.945

29-Jun 54.947 40.619 82.448 34.307 58.378 82.663

30-Jun 58.806 40.602 89.648 40.328 53.557 59.334

1-Jul 39.933 37.372 46.079 46.377 37.911 38.859

2-Jul 31.779 31.256 42.059 42.875 53.276 33.021

3-Jul 28.970 30.295 41.395 41.661 65.009 32.403

Figure 7. Porous Pot Effluents DOC Pre Unacclimated ALFO 23-4S (mg/l)
12/14/2011 Page 55 of 66

POT 1 2 3 4 5 6

4-Jul 23.856 24.307 29.441 32.022 38.571 30.051

5-Jul 14.309 11.763 17.811 14.068 19.159 16.284

6-Jul 11.747 8.449 15.297 11.373 17.490 13.665

7-Jul 11.830 8.623 15.178 13.382 16.489 16.560

8-Jul 11.570 8.991 15.406 13.794 16.002 15.762

9-Jul 9.496 8.664 15.919 11.637 13.499 15.759

10-Jul 10.925 10.282 16.795 10.027 10.059 17.069

11-Jul 12.123 10.257 23.031 10.784 11.208 21.277

12-Jul 10.177 9.635 24.781 9.901 11.296 21.062

13-Jul 9.601 7.604 21.760 12.228 9.700 20.971

14-Jul 10.750 8.695 19.603 14.165 11.253 21.720

15-Jul 11.782 9.664 20.341 13.150 12.519 20.116

16-Jul

17-Jul 12.684 10.059 20.030 12.014 11.731 19.936

18-Jul 11.512 9.846 18.593 9.657 10.122 18.202

19-Jul 10.142 9.585 17.149 8.769 8.777 16.910

20-Jul 10.135 10.248 17.453 9.646 9.934 18.026

21-Jul 9.948 9.276 18.599 10.538 9.871 19.865

22-Jul 10.278 8.374 18.656 10.342 10.016 19.230

23-Jul 11.071 10.128 18.546 12.114 12.021 17.728

24-Jul 12.209 11.293 19.924 13.765 12.585 19.956

25-Jul 12.292 10.240 19.288 12.273 10.609 19.466

26-Jul 11.954 8.959 17.847 9.425 9.459 17.849

27-Jul 10.939 7.975 18.242 8.797 7.508 18.240

28-Jul 9.882 8.158 17.745 9.287 7.811 18.582

29-Jul 9.548 8.711 18.532 9.386 7.597 19.619

30-Jul 9.377 8.547 18.041 9.578 7.721 18.567

31-Jul 10.393 9.521 19.124 9.134 7.809 18.238

1-Aug 9.078 16.723 18.275 9.476 13.278 17.869

2-Aug 10.443 21.702 19.466 9.916 19.507 17.994

3-Aug 10.186 19.422 19.289 9.594 19.641 19.032

4-Aug 9.462 19.051 19.051 9.397 19.272 18.114

5-Aug 8.544 18.627 19.138 8.548 18.773 17.710

6-Aug 8.349 17.032 17.478 7.592 17.735 16.699

7-Aug 8.612 16.222 17.900 8.730 18.433 17.053

8-Aug 8.506 17.371 21.936 9.676 18.421 17.222

9-Aug 8.633 18.979 23.501 10.896 18.210 17.241

10-Aug 9.122 19.029 22.332 11.119 21.136 18.452

11-Aug 9.647 18.593 15.954 11.359 23.890 12.555

12-Aug 9.763 18.736 11.577 11.251 22.779 9.150

13-Aug 10.869 18.046 10.947 12.736 22.367 9.964

14-Aug 12.368 17.517 10.210 11.895 21.079 10.961

15-Aug 11.797 16.757 8.796 11.953 20.191 10.419

16-Aug 10.478 21.293 8.290 10.862 19.181 9.695

17-Aug 11.509 17.302 8.570 14.866 17.302 11.568

18-Aug 11.688 18.969 9.307 15.705 22.359 12.018

19-Aug 11.848 19.597 10.053 15.251 22.014 13.800

20-Aug 17.829 15.622 11.347 21.568 17.571 14.611

21-Aug 14.716 17.644 11.615 19.332 18.968 16.138

22-Aug 12.209 23.570 12.708 17.113 21.705 17.584

23-Aug 11.595 25.438 13.405 16.266 21.192 16.477

24-Aug 12.185 26.305 13.977 15.317 19.852 14.895

25-Aug 11.531 26.019 14.925 14.278 19.862 15.076

26-Aug 10.975 24.315 13.342 12.606 18.374 14.906

Figure 7. Porous Pot Effluents DOC Pre Unacclimated ALFO 23-4S (mg/l)
12/14/2011 Page 56 of 66

POT 1 2 3 4 5 6

27-Aug 10.818 23.274 13.905 12.286 18.981 15.615

28-Aug 10.349 23.970 14.862 11.129 18.040 12.299

29-Aug 10.875 24.091 16.367 11.683 18.669 13.277

30-Aug 11.779 25.882 16.180 12.318 22.884 12.634

31-Aug 20.714 27.283 15.182 20.531 24.304 12.591

1-Sep 15.978 26.113 13.724 15.113 22.066 13.315

2-Sep 11.366 25.023 13.010 10.581 21.006 13.188

3-Sep 10.427 25.200 12.917 10.439 21.760 14.452

4-Sep 10.066 24.601 12.045 9.636 21.613 13.879

5-Sep 9.901 23.713 11.765 9.223 21.243 12.706

6-Sep 10.149 23.138 10.926 9.594 21.749 12.154

7-Sep 10.503 22.527 11.856 10.411 22.286 12.281

8-Sep 12.498 23.158 13.973 11.436 23.409 13.398

9-Sep 12.835 23.764 16.722 11.529 24.306 14.156

10-Sep 12.443 22.332 20.085 11.069 23.412 13.958

11-Sep 12.641 22.748 17.689 10.469 22.751 13.364

12-Sep 11.709 20.968 13.183 10.070 22.204 12.454

13-Sep 11.608 21.284 11.801 9.767 22.282 11.568

14-Sep 11.620 24.308 11.753 9.710 22.193 10.784

15-Sep 16.719 24.853 11.740 10.765 22.927 11.152

16-Sep 17.648 22.944 11.996 10.704 22.809 10.475

17-Sep 17.593 23.992 13.251 10.528 23.299 10.270

18-Sep 16.903 25.509 12.364 10.486 23.497 10.234

19-Sep 14.706 23.861 11.515 10.180 22.900 10.007

20-Sep 13.268 23.232 10.770 10.482 22.389 9.932

21-Sep 13.377 23.479 11.675 11.486 23.480 11.445

22-Sep 16.809 23.499 12.035 15.046 24.853 11.901

23-Sep 12.245 23.338 15.133 13.439 23.302 13.533

24-Sep 12.698 23.617 13.262 15.840 23.046 11.314

25-Sep 11.130 22.490 11.974 15.955 22.102 11.065

26-Sep 10.436 22.485 11.243 16.001 21.810 10.419

27-Sep 10.373 22.744 10.329 15.872 20.982 11.089

28-Sep 10.404 23.165 10.408 14.406 21.075 11.006

29-Sep 10.932 23.759 10.758 13.608 20.633 10.953

30-Sep 10.891 22.285 10.801 13.099 20.355 11.242

1-Oct 10.495 22.540 10.844 14.001 19.488 10.390

2-Oct 11.365 26.022 11.399 12.684 20.468 10.976

3-Oct 12.626 20.104 10.331 11.082 19.193 11.403

4-Oct 12.244 20.534 10.092 11.092 19.931 12.310

5-Oct 12.810 19.083 8.635 10.097 19.076 12.698

6-Oct 13.882 19.841 9.215 10.321 18.704 13.530

7-Oct 12.498 20.916 8.630 10.060 19.672 13.849

8-Oct

9-Oct

10-Oct

11-Oct

27-Jan 7.838 24.293 7.210 18.717 6.927 19.858

28-Jan 7.041 22.283 6.789 18.129 7.411 17.741

29-Jan 7.092 21.705 7.836 18.065 6.949 16.841

30-Jan 10.384 23.765 7.704 20.328 10.819 18.908

31-Jan 10.251 25.144 7.103 19.607 10.053 18.549

1-Feb 8.195 22.245 12.655 20.611 11.038 18.989

2-Feb 10.221 19.207 5.564 16.981 8.556 15.924

Figure 7. Porous Pot Effluents DOC Pre Unacclimated ALFO 23-4S (mg/l)
12/14/2011 Page 57 of 66

POT 1 2 3 4 5 6

3-Feb 10.358 18.358 5.109 16.509 14.327 14.957

4-Feb - - - - - 15.533

5-Feb 12.659 21.732 9.762 20.251 28.741 19.345

6-Feb 12.166 21.871 9.975 20.913 25.082 19.323

7-Feb 11.517 20.609 9.982 21.598 22.925 18.706

8-Feb 11.990 19.378 9.790 20.600 22.808 17.971

9-Feb 12.095 19.527 10.109 19.721 21.545 18.237

10-Feb 10.137 18.812 9.586 19.784 21.942 18.208

11-Feb 9.497 17.350 8.865 19.106 21.082 17.630

12-Feb 9.997 17.536 9.153 19.341 21.871 17.226

13-Feb 10.032 16.785 8.374 18.517 21.087 16.087

14-Feb 10.476 17.272 8.270 20.282 24.402 16.910

15-Feb 11.584 17.668 8.407 20.103 24.752 17.514

16-Feb 11.927 17.335 8.790 19.784 24.524 17.137

17-Feb 10.369 16.625 7.578 18.733 24.478 16.751

18-Feb 9.327 16.554 8.015 19.235 25.926 17.089

19-Feb 8.934 16.010 8.114 18.964 25.227 17.168

20-Feb 7.545 15.258 7.747 17.636 21.555 16.359

21-Feb 7.530 14.440 7.186 17.531 20.581 15.640

22-Feb 7.497 13.977 7.469 16.378 20.017 15.460

23-Feb 9.161 15.515 11.841 17.481 20.528 17.275

24-Feb 8.660 15.284 9.224 17.862 19.937 17.141

25-Feb 9.038 15.848 9.130 19.524 19.951 16.359

26-Feb 8.424 14.550 8.773 20.421 17.607 15.653

27-Feb 9.765 15.294 9.459 20.583 17.964 16.959

28-Feb 7.665 15.685 11.315 19.864 17.700 16.761

1-Mar 10.946 17.418 13.874 20.555 18.541 18.291

2-Mar 10.296 18.553 13.816 20.334 18.462 19.059

3-Mar 8.601 17.760 10.301 20.134 17.334 16.922

4-Mar 9.806 18.244 10.668 22.056 17.343 17.279

5-Mar 9.758 17.889 9.865 20.952 17.186 17.622

6-Mar 9.325 18.858 9.108 19.876 15.658 16.091

7-Mar 9.454 21.583 10.190 20.830 16.501 17.315

8-Mar 8.923 20.579 10.887 20.855 17.797 17.471

9-Mar 9.719 20.555 12.137 24.563 19.187 18.293

10-Mar 10.355 20.227 11.715 22.588 17.553 16.323

11-Mar 8.056 20.842 11.683 20.263 18.006 16.174

12-Mar 7.939 21.044 10.028 19.523 13.407 15.065

13-Mar 8.215 20.371 9.271 20.698 18.992 14.248

14-Mar 8.206 20.036 8.356 18.752 19.452 13.680

15-Mar 8.368 19.445 7.713 18.704 18.478 13.063

16-Mar 7.378 19.223 7.730 17.882 17.791 12.846

17-Mar 7.465 18.520 7.977 17.236 18.189 12.594

18-Mar 7.618 18.036 8.110 17.691 17.503 12.635

19-Mar 8.326 17.975 8.432 15.766 17.404 12.503

20-Mar 8.933 16.577 8.026 15.793 17.617 12.499

21-Mar 10.063 16.427 7.689 15.924 17.235 12.687

22-Mar 10.129 15.075 7.977 16.070 16.588 12.476

23-Mar 8.997 15.431 7.815 15.484 16.694 12.563

24-Mar 8.445 15.294 7.937 15.964 16.825 13.244

25-Mar 8.544 15.374 7.494 16.134 16.928 13.739

26-Mar 8.237 14.972 7.345 15.932 17.945 13.896

Figure 7. Porous Pot Effluents DOC Pre Unacclimated ALFO 23-4S (mg/l)
12/14/2011 Page 58 of 66

Appendix 8: Figure 8 - PPS for Fragment Analysis Effluent

Blank (1) Alfo (2) Blank (3) Alfo (6)

27-Jan 7.84 24.29 7.21 19.86

28-Jan 7.04 22.28 6.79 17.74

29-Jan 7.09 21.71 7.84 16.84

30-Jan 10.38 23.77 7.70 18.91

31-Jan 10.25 25.14 7.10 18.55

1-Feb 8.19 22.25 12.66 18.99

2-Feb 10.22 19.21 5.56 15.92

3-Feb 10.36 18.36 5.11 14.96

5-Feb 12.66 21.73 9.76 19.35

6-Feb 12.17 21.87 9.97 19.32

7-Feb 11.52 20.61 9.98 18.71

8-Feb 11.99 19.38 9.79 17.97

9-Feb 12.10 19.53 10.11 18.24

10-Feb 10.14 18.81 9.59 18.21

11-Feb 9.50 17.35 8.86 17.63

12-Feb 10.00 17.54 9.15 17.23

13-Feb 10.03 16.79 8.37 16.09

14-Feb 10.48 17.27 8.27 16.91

15-Feb 11.58 17.67 8.41 17.51

16-Feb 11.93 17.33 8.79 17.14

17-Feb 10.37 16.63 7.58 16.75

18-Feb 9.33 16.55 8.02 17.09

19-Feb 8.93 16.01 8.11 17.17

20-Feb 7.55 15.26 7.75 16.36

21-Feb 7.53 14.44 7.19 15.64

22-Feb 7.50 13.98 7.47 15.46

23-Feb 9.16 15.52 11.84 17.27

24-Feb 8.66 15.28 9.22 17.14

25-Feb 9.04 15.85 9.13 16.36

26-Feb 8.42 14.55 8.77 15.65

27-Feb 9.77 15.29 9.46 16.96

28-Feb 7.67 15.69 11.31 16.76

1-Mar 10.95 17.42 13.87 18.29

2-Mar 10.30 18.55 13.82 19.06

3-Mar 8.60 17.76 10.30 16.92

4-Mar 9.81 18.24 10.67 17.28

5-Mar 9.76 17.89 9.86 17.62

6-Mar 9.32 18.86 9.11 16.09

7-Mar 9.45 21.58 10.19 17.31

8-Mar 8.92 20.58 10.89 17.47

9-Mar 9.72 20.55 12.14 18.29

10-Mar 10.35 20.23 11.72 16.32

11-Mar 8.06 20.84 11.68 16.17

12-Mar 7.94 21.04 10.03 15.06

13-Mar 8.21 20.37 9.27 14.25

14-Mar 8.21 20.04 8.36 13.68

15-Mar 8.37 19.45 7.71 13.06

16-Mar 7.38 19.22 7.73 12.85

17-Mar 7.46 18.52 7.98 12.59

18-Mar 7.62 18.04 8.11 12.64

19-Mar 8.33 17.98 8.43 12.50

20-Mar 8.93 16.58 8.03 12.50

21-Mar 10.06 16.43 7.69 12.69

22-Mar 10.13 15.08 7.98 12.48

23-Mar 9.00 15.43 7.81 12.56

24-Mar 8.44 15.29 7.94 13.24

25-Mar 8.54 15.37 7.49 13.74

26-Mar 8.24 14.97 7.34 13.90

Porous Pot Study for Fragment Analysis Effluent 
12/14/2011 Page 59 of 66

Appendix 9: Figure 9 - Return to Steady State: Post Un-acclimated ALFO 23-4S

Pot 1 2 3 4 5 6

Blnk U-Alfo A-Alfo Blnk U-Alfo A-Alfo

30-Jul 9.38 8.55 18.04 9.58 7.72 18.57

31-Jul 10.39 9.52 19.12 9.13 7.81 18.24

1-Aug 9.08 16.72 18.28 9.48 13.28 17.87

2-Aug 10.44 21.70 19.47 9.92 19.51 17.99

3-Aug 10.19 19.42 19.29 9.59 19.64 19.03

4-Aug 9.46 19.05 19.05 9.40 19.27 18.11

5-Aug 8.54 18.63 19.14 8.55 18.77 17.71

6-Aug 8.35 17.03 17.48 7.59 17.73 16.70

7-Aug 8.61 16.22 17.90 8.73 18.43 17.05

8-Aug 8.51 17.37 21.94 9.68 18.42 17.22

9-Aug 8.63 18.98 23.50 10.90 18.21 17.24

Figure 9. Return to Steady State: Post Unacclimated ALFO 23-4S

Pre AES (mg/l DOC)
12/14/2011 Page 60 of 66

Appendix 10: Figure 10 - Porous Pot DOC: Comparison of 1214E4S and ALFO 23-4S

Pot 1 2 3 4 5 6

AES Alfo Blnk AES Alfo Blnk

14-Aug 12.37 17.52 10.21 11.89 21.08 10.96

15-Aug 11.80 16.76 8.80 11.95 20.19 10.42

16-Aug 10.48 21.29 8.29 10.86 19.18 9.69

17-Aug 11.51 17.30 8.57 14.87 17.30 11.57

18-Aug 11.69 18.97 9.31 15.70 22.36 12.02

19-Aug 11.85 19.60 10.05 15.25 22.01 13.80

20-Aug 17.83 15.62 11.35 21.57 17.57 14.61

21-Aug 14.72 17.64 11.62 19.33 18.97 16.14

22-Aug 12.21 23.57 12.71 17.11 21.70 17.58

23-Aug 11.60 25.44 13.40 16.27 21.19 16.48

24-Aug 12.18 26.30 13.98 15.32 19.85 14.90

25-Aug 11.53 26.02 14.93 14.28 19.86 15.08

26-Aug 10.98 24.32 13.34 12.61 18.37 14.91

27-Aug 10.82 23.27 13.91 12.29 18.98 15.62

28-Aug 10.35 23.97 14.86 11.13 18.04 12.30

29-Aug 10.87 24.09 16.37 11.68 18.67 13.28

30-Aug 11.78 25.88 16.18 12.32 22.88 12.63

31-Aug 20.71 27.28 15.18 20.53 24.30 12.59

1-Sep 15.98 26.11 13.72 15.11 22.07 13.31

2-Sep 11.37 25.02 13.01 10.58 21.01 13.19

3-Sep 10.43 25.20 12.92 10.44 21.76 14.45

4-Sep 10.07 24.60 12.05 9.64 21.61 13.88

5-Sep 9.90 23.71 11.77 9.22 21.24 12.71

6-Sep 10.15 23.14 10.93 9.59 21.75 12.15

7-Sep 10.50 22.53 11.86 10.41 22.29 12.28

8-Sep 12.50 23.16 13.97 11.44 23.41 13.40

9-Sep 12.84 23.76 16.72 11.53 24.31 14.16

10-Sep 12.44 22.33 20.08 11.07 23.41 13.96

11-Sep 12.64 22.75 17.69 10.47 22.75 13.36

12-Sep 11.71 20.97 13.18 10.07 22.20 12.45

13-Sep 11.61 21.28 11.80 9.77 22.28 11.57

14-Sep 11.62 24.31 11.75 9.71 22.19 10.78

15-Sep 16.72 24.85 11.74 10.76 22.93 11.15

16-Sep 17.65 22.94 12.00 10.70 22.81 10.47

17-Sep 17.59 23.99 13.25 10.53 23.30 10.27

18-Sep 16.90 25.51 12.36 10.49 23.50 10.23

19-Sep 14.71 23.86 11.51 10.18 22.90 10.01

20-Sep 13.27 23.23 10.77 10.48 22.39 9.93

21-Sep 13.38 23.48 11.68 11.49 23.48 11.44

22-Sep 16.81 23.50 12.03 15.05 24.85 11.90

23-Sep 12.25 23.34 15.13 13.44 23.30 13.53

24-Sep 12.70 23.62 13.26 15.84 23.05 11.31

Figure 10. Porous Pot DOC: A Comparison of 1214E4S and ALFO 23-4S

(mg/l DOC)
12/14/2011 Page 61 of 66

Appendix 11: Figure 11 - PPS: Increased HRT

Pot 1 2 3 4 5 6

AES Alfo Blnk AES Alfo Blnk

20-Sep 13.27 23.23 10.77 10.48 22.39 9.93

21-Sep 13.38 23.48 11.68 11.49 23.48 11.44

22-Sep 16.81 23.50 12.03 15.05 24.85 11.90

23-Sep 12.25 23.34 15.13 13.44 23.30 13.53

24-Sep 12.70 23.62 13.26 15.84 23.05 11.31

25-Sep 11.13 22.49 11.97 15.96 22.10 11.07

26-Sep 10.44 22.48 11.24 16.00 21.81 10.42

27-Sep 10.37 22.74 10.33 15.87 20.98 11.09

28-Sep 10.40 23.17 10.41 14.41 21.08 11.01

29-Sep 10.93 23.76 10.76 13.61 20.63 10.95

30-Sep 10.89 22.29 10.80 13.10 20.35 11.24

1-Oct 10.50 22.54 10.84 14.00 19.49 10.39

2-Oct 11.36 26.02 11.40 12.68 20.47 10.98

3-Oct 12.63 20.10 10.33 11.08 19.19 11.40

4-Oct 12.24 20.53 10.09 11.09 19.93 12.31

5-Oct 12.81 19.08 8.63 10.10 19.08 12.70

6-Oct 13.88 19.84 9.21 10.32 18.70 13.53

7-Oct 12.50 20.92 8.63 10.06 19.67 13.85

8-Oct

9-Oct 12.50 20.92 8.63 10.06 19.67 13.85

10-Oct 12.64 21.18 8.30 10.06 17.86 13.49

Figure 11. Porous Pot: Increased Hydraulic Rentenion Time 

(6 Hr to 12 hr) mg/l DOC

Appendix 12: Figure 12 - Die-Away Porous Pot Effluent

Figure 12. Die-Away Porous Pot Effluent (mg/l)

Days 0 1 4 7 12 17 24 28

PP Blank 16.854 15.159 12.4125 11.1985 9.7205 7.8225 7.4825 6.9545

PP AES 11.659 12.634 9.7695 8.5245 7.7485 6.3905 5.189 5.0195

PP ALFO 23-4S 23.473 22.232 21.461 20.473 19.2685 17.359 16.591 16.436

Appendix 13: Figure 13 - OECD Die-Away DOC Loss %

Figure 13. OECD Die-Away DOC Loss (%)

Days 0 1 4 7 12 17 24 28

AES 0 8.26 54.29 60.68 93.28 99.49 98.99 98.9

ALFO 23-4S 0 1.62 9.7 9.67 9.21 17.87 19.81 20.5

Benz 0 17.75 95.9 97.4 98.5 100 99.63 99.3
12/14/2011 Page 62 of 66

Appendix 14: Figure 14 - Porous Pot: MBAS Primary Biodegradation

Influent Effluent

Blank 1.2 0.1

AES 44.0 0.1

ALFO 23-4S 39.0 2.3

Influent Effluent

Porous Pot 39.0 2.3

Die-away 46.0 4.6

Figure 14. Porous Pot: MBAS Primary 

Biodegradation (mg/l)

Figure 15. ALFO 23-4S Comparison Porous Pot

and Die-away 

MBAS (mg/l)

Appendix 15: ALFO 23-4S Comparison Porous Pot and Die Away

Appendix 16: Figure 16 - AES and ALFO 23-4S Acute Toxicity

Appendix 17: Figure 17 - Porous Pot Aquatic Toxicity 96 Hour Minnow

Appendix 18: Figure 18- OECD Die-Away Test ALFO 23-4S

LC50 LOEC NOEC

AES 5.5 2.4 4.8

ALFO 23-4S 3.1 1.2 2.4

LC50 LOEC NOEC

Blank Feed 14.7 25.0 12.0

Blank Effluent 100.0 100.0 100.0

AES + Feed 5.5 6.0 3.0

AES Effluent 100.0 100.0 100.0

ALFO 23-4S + Feed 9.5 12.0 6.0

ALFO 23-4S Effluent 100.0 100.0 100.0

LC50 LOEC NOEC

ALFO 23-4S Day 0 8.8 12.0 6.0

ALFO 23-4S Day 28 100.0 100.0 100.0

Acute Toxicity 96 hr Fathead Minnow (mg/l)

Figure 16. AES and ALFO 23-4S

Figure 17 - Porous Pot Aquatic Toxicity 96 hr. Minnow (mg/l)

40 mg/l AES and ALFO 23-4S active

Figure 18. OECD Die-Away Test ALFO 23-4S

50 mg/l active or 22 mg/l DOC
12/14/2011 Page 63 of 66

Appendix 19: Figure 19: ALFOTERRA Hydrophobe Biodegradation Study - OECD Demand

Isalchem

23

Alchem

23 TDA Safol 23 Lial 23 Alfol 12 Ester

0 0 0 0 0 0 0 0

1 10.30 13.14 2.34 12.90 14.43 14.31 -0.13

2 18.92 25.66 9.87 25.39 23.75 27.47 0.27

3 23.13 37.62 17.48 33.67 29.10 38.85 0.44

4 27.75 47.88 22.57 41.31 35.01 49.49 0.61

5 33.45 56.08 27.13 48.47 40.51 57.01 0.81

6 38.46 61.58 31.92 53.53 45.21 61.79 0.96

7 43.08 65.54 36.19 58.02 49.67 66.37 0.96

8 47.99 68.90 40.63 61.31 54.02 70.68 0.93

9 53.21 71.72 45.31 63.46 57.69 74.03 0.95

10 57.97 73.63 49.83 65.43 60.73 76.40 1.00

11 61.95 75.36 53.71 67.65 63.31 78.51 1.14

12 65.29 77.04 56.84 69.77 65.48 80.38 1.27

13 68.15 78.71 59.40 71.74 67.38 82.48 1.40

14 70.92 80.82 61.66 73.60 69.13 84.78 1.50

15 73.54 82.54 63.56 75.31 70.73 86.98 1.62

16 76.03 84.41 65.24 76.86 72.50 89.28 1.74

17 78.28 86.04 66.67 78.71 73.98 91.17 1.87

18 80.58 87.28 68.19 80.61 75.68 92.68 1.97

19 82.75 88.28 69.72 82.52 77.63 94.21 2.07

20 84.79 89.14 71.17 84.10 79.43 95.38 2.19

21 86.78 89.89 72.44 85.36 80.64 96.28 2.30

22 88.77 90.77 73.73 86.75 81.63 97.14 2.44

23 90.58 91.72 75.22 88.08 82.43 97.82 2.56

24 92.25 92.52 76.73 89.33 83.13 98.49 2.68

25 93.84 93.14 78.26 90.66 83.82 99.16 2.80

26 95.31 93.62 79.74 91.55 84.52 99.84 2.94

27 96.74 94.08 81.26 92.32 85.46 100.52 3.05

28 98.14 94.48 82.70 92.97 86.46 101.26 3.18

Figure 19. ALFOTERRA Hydrophobe Biodegradation Study

OECD 301F Theoretical Oxygen demand
12/14/2011 Page 64 of 66

Appendix 20: Figure 20 - ALFOTERRA Hydrophobe 4PO Biodegradation Study - OECD

Day

Isalchem

23

Alchem

23 TDA Safol 23 Lial 23 Alfol 12 Benzoate

0 0 0 0 0 0 0 0

1 0.97 5.60 0.28 4.48 3.74 7.74 30.41

2 3.64 18.64 1.29 16.24 11.93 18.98 57.22

3 9.64 22.32 2.81 21.61 18.57 22.34 66.56

4 16.92 27.55 3.73 26.46 24.72 26.49 71.08

5 22.86 32.40 5.95 31.69 29.96 31.20 75.09

6 29.42 37.16 11.70 36.57 35.10 35.36 78.74

7 35.81 41.50 20.36 40.47 40.46 39.11 81.26

8 41.09 45.68 31.21 44.46 45.26 42.74 83.09

9 45.94 49.76 39.65 48.67 49.78 46.67 84.70

10 50.34 53.41 46.04 52.66 53.74 51.72 86.08

11 54.80 56.95 50.94 56.60 57.71 53.64 87.51

12 58.91 60.29 55.37 60.29 61.42 57.09 88.76

13 62.74 63.25 59.24 63.59 64.80 60.19 89.88

14 66.31 65.88 62.96 66.56 67.89 62.87 90.86

15 69.79 68.25 66.27 69.55 70.86 65.37 91.86

16 73.03 70.50 68.91 72.45 73.64 67.73 92.90

17 75.95 72.68 71.40 75.13 76.19 69.90 93.76

18 78.80 75.02 74.31 77.78 78.72 72.09 94.74

19 81.52 77.36 77.24 80.22 81.08 74.14 95.54

20 84.21 79.69 79.98 82.71 83.47 76.30 96.47

21 86.83 82.02 82.32 85.18 85.78 78.36 97.40

22 89.30 84.15 84.43 87.26 88.02 80.14 98.02

23 92.08 86.80 86.79 89.56 90.51 82.02 99.23

24 94.61 89.21 89.61 91.41 92.67 83.45 100.06

25 97.12 91.39 91.79 93.07 94.73 84.76 101.84

26 99.49 93.47 93.98 94.55 96.64 85.94 102.25

27 101.92 95.81 96.29 95.97 98.44 87.03 102.94

28 104.22 98.51 98.31 97.44 100.21 88.25 104.18

Figure 20. ALFOTERRA Hydrophobe 4PO Biodegradation Study

OECD 301F Theoretical Oxygen demand (%)
12/14/2011 Page 65 of 66

Appendix 21: Figure 21- ALFOTERRA Hydrophobe 4PO Sulfate Biodegradation Study - OECD

DAY

Isalchem

23

Alchem

23 TDA Lial 23 Safol 23 Alfol 12 Benzoate

0 0 0 0 0 0 0 0

1 1.25 7.86 0.55 3.54 4.99 7.44 26.91

2 4.08 13.22 0.54 9.51 12.14 13.72 49.60

3 14.27 16.76 4.05 13.56 15.33 17.18 60.60

4 17.71 19.88 8.41 17.45 19.47 20.48 67.90

5 19.48 22.42 11.90 20.79 22.89 22.84 73.19

6 20.99 25.53 14.09 23.67 25.85 25.53 76.91

7 22.23 30.40 15.55 26.55 29.36 30.10 79.54

8 24.20 34.59 16.73 28.38 32.00 35.28 81.26

9 25.95 36.97 17.75 29.76 33.74 37.88 82.50

10 27.33 38.79 18.60 31.06 35.23 39.47 83.66

11 28.39 40.01 19.27 32.03 36.29 40.72 84.57

12 29.21 40.85 19.82 32.77 37.10 41.53 85.45

13 29.94 41.53 20.33 33.32 37.88 42.20 86.25

14 30.61 42.19 20.86 33.73 38.65 42.92 86.79

15 31.34 42.97 21.44 34.61 39.49 43.84 87.82

16 32.08 43.77 21.96 35.44 40.25 44.77 88.54

17 32.93 44.60 22.52 36.20 41.13 45.60 89.33

18 33.62 45.36 23.01 37.05 41.95 46.49 89.98

19 34.22 46.04 23.48 37.83 42.73 47.71 90.58

20 34.74 46.61 23.90 38.28 43.44 48.76 91.18

21 35.31 47.23 24.45 38.82 44.16 49.72 91.78

22 35.71 47.67 24.86 39.32 44.66 50.36 92.08

23 36.14 48.11 25.27 39.78 45.15 51.05 92.50

24 36.64 48.63 25.80 40.10 45.76 51.86 93.04

25 37.07 49.09 26.26 40.61 46.29 52.59 93.44

26 37.45 49.53 26.66 41.09 46.76 53.21 93.83

27 37.82 49.93 26.99 41.34 47.16 54.13 94.26

28 38.28 50.44 27.42 41.63 47.60 54.28 94.70

Figure 21. ALFOTERRA Hydrophobe 4PO Sulfate Biodegradation Study

OECD 301F Theoretical Oxygen demand
12/14/2011 Page 66 of 66

Appendix 22: ALFOTERRA Hydrophobe Biodegradation Study, Comparison of Alcohol, Nonionic, 

and Anionic

Isalchem 

23

Isalchem

23 4PO

Isalchem 23 

4PO S

0 0 0 0

1 10.30 0.97 1.25

2 18.92 3.64 4.08

3 23.13 9.64 14.27

4 27.75 16.92 17.71

5 33.45 22.86 19.48

6 38.46 29.42 20.99

7 43.08 35.81 22.23

8 47.99 41.09 24.20

9 53.21 45.94 25.95

10 57.97 50.34 27.33

11 61.95 54.80 28.39

12 65.29 58.91 29.21

13 68.15 62.74 29.94

14 70.92 66.31 30.61

15 73.54 69.79 31.34

16 76.03 73.03 32.08

17 78.28 75.95 32.93

18 80.58 78.80 33.62

19 82.75 81.52 34.22

20 84.79 84.21 34.74

21 86.78 86.83 35.31

22 88.77 89.30 35.71

23 90.58 92.08 36.14

24 92.25 94.61 36.64

25 93.84 97.12 37.07

26 95.31 99.49 37.45

27 96.74 101.92 37.82

28 98.14 104.22 38.28

Figure 22. ALFOTERRA Hydrophobe 

Biodegradation Study

Appendix 23: Aquatic Toxicity Reports from Environmental Enterprise USA Laboratory TSR No. 

R&D ENV 10-004 01-10

Attached