Total Sulfide, Methylene Blue Method 1. Scope and Application

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Alpha Analytical, Inc. ID No.: 2221 Facility: Westborough Revision 3 Department: Wet Chemistry Published Date:11/2/2012 7:58:09 AM Title: Total Sulfide, Methylene Blue Method Page 1 of 10

Total Sulfide, Methylene Blue Method

References: EPA 376.2: Methods for the Chemical Analysis of Water and Wastes, EPA 600/ 4-82-055, 1982. SM 4500S2-AD: Standard Methods for the Examination of Water and Wastewater, APHA-AWWA-WPCF, 21 st Edition, 2000. EPA 9030B: SW-846, Test Methods for Evaluating Solid Waste: Physical/Chemical Methods, EPA SW-846, Update III, 1997.

1. Scope and Application Matrices: This method is applicable to water, wastewater and seawater samples. Definitions: Refer to Alpha Analytical Quality Manual. Sulfide often is present in groundwater, especially in hot springs. Its common presence in wastewaters comes partly from the decomposition of organic matter, sometimes from industrial wastes, but mostly from the bacterial reduction of sulfate. Hydrogen sulfide escaping into the air from sulfide-containing wastewater causes odor nuisances. The threshold odor concentration of H 2S in clean water is between 0.025 and 0.25µg/L. H 2S is very toxic and has claimed the lives of numerous workers in sewers. It attacks metals directly and has indirectly caused serious corrosion of concrete sewers because it is oxidized biologically to H 2SO 4 on the pipe wall. From an analytical standpoint, three categories of sulfide in water and wastewater are distinguished:

♦ Total sulfide includes dissolved H 2S and HS-, as well as acid-soluble metallic sulfides present in suspended matter. The S -2 is negligible, amounting to less than 0.5% of the dissolved sulfide at pH 12, less than 0.05% at pH 11, etc. Copper and silver sulfides are so insoluble that they do not respond in ordinary sulfide determinations; they can be ignored for practical purposes. ♦ Dissolved sulfide is that remaining after suspended solids have been removed by flocculation and settling. Un-ionized hydrogen sulfide may be calculated from the concentration of dissolved sulfide, the sample pH, and the practical ionization constant of H 2S. The data report packages present the documentation of any method modification related to the samples tested. Depending upon the nature of the modification and the extent of intended use, the laboratory may be required to demonstrate that the modifications will produce equivalent results for the matrix. Approval of all method modifications is by one or more of the following laboratory personnel before performing the modification: Area Supervisor, Department Supervisor, Laboratory Director, or Quality Assurance Officer. This method is restricted to use by or under the supervision of trained analysts. Each analyst must demonstrate the ability to generate acceptable results with this method by performing an initial demonstration of capability, analyzing a proficiency test sample and completing the record of training. After initial demonstration, ongoing demonstration is based on acceptable laboratory performance of at least a quarterly laboratory control sample or acceptable performance from an annual proficiency test sample. A major modification to this procedure requires demonstration of performance. The identification of major method modification requiring performance demonstration is directed by the Quality Assurance Officer and/or Laboratory Director on a case-by-case basis.

Printouts of this document may be out of date and should be considered uncontrolled. To accomplish work, the published version of the document should be viewed online. Document Type: SOP-Technical Pre-Qualtrax Document ID: SOP 07-30 Issue 2 Rev 1 Alpha Analytical, Inc. ID No.: 2221 Facility: Westborough Revision 3 Department: Wet Chemistry Published Date:11/2/2012 7:58:09 AM Title: Total Sulfide, Methylene Blue Method Page 2 of 10

2. Summary of Method Sulfide is separated from the sample matrix by suspending the sample in a hydrochloric acid solution and vigorous agitation. Hydrogen sulfide gas is released from the sample and collected in a gas scrubbing bottle cotaining sodium hydroxide. The methylene blue method is based on the reaction of sulfide, mild oxidizing agent potassium dichromate, and dimethyl-p- phenylenediamine to produce methylene blue. 2.1 Method Modifications from Reference None. 3. Reporting Limits The reported detection limit is 0.1 mg/L. 4. Interferences 4.1 Strong reducing agents interfere in the methylene blue test by preventing formation of the blue color. Thiosulfate at concentrations about 10mg/L may retard color formation or completely prevent it. Sulfide itself prevents the reaction if its concentration is very high, in the range of several hundred milligrams per liter. To avoid the possibility of false negative results, use the antimony method to obtain a qualitative result in industrial wastes likely to contain sulfide but showing no color by the methylene blue method. Iodide, which is likely to be present in oil-field wastewaters, may diminish color formation if its concentration exceeds 2mg/L. Ferrocyanide produces a blue color.

4.2 Eliminate interferences due to sulfite, thiosulfate, iodide, and many other soluble substances (but not ferrocyanide), by precipitating ZnS.

4.3 Interference Removal or Sample Concentration: Put 0.15mL (3 drops) zinc acetate solution into a 100mL glass bottle, fill with sample, and add 0.10mL (2 drops) 6N NaOH solution. Stopper, making sure no air bubbles are present and mix by rotating back and forth vigorously about a transverse axis. Add enough NaOH to produce a pH above 9. Let precipitate settle for 30 minutes. The treated sample is relatively stable and can be held for several hours. However, if much iron is present, oxidation may be fairly rapid.

5. Health and Safety The toxicity or carcinogenicity of each reagent and standard used in this method is not fully established; however, each chemical compound should be treated as a potential health hazard. From this viewpoint, exposure to these chemicals must be reduced to the lowest possible level by whatever means available. A reference file of material safety data sheets is available to all personnel involved in the chemical analysis. Additional references to laboratory safety are available in the Chemical Hygiene Plan. All personnel handling environmental samples known to contain or to have been in contact with municipal waste must follow safety practices for handling known disease causative agents.

6. Sample Collection, Preservation, Shipping and Handling 6.1 Sample Collection Take samples with minimum aeration in either a glass or plastic bottle. Fill the bottle completely and cover. Aqueous samples are collected in a 250mL plastic bottle, soil/solid samples are collected in an amber glass container. 6.2 Sample Preservation All samples must be preserved with 2N zinc acetate (Section 8.8) and 6N sodium hydroxide (Section 8.9). Use four drops of 2N zinc acetate solution per 100mL aqueous sample. Adjust the pH to greater than 9.0 with 6N NaOH. 6.3 Sample Shipping No specific requirements. 6.4 Sample Handling Samples are stored refrigerated at 4 ± 2° C. Soil samples are analyzed within a 28-day holding time. Aqueous samples are analyzed within a 7-day holding time. 7. Equipment and Supplies 7.1 Spectrophotometer, Genesys 10vis: For use at a wavelength of 664nm with cells providing light paths of 1cm.

7.2 Test Tubes: 25mL capacity

7.3 Pipets: Glass, volumetric, various volumes

7.4 Volumetric Flasks: 100mL and 1L volumes

7.5 Erlenmeyer Flasks: 125mL volume

7.6 Three-neck Flask: 500mL volume, 24/40 standard taper joints.

7.7 Dropping funnel: 100mL volume, 24/40 joint.

7.8 Purge gas inlet tube: 24/40 joint with coarse frit.

7.9 Purge gas outlet: 24/40 joint reduced to ¼ in. tube.

7.10 Gas scrubbing bottles: 125mL volume, with ¼ inch o.d. inlet and outlet tubes.

7.11 Tubing: ¼ inch o.d. polytetrafluoroethylene (PTFE) or polypropylene.

7.12 Magnetic Stirrer

7.13 Nitrogen gas: 5.0 Ultra pure.

7.14 Nitrogen regulator

7.15 Flowmeter

7.16 Teflon-coated stirring bar

7.17 Analytical Balance: Capable of weighing 0.1g.

7.18 Glass fiber filter disks: 90mm

7.19 Filtration Apparatus: With membrane filter funnel.

7.20 Suction flask

7.21 Vacuum pump

8. Reagents and Standard 8.1 2 different sources. Stock Sulfide Solution, approximately 0.05M: Weigh 12 grams of Na 2S · 9H 2O (Section 8.13) and add to a 1L volumetric flask. Bring to volume with DI water. Mix by swirling gently to prevent oxygen entrapment. Transfer into a 1L amber glass container and store at room temperature. This solution reacts slowly with oxygen but the change is unimportant within a few hours. 8.1.1 Standardization against thiosulfate solution Immediately after preparation, standardize the Stock Sulfide Solution as follows. Also standardize the Stock Solution if the % recovery falls below 75%. 8.1.1.1 Pipet 10mL of 0.025N standard iodine solution (Section 8.5) into each of two 125mL Erlenmeyer flasks. 8.1.1.2 Pipet 2mL of sulfide stock standard solution (Section 8.1) into one flask. Pipet 2mL of DI water, as a reagent blank, into the other flask. 8.1.1.3 Add 5mL of 6M HCl (Section 8.12) into each flask, swirl slightly, then cover and place in the dark for 5 minutes. 8.1.1.4 Titrate each with 0.025N thiosulfate (Section 8.4) until the yellow iodine color fades to a pale straw color. Just before all the iodine has been titrated, add starch indicator (Section 8.6) dropwise to form a pale blue color. Continue the titration with the thiosulfate. The end point is reached when the blue color first disappears.

8.1.1.5 Calculate the sulfide concentration as follows:

2- 2- Sulfide (µmol/mL) = ( T blank - T sample ) X N S 2O3 1 mole S 1000 µmoles 2- V sample 2 equiv S 1 mmole Where: T = volume of titrant used for the blank and sample (mL) 2- N = concentration of S 2O3 titrant V = volume of sample used (mL), 2mL recommended Calculation: 1.1045 g Na2S. 9H2O in 140-ml DI= 1097mg/l. 8.2 Sulfide 1 Reagent: Available commercially from HACH Company. Store at 4 ± 2°. Reagent expires upon manufacturer’s specified date.

8.3 Sulfide 2 Reagent: Available commercially from HACH Company. Store at 4 ± 2°. Reagent expires upon manufacturer’s specified date.

8.4 Sodium Thiosulfate, 0.025N: Purchased commercially prepared and standardized. Store at room temperature, away from light. Expires upon manufacturer’s specified date.

8.5 Standard Iodine Solution, approximately 0.025N: Dissolve 25g of potassium iodide (KI) (Section 8.14) in 700mL of DI water in a 1L volumetric flask. Add 3.2g of iodine (Section 8.15). Allow to dissolve. Dilute to 1L with DI water and standarize exactly 20mL of the iodine solution . Titrate with 0.025N sodium thiosulfate (Section 8.4) until the amber floor fades. Add starch indicator solution (Section 8.6) until the solution turns a deep blue. Continue titration drop by drip until the blue color disappears. Prepare solution fresh as needed.

Standard Iodine Solution, 0.025 N; Purchased commercially.

8.5.1 LCS/ICV/CCV Solution: The sulfide stock soln. (Section 8.1) is diluted with 0.2N NaOH soln.to a concentration at about the mid-point of the calibration curve.

8.6 Starch indicator solution: Purchased commercially prepared. Store at room temperature. There is no expiration date.

8.7 Sulfide Calibration Solutions: These solutions are used to generate the calibration curve in Section 10.2. Prepare a series of sulfide solutions by diluting the Stock Sulfide Standard second source(Section 8.1). These solutions are prepared fresh prior to generating a calibration curve. Into seven 100mL volumetric flasks, pipet 0mL, 0.5mL, 1mL, 2.0mL, 4mL, 6mL and 8mL of Stock Sulfide Solution (Section 8.1). Bring each to volume with 0.2N NAOH Solution.

8.8 Zinc Acetate Solution: Dissolve 220g Zn(C 2H3O2)2 · 2H 2O in 500mL water. This solution is used as the preservative during sampling. Store at room temperature. Solution is stable.

8.9 Sodium Hydroxide Solution, NaOH, 1N and 6N: For 1N NaOH, dissolve 40g NaOH in 1L of DI water. For 6N, dissolve 240g NaOH in 1L of DI water. This solution is used as a preservative during sampling. Store at room temperature. Solution is stable.

8.10 Sodium Hydroxide Solution, NaOH, 0.2N: In a 100mL volumetric flask, pipet 20mL of 1N NaOH (Section 8.8). Bring to volume with DI water. Store at room temperature. Solution is stable.

8.11 HCL Solution, 2N: Add 175mL concentrated HCL (Section 8.16) to 500mL of DI water. Dilute to 1L with DI water. Store at room temperature.

8.12 HCL (50%): Carefully add 250mL of concentrated HCL (Section 8.16) to 250mL DI water. Store at room temperature.

8.13 Na 2S · 9H 2O: Store at room temperature, out of direct light. Expires one year from date of opening.

8.14 Potassium Iodide (KI): Store at room temperature, away from light. No expiration date.

8.15 Iodine: Store at room temperature, away from light. No expiration date.

8.16 Concentrated HCL: Store at room temperature.

9. Quality Control The laboratory must maintain records to document the quality of data that is generated. Ongoing data quality checks are compared with established performance criteria to determine if the results of analyses meet the performance characteristics of the method. 9.1 Blanks 9.1.1 Method Blank: Distill and analyze one Method Blank of DI water per batch of 20 samples or less. The Method Blank results must be less than the Reporting Limit. Refer to Section 12 for any corrective actions. 9.1.2 ICB/CCB: The ICB/CCB is not distilled. It consists of 0.2N NaOH soln and is analyzed following each ICV/CCV. Results must be less than the Reporting Limit. Refer to Section 12 for any corrective actions. 9.2 Laboratory Control Sample (LCS) Distill and analyze one LCS (Section 8.5.1) per batch of 20 samples or less. % Recovery must be within 75 – 125%. Refer to Section 12 for any corrective actions. 9.3 Initial Calibration Verification (ICV) The ICV is not distilled. It consists of the ICV Standard (Section 8.5.1) and is analyzed after every 10 samples in the batch. % Recovery must be within 75 – 125%. Refer to Section 12 for any corrective actions. 9.4 Continuing Calibration Verification (CCV) The CCV is not distilled. It consists of the CCV Standard (Section 8.5.1) and is analyzed after every 10 samples in the batch. % Recovery must be within 75 – 125%. Refer to Section 12 for any corrective actions. 9.5 Matrix Spike Prepare and analyze one matrix spike per batch of 20 samples or less.

9.6 Laboratory Duplicate Prepare and analyze one sample in duplicate per batch of 20 samples or less. %RPD must be < 25%. Refer to Section 12 for any corrective actions. 9.7 Method-specific Quality Control Samples None. 9.8 Control Limits The laboratory maintains performance records to document the quality of data that is generated. Method accuracy for samples is assessed and records maintained. After the analysis of 20 spiked samples, and 20 laboratory control samples, calculate the average percent recovery (R) and the standard deviation of the percent recovery (S). Control limits for the method parameters are generated. The control limits are calculated based on in-house performance data. The limits are compared to the control limits found in the reference method. 9.9 Method Sequence • Standardize the Stock Sulfide Solution. • Generate a calibration curve. • Add 1mL Sulfide 1 Reagent to 25mL of sample and swirl gently to mix. • Add 1mL of Sulfide 2 Reagent. • Wait 5 – 15 minutes. • Read the absorbances of samples and QC samples. • Calculate results. 10. Procedure 10.1 Initial Calibration Prepare a calibration curve using the selected standards from Section 8.7. Analyze the standards as in Section 10.3 below, and plot concentration vs. absorbance. Determine the slope and the y-intercept. The correlation coefficient must be > 0.995.

10.2 Equipment Operation and Sample Processing 10.2.1 Sample Preparation 10.2.1.1 Soil samples: Weigh 10 grams of the sample on parafilm and place into the reaction flask with a clean stirbar. Record the weight in the laboratory notebook. 10.2.1.2 Aqueous samples: Filter entire sample volume through a 90mm glass fiber filter and rinse with DI water. Place filter into the reaction flask with a clean Teflon stirbar. Record the volume of sample filtered in the laboratory notebook. 10.2.1.3 Add 200mL of deaerated 2N HCL Solution (Section 8.11) into the thistle tube, making sure that the stopcock is closed. 10.2.1.4 Add 20mL of deaerated 1N NaOH (Section 8.9) and 50mL of deaerated DI to the 250mL collection vessel. 10.2.1.5 Assemble the distillation train.

10.2.1.6 Connect the thistle flask to the center position of the reaction flask 10.2.1.7 Connect the collection vessel to the left side of the reaction flask, and the impinger tube to the collection vessel. The impinger tube must be below the surface of the NaOH solution. Make sure that all ground glass joints are sealed. 10.2.1.8 Connect the Nitrogen gas to the right side of the collection vessel. The gas inlet must be above the surface of the reaction flask solution (about two inches). 10.2.1.9 Making sure that the train is a closed system, purge the train with Nitrogen for 5 minutes. Open the valve on the Nitrogen tank (Section 7.13) and open up the regulator so that 45 psi of Nitrogen is flowing (this will effectively give a flow rate of about 150mL/min). 10.2.1.10 Once the system has been purged for 5 minutes, open the stopcock fully on the thistle tube and release the 2N HCL solution into the reaction vessel. At this point the sample should be stirring. Close the stopcock as soon as all of the 2N HCL solution has been delivered to the reaction flask. Let the reaction progress for 1 hour. 10.2.1.11 After one hour has passed, dissemble the train and bring the total volume of the collection vessel to 100mL with deaerated DI . 10.2.2 Color Development: Transfer 25mL sample to a test tube using a special wide-tip pipet. Add 1.0mL Sulfide 1 reagent (Section 8.2) and swirl to mix. Add 1mL of Sulfide 2 reagent (Section 8.3). Let the reaction continue for 5 minutes. Excessive mixing causes low results by loss of H 2S as a gas before it has had time to react. The presence of sulfide will be indicated by the appearance of blue color. Color development usually is complete in about 1 minute, but a longer time is often required for fading out of the initial pink color. Wait 5 to 15 minutes and make color comparisons. If zinc acetate was used, wait at least 10 minutes before making a visual color comparison. 10.2.3 Absorbance Determination: Set the spectrophotometer to a wavelength of 664nm. Zero the instrument with a portion of the blank sample using a 1 cm cell. Read the absorbances of the samples and QC samples and record this information in the laboratory notebook. If the absorbance is greater than the highest standard in the calibration curve, the sample must be diluted with 0.2N NaOH (Section 8.10) and reanalyzed as in Section 10.3.1 above. Record in the laboratory notebook any dilutions that are prepared. 10.3 Continuing Calibration See Section 9.4

10.4 Preventive Maintenance An instrument service company calibrates the spectrophotometers on a semi-annual basis. Certificates are kept on file.

11. Calculations Calculate sulfide concentration using the calibration curve generated in Section 10.2:

Sulfide (mg/L) = Absorbance – y intercept X Dilution factor

Slope

12. Contingencies for Handling Out-of-Control Data or Unacceptable Data Holding time exceedence and improper preservation are noted on the nonconformance report form. Perform routine preventative maintenance following manufacturer's specification. Record all maintenance in the instrument logbook. Review of standards, blanks and standard response for acceptable performance occurs for each batch of samples. Record any trends or unusual performance on a nonconformance action form. If the CV or LCS recovery of any parameter falls outside the designated acceptance range, the laboratory performance for that parameter is judged to be out of control, and the problem must be immediately identified and corrected. The analytical result for that parameter in the unspiked samples is suspect and is only reported for regulatory compliance purposes with the appropriate nonconformance action form. Immediate corrective action includes reanalyzing all affected samples by using any retained sample before the expiration of the holding time.

If the Method Blank result is greater than the reporting limit, and all associated sample results are non-detect, or greater than 10x the concentration of the Method Blank, the data may be reported and a narrative submitted for inclusion on the final report. If associated sample results are positive, but at a concentration less than 10x the concentration of the Method Blank, those samples are redigested and reanalyzed.

If the LCS fails acceptance criteria, the entire batch must be redigested and reanalyzed. If there is insufficient sample volume to redigest, then the Client Services Department is notified and the sample results are reported as estimated, with a narrative to be included on the final report.

If %RPD for the Duplicate fails acceptance criteria, the non-conformance is narrated and included on the final report.

13. Method Performance 13.1 Method Detection Limit Study (MDL) / Limit of Detection Study (LOD) / Limit of Quantitation (LOQ) The laboratory follows the procedure to determine the MDL, LOD, and/or LOQ as outlined in Alpha SOP/08-05. These studies performed by the laboratory are maintained on file for review. 13.2 Demonstration of Capability Studies Refer to Alpha SOP/08-12 for further information regarding IDC/DOC Generation. 13.2.1 Initial (IDC) The analyst must make an initial, one-time, demonstration of the ability to generate acceptable accuracy and precision with this method, prior to the processing of any samples. 13.2.2 Continuing (DOC) The analyst must make a continuing, annual, demonstration of the ability to generate acceptable accuracy and precision with this method.

14. Pollution Prevention and Waste Management Refer to Alpha’s Chemical Hygiene Plan and Waste Management and Disposal SOP for further pollution prevention and waste management information.

15. Referenced Documents Chemical Hygiene Plan SOP/08-05 MDL/LOD/LOQ Generation SOP/08-12 IDC/DOC Generation SOP/14-01 Waste Management and Disposal SOP

16. Attachments None.