Problem Statement:

The ETP plant of ASK chemicals stops getting disturbed at intervals of time. This disturbance is caused because the factors facilitating the growth of microbial culture in the ETP plant are disturbed. This results in not dropping of COD (Chemical Oxygen Demand) of the effluent as per norms.

Objectives:

  1. Finding out and rectifying the factors causing disturbance of ETP
  2. Finding out the Nutrient Requirement and its level for effective functioning of microbial culture in ETP
  3. To drop the COD level of ETP as per norms
  4. Setting up SOP for culture maintenance in the ETP plant

Actionable Item:

Discussion on understanding the Project- 26 Feb 2022:

To understand the problem statement of the ETP project and the work required to be done a meeting was held with Dr Arun Dixt Sir, Dr Anuradha Jape Ma’am, & Sonal Shinde Ma’am. Following are the bullet points that were discussed/decided in the meeting:

  • The biomass left after the elute treatment is taken to the dumping yard without its retreatment and reuse. To treat biomass and find its further applications, its biological and chemical analysis is needed to be done.
  • By chemical analysis we can find out N, P, K, Ca, Mg, Fe, Zn, Cu, Mn, Ni, Co, Mo, etc. content.
  • Also, Total Biomass dry Weight has to be determined by doing its centrifugation & taking filtrate.
  • Identification of bacteria and fungi contributing to the effective functioning of the ETP plant needs to be done. For this identification step, we can approach a nearby lab.
  • Doing BOD and COD analysis to know the efficiency of the ETP plant.
  • Design of Experiment (DOE) has to be done to standardize the media Composition and chemical composition for running the ETP plant effectively.

Discussion on Design of Experiment (DOE)- 17 March 2022:

To resolve the problem of disturbance in ETP functioning, DOE was decided in a meeting with Dr Arun Dixit Sir and Dr Anuradha Jape Ma’am. Following are the bullet points that were discussed and decided in the meeting:

  • Batch setup can be done in a bucket or small IBC tank.
  • On evaporation of water from culture can be made up by adding water free from any salts or additional nutrients.
  • Levels of C, N, and P will not be changed.
  • The ratio of C: N:P: K in ETP is 100:10:1:1 respectively.
  • There is no heavy requirement for Phosphorus, a trace amount of 1m mg/L is sufficient.
  • Mn, Mg, and S will not be added as it is likely to get precipitated.
  • Ca not be added as it is already available in the ETP.
  • In 1st trial, only standard trace elements should be added to the reactor viz. are Fe, Cu, Mo, Zn, Co.
  • Following are the nutrients and their concentration level to be added:
Iron0.3 mg/L
Copper0.03 mg/L
Molybdenumo.5 mg/L
Zinc0.8 mg/L
Cobalt3 mg/L
  • BOD & COD analysis should be done on the 1st day, 3rd day, and day when the COD reduction stops.
  • Find out the COD, BOD ratio to know the biodegradability.

Listing of requirements required for the Experiment Setup (29 March 2022):

Following is the list of chemicals, glassware, and other materials required for the experiment:

Sr. No.Chemical / GlasswareQuantity
1Potassium dichromate (K2Cr2O7).500 gm
2Mercuric sulphate (HgSO4)100 gm
3Sulfuric acid (H2SO4)5 Liter
4Ferrous ammonium sulphate (FAS)500 gm
5Ferron indicator100 ml
6Potassium hydrogen phosphate (KHP)100 gm
7Sodium hydroxide500 gm
8Sodium sulfite500 gm
9Acetic acid 500 ml
10Potassium iodide500 gm
11Starch indicator250 ml
12Potassium dihydrogen phosphate (KH2PO4)500 gm
13Dipotassium Hydrogen Phosphate (K2HPO4)500 gm
14Disodium hydrogen phosphate                                 ( Na4HPO4.7H2O)500 gm
15Ammonium chloride (NH4Cl)500 gm
16Sodium iodide250 gm
17Calcium Carbonate500 gm
18Magnesium Sulphate500 gm
19Ferric Chloride500 gm
20Manganous sulphate (MnSO4.H2O)500 gm
21Sodium thiosulphate500 gm
22BOD Bottles10
23Erlenmeyer Flask (1000 ml)2
24Erlenmeyer Flask (500 ml)2
25Erlenmeyer Flask (250 ml)6
26Burette1
27Reflux condenser6
28COD tubes6
29Glass Measuring Cylinder (100 ml)1
30Hand Gloves1 Box
31Buckets5
32Motor5
33Air Bubbles5
34Ferrous sulphate100 gm
35Cupric Sulphate100 gm
36 Zinc suphate100 gm
37Molybdenum Trioxide100 gm
38Cobalt Sulphate100 gm
39NH4NO3100 gm

Experiment Setup 1 at ASK Chemicals (1 April 2022):

As per discussions on Project in a meeting, we arranged a setup of the experiment at ASK Chemicals. Following is a detailed procedure of the Experiment to know the nutrient requirement of the Microbial culture.

A) Multiplication of Microbial Culture (30 March 2022) :

  1. 5 Liters of tap water were taken and boiled for 30 min and allowed to cool again.
  2. On the cooling of water, add 50 g Jaggery and 0,5 g Urea to it.
  3. Inoculate 250 ml microbial mother culture to it. This culture is used for experiments later.

B) Experiment Setup at ASK Chemicals (1 April 2022):

  1. 5 Buckets were taken.
  2. Addition in 5 buckets is done as follows:
Bucket 1Bucket 2Bucket 3Bucket 4Bucket 5
ETP Effluent5 L5 L1 L1 L1 L
Tap Water10 L10 L14 L14 L14 L
Fe0.3 mg/L0.3 mg/L
Cu0.06 mg/L0.06 mg/L
Zn0.8 mg/L0.8 mg/L
Mix Nutrients14 g14 g
Soil 1 kg
Microbial culture1 L1 L1 L1 L1 L

Note:

  1. In Bucket No. 1 & Bucket No. 2; 5 Lit of effluent and 10 L of tap water were added. This dilution was done to get the effluent COD 5000 mg/L.
  2. In Bucket No. 3, Bucket No,4 & Bucket No. 5; 5 Lit of effluent and 10 L of tap water was added. This dilution was done to get the effluent COD 1000 mg/L.
  3. As a source of Fe, Cu, and Zn; Ferrous sulphate, Cupric Sulphate, and Zinc suphate salts were taken respectively.
  4. By calculation: Ferrous sulphate- 0.045 gm, Cupric Sulphate- 0.005 gm, and Zinc suphate- 0.15 gm was weighed (to get 0.3mg/L of Fe, 0.03 mg/L of Cu, and 0.8 mg/L of Zn) and taken in a beaker. Made up volume up to 100ml using distilled water. 50 ml of this solution was added to Bucket No.1 and another 50ml was added to Bucket No. 3.
  5. After the addition of all components given in the above table, aeration was provided in the bucket using aero tubes.
  6. COD (Chemical Oxygen Demand) analysis was done after every 24 hours.

COD Readings:

COD analysis was done after every 24 hours. Following are the readings of COD of effluent in all buckets:

Bucket No.1/4/20222/4/20223/4/20224/4/20225/4/20226/4/2022
15 lit Diluted to 15+ Micro Nutrients
3828
39043864351330063279
25 lit Diluted to 15+ Mix Nutrients
3368

3864

3825

3201
28493006
31 lit Diluted to 15 + Micro Nutrients
918
9181140132711321249
41 lit Diluted to 15 + Mix Nutrients
918
128811671171702858
51 lit Diluted to 15 + Soil
918
124910961015624741
% of COD Decrease From the previous day
1.029.086.668.33
1.0116.316.095.22
14.09-16.405.889.37
9.39-0.3426.7318.18
12.257.3927.0015.79
% of COD Decrease From 2 April
1.0210.0216.0123.00
1.0117.1622.2026.27
14.090.005.8814.69
9.399.0833.3945.50
12.2518.7340.6750.04

Observations:

  1. 16% COD reduction was observed in Bucket No. 1
  2. 22% COD reduction was observed in Bucket No. 2
  3. 6% COD reduction was observed in Bucket No. 3
  4. 33% COD reduction was observed in Bucket No. 4
  5. 40% COD reduction was observed in Bucket No. 5

Understanding and Learning outcome from Experiment Setup No.1:

  • COD reduction was observed more in Bucket no. 5. This could be because of:- a) Microbes which we introduced through microbial culture have got nutrients from the soil that contributed to the growth of microbes that also fed on effluent in bucket resulting in a decrease in COD; b) Microbes from soil got introduced in the system from soil which fed on effluent in bucket resulting in a decrease in COD; c) The content in the effluent may have got adsorbed on soil resulting in COD reduction.
  • COD in Bucket No. 1, 2, 3, and 4 had not reduced remarkably as compared to Bucket No. 5. This could be because we just gave micronutrients to the microbes and failed to provide an NPK source. Also in Bucket No. 1 & Bucket No. 3, micronutrients like Mo and Co were not added (Mo and Co were not added because it was not available in the lab during the experiment setup); because of this enzyme activity of microbes may not be seen at the optimum level.
  • Hence, considering the above mistakes and learning outcomes we decided to set up a 2nd Experiment trial.

B) Experiment Setup 2 at ASK Chemicals (9 April 2022):

  1. 5 Buckets were taken.
  2. Addition in 5 buckets is done as follows:
Bucket 1Bucket 2Bucket 3Bucket 4Bucket 5
ETP Effluent5 L5 L1 L1 L1 L
Tap Water10 L10 L14 L14 L14 L
Fe0.3 mg/L0.3 mg/L
Cu0.06 mg/L0.06 mg/L
Zn0.8 mg/L0.8 mg/L
Mix Nutrients14 g14 g
KH2PO40.5 gm/L0.5 gm/L0.5 gm/L0.5 gm/L
K2HPO40.5 gm/L0.5 gm/L0.5 gm/L0.5 gm/L
NH4NO30.1 gm/L0.1 gm/L0.1 gm/L0.1 gm/L
Soil 1 kg
Microbial culture750 ml750 ml750 ml750 ml750 ml

Note:

  1. In Bucket No. 1 & Bucket No. 2; 5 Lit of effluent and 10 L of tap water were added. This dilution was done to get the effluent COD 5000 mg/L.
  2. In Bucket No. 3, Bucket No,4 & Bucket No. 5; 5 Lit of effluent and 10 L of tap water was added. This dilution was done to get the effluent COD 1000 mg/L.
  3. As a source of Nitrogen, NH4NO3 was added at 0.1 gm/L (1.5 gm for 15 Liters Bucket) in Buckets No. 1, 2, 3, and 4.
  4. As a source of Potassium and Phosphorous, KH2PO4 and K2HPO4 were added at 0.5 gm/L each (7.5 gm for 15 Liters Bucket) in Buckets No. 1, 2, 3, and 4.
  5. As a source of Fe, Cu, and Zn; Ferrous sulphate, Cupric Sulphate, and Zinc suphate salts were taken respectively.
  6. By calculation: Ferrous sulphate- 0.045 gm, Cupric Sulphate- 0.005 gm, and Zinc suphate- 0.15 gm was weighed (to get 0.3mg/L of Fe, 0.03 mg/L of Cu, and 0.8 mg/L of Zn) and taken in a beaker. Made up volume up to 100ml using distilled water. 50 ml of this solution was added to Bucket No.1 and another 50ml was added to Bucket No. 3.
  7. 0.75 g Urea was added as a Nitrogen source to Buckets 1, 2, 3 and 4.
  8. After the addition of all components given in the above table, aeration was provided in the bucket using aero tubes.
  9. COD (Chemical Oxygen Demand) analysis was done after every 24 hours.

COD Readings:

COD analysis was done after every 24 hours. Following are the readings of COD of effluent in all buckets:

Bucket No.10/4/202211/4/202212/4/202213/4/202214/4/202215/4/202216/4/202219/4/202220/4/202221/4/202222/4/2022
15 lit Diluted to 15+ Micro Nutrients + NPK
6910
6026503755444782568455404664453041323817
25 lit Diluted to 15+ Mix Nutrients + NPK
7027

6531

6136
61365448529250995213519650774526
31 lit Diluted to 15 + Micro Nutrients + NPK
3474
3382330432522273207720201646163012591102
41 lit Diluted to 15 + Mix Nutrients + NPK
4099
4043408239483880388037703449329628732715
51 lit Diluted to 15 + Soil
967
505583493274392320274236196118

Per day COD Reduction Rate:

In 5 days of experiment setup 2 COD reduction was observed as follows:

Bucket No.12/4/202213/4/202214/4/202215/4/202216/4/202219/4/202220/4/202221/4/202222/4/2022
118.42 %19.77%30.8 %17.74 %19.83 %32.50 %34.44 %40.20 %44.76 %
28.71 %12.68 %22.47 %24.69 %27.44 %25.81 %26.06 %27.75 %35.59 %
34.89 %6.39 %34.57 %40.21 %41.85 %52.62 %53.08 %63.76 %68.28 %
40.41 %3.68 %5.34 %5.34 %8.03 %15.86 %19.59 %29.91 %33.76 %
54.27 %49.49 %71.93 %59.64 %67.21 %71.93 %75.82 %66.38 %79.76 %

Understanding and Learning outcome from Experiment Setup No.2:

  • Buckets No. 1 & 3 are showing a remarkable reduction in COD of ETP plant effluent. This bucket contained N, P, K, Cu, Fe, & Zn.
  • Bucket No. 5 showed the highest rate of COD reduction. But, it can be due to- 1) Content in effluent got adsorbed on the soil, 2) Microorganisms from soil got introduced into the bucket that fed on the organic and inorganic matter in effluent resulting in the dropping of COD.

Note:

  • Bucket No. 5 having soil showed a remarkable result in dropping COD of the effluent of the ETP plant; soil cannot be used as a solution to drop COD at the Industrial level as the amount of soil required is huge and getting the same quality of soil is not possible.
  • But, we can isolate the microbes responsible for the reduction of ETP effluent COD using appropriate media and prepare an effective microbial consortium for ETP effluent treatment.
  • In Bucket No. 1 & 3, micronutrients like Co and Mo were not added because of their unavailability. Therefore, we had made a setup of Bucket No. 2 & 4 where we used mixed nutrients. But now Co and Mo micronutrients are available. So, in Experiment 3 setup we can add all the micronutrients decided in the ‘Design of Experiment’ viz. were Cu, Co, Mo, Fe, & Zn along with N, P and K.
  • When calculations were reviewed again for the addition of Cupric Sulphate, Iron Sulphate and Zinc Sulphate as a source of Cu, Fe and Zn it was understood that the calculation for Cu and Zn went wrong. According to the correction, Copper sulphate and Zinc sulphate had to be added at 0.0036 gm and 0.108 gm respectively for 30 litres.
  • As all the micronutrients as per decided in DOE of the experiment are available, no need to set an experiment setup of mixed nutrients separately.

Discussion on Design of Experiment (DOE)- 22 April 2022:

Results of ETP experiment setup 2 were recorded, studied and shared with Jape maám. The aphonic meeting was scheduled with Jape Maám to discuss the results, understanding and learning outcomes, and the next steps to be taken in the ETP project. The points raised through the discussion and Experiment Setup 3 were designed and decided to be done at ASK Chemicals as follows:

  • Macronutrients and Micronutrients Levels: Levels of NH4NO3, K2HPO4, KH2PO4, Cupric Sulphate, Zinc Sulphate, and Iron Sulphate as a source of N, P, K, Cu, Zn, Fe respectively should not be changed and should be added just like added in Experiment Setup no.2. Also, Micronutrients Molybdenum Trioxide and Cobalt sulphate should be added as a source of Mo and Co in next Experiment Setup No.3 that was left to be added in previous experiment setup no.2; it should be added such that we should get Mo 0.5 mg/L and Co 3 mg/L. Hence, the addition of Macronutrients and Micronutrients should be as follows:
Sr.No.NutrientsComposition (for 15 Liters )
1.NH4NO31.5 gm (0.1 gm/L)
2.KH2PO47.5 gm (0.5 gm /L)
3.K2HPO47.5 gm (0.5 gm /L)
4.Cupric Sulphate0.0025 gm
5.Zinc sulphate0.075 gm
6.Ferrous Sulphate0.0225 gm
7.Molybdenum Trioxide0.1136 gm
8.Cobalt Sulphate0.215 gm
  • The soil that was used in Bucket No. 5 should be taken and reused for effluent treatment. From this, we can understand whether the COD of effluent in Bucket No.5 was dropped due to adsorption or not.
  • The treated effluent of Bucket No.5 can be used as an inoculum and study its effect on the COD of effluent. From this, we can understand if there is any effect of Microbes that were introduced from the soil used in Experiment Setup 2 in dropping the COD of effluent.
  • BOD analysis should be done immediately on the day of Experiment Setup 3 arranged and on the day when COD drop is stabilized. ( Procedure followed for BOD: http://vadic.vigyanashram.blog/2022/04/13/experiment-setup-of-biological-oxygen-demand-bod/)

C) Experiment Set 3 at ASK Chemicals (03/05/2022):

As per understanding from the experiments setup 1 & 2 done at ASK Chemicals, and discussions held with Dr Arun Dxit Sir and Dr Jape Maám; Experiment Setup 3 was designed as follows:


Bucket No. 1Bucket No.2Bucket No.3Bucket No. 4Bucket No. 5Bucket No.6
Nutrients+ NPK+ Culture (5000 COD)Nutrients+ NPK+ Culture (1000 COD) Mix Nutrients+ NPK+ Culture (1000 COD)Nutrients+ NPK+ Effluent from Bucket No.5, Exp No.2 (1000 COD)Nutrients+ NPK+ Culture + Soil from Bucket No.5, Exp No.2 (1000 COD)NPK+ Culture + New Soil (1000 COD)
ETP Effluent5 L1 L1 L1 L1 L1 L
Tap Water10 L14 L14 L14 L14 L14 L
KH2PO47.5 g7.5 g7.5 g7.5 g7.5 g7.5 g
K2HPO47.5 g7.5 g7.5 g7.5 g7.5 g7.5 g
Urea0.75 g0.75 g0.75 g0.75 g0.75 g0.75 g
Ferrous Sulphate0.0225 g0.0225 g0.0225 g0.0225 g
Cupric Sulphate0.0025 g0.0025 g0.0025 g0.0025 g
Zinc Sulphate0.075 g0.075 g0.075 g0.075 g
Molybdenum Trioxide0.1136 g0.1136 g0.1136 g0.1136 g
Cobalt Sulphate0.215 g0.215 g0.215 g0.215 g
Mix micronutrients14 g
Culture750 ml750 ml750 ml750 ml750 ml
The effluent of Bucket No.5 in Exp Setup No.2 (as inoculum)750 ml
The soil from Bucket No.5 in Exp Setup No.2Complete Soil
Soil 1 Kg

COD Readings:

Following are the COD readings day wise:

Bucket No.6/5/20227/5/20228/5/20229/5/2022
1Nutrients+ NPK+ Culture (5000 COD)3825370837083669
2Nutrients+ NPK+ Culture (1000 COD)741585507429
3Mix Nutrients+ NPK+ Culture (1000 COD)1991175616391561
4Nutrients+ NPK+ Effluent from Bucket 5, Exp No. 2 (1000 COD)468312273195
5Nutrients+ NPK+ Culture+ Soil from Bucket 5, Exp No. 2 (1000 COD)1522124912101054
6NPK+ Culture+ New Soil (1000 COD)741546507390