"test methods for evaluating solid waste, physical/chemical methods, method 3620c: florisil cleanup"

SW-846 is not intended to be an analytical training manual. Therefore, method procedures are written based on the assumption that they will be performed by analysts who areformally trained in at least the basic principles of chemical analysis and in the use of the subjecttechnology. In addition, SW-846 methods, with the exception of required method use for the analysis of method-defined parameters, are intended to be guidance methods which contain generalinformation on how to perform an analytical procedure or technique which a laboratory can useas a basic starting point for generating its own detailed Standard Operating Procedure (SOP),either for its own general use or for a specific project application. The performance dataincluded in this method are for guidance purposes only, and are not intended to be and mustnot be used as absolute QC acceptance criteria for purposes of laboratory accreditation.
Florisil®, a registered trade name of U. S. Silica Co., is a magnesium silicate with basic properties. It is used to separate analytes from interfering compounds prior to sampleanalysis by a chromatographic method. Florisil® has been used for the cleanup of pesticide residues and other chlorinated hydrocarbons; the separation of nitrogen compounds from hydrocarbons; the separation ofaromatic compounds from aliphatic-aromatic mixtures; and similar applications for use with fats,oils, and waxes. Additionally, Florisil® is considered good for separations with steroids, esters,ketones, glycerides, alkaloids, and some carbohydrates.
Florisil® cleanup may be accomplished by either using a glass chromatographic column packed with Florisil® or using solid-phase extraction cartridges containing Florisil®.
This method includes procedures for cleanup of sample extracts containing the Other analytes may potentially be cleaned up using this method provided that adequate Prior to employing this method, analysts are advised to consult the base method for each type of procedure that may be employed in the overall analysis (e.g., Methods 3500,3600, 5000, and 8000) for additional information on quality control procedures, development ofQC acceptance criteria, calculations, and general guidance. Analysts also should consult thedisclaimer statement at the front of the manual and the information in Chapter Two for guidanceon the intended flexibility in the choice of methods, apparatus, materials, reagents, and supplies, and on the responsibilities of the analyst for demonstrating that the techniquesemployed are appropriate for the analytes of interest, in the matrix of interest, and at the levelsof concern. In addition, analysts and data users are advised that, except where explicitly specified in a regulation, the use of SW-846 methods is not mandatory in response to Federal testingrequirements. The information contained in this method is provided by EPA as guidance to beused by the analyst and the regulated community in making judgments necessary to generateresults that meet the data quality objectives for the intended application.
This method is restricted to use by, or under the supervision of, appropriately experienced and trained analysts. Each analyst must demonstrate the ability to generateacceptable results with this method.
This method describes procedures for Florisil® cleanup of solvent extracts of environmental samples. It provides the option of using either traditional columnchromatography techniques or solid-phase extraction cartridges. Generally, the traditionalcolumn chromatography technique uses larger amounts of adsorbent and, therefore, has agreater cleanup capacity.
In the column cleanup protocol, the column is packed with the appropriate amount of adsorbent, topped with a water adsorbent, and then loaded with the sample extract. Elutionof the analytes is effected with a suitable solvent(s), leaving the interfering compounds on thecolumn. The eluate may be further concentrated prior to gas chromatographic analysis.
The cartridge cleanup protocol uses solid-phase extraction cartridges containing 40 µm particles of Florisil® (60 D pores). Each cartridge is washed with solvent immediately prior to use. The sample extract is loaded onto the cartridge which is then eluted with suitablesolvent(s). A vacuum manifold is necessary to obtain reproducible results. The eluate may befurther concentrated prior to gas chromatographic analysis.
Refer to Chapter One and the manufacturer's instructions for definitions that may be Solvents, reagents, glassware, and other sample processing hardware may yield artifacts and/or interferences to sample analysis. All of these materials must be demonstratedto be free from interferences under the conditions of the analysis by analyzing method blanks. Specific selection of reagents and purification of solvents by distillation in all-glass systems maybe necessary. Refer to each method to be used for specific guidance on quality controlprocedures and to Chapter Four for general guidance on the cleaning of glassware.
A reagent blank should be prepared and analyzed for the compounds of interest prior to the use of this method. The level of interferences must meet the QC acceptance criteriafor blanks that are specified in a project planning document, e.g. QAPP, SAP, or in a laboratorySOP.
The procedures for reagent purification outlined here should be considered to be the minimum needed for successful use of this method. More extensive procedures may benecessary to achieve acceptable levels of interferences for some analytes. However, during theevaluation of the cartridge cleanup procedure, phthalate esters were detected in the Florisil®cartridge method blanks at concentrations of up to 400 ng per cartridge. Therefore, completeremoval of the phthalate esters from Florisil® cartridges may not be possible. Phthalate estercontamination may be a problem with certain cartridges The more inert the column and/orcartridge material (i.e., glass or polytetrafluoroethylene (PTFE)), the less problem withphthalates. Phthalates create interference problems for all method analytes, not just thephthalate esters themselves.
This method does not address all safety issues associated with its use. The laboratory is responsible for maintaining a safe work environment and a current awareness file of OSHAregulations regarding the safe handling of the chemicals listed in this method. A reference fileof material safety data sheets (MSDSs) should be available to all personnel involved in theseanalyses.
The mention of trade names or commercial products in this manual is for illustrative purposes only, and does not constitute an EPA endorsement or exclusive recommendation foruse. The products and instrument settings cited in SW-846 methods represent those productsand settings used during method development or subsequently evaluated by the Agency. Glassware, reagents, supplies, equipment, and settings other than those listed in this manualmay be employed provided that method performance appropriate for the intended applicationhas been demonstrated and documented.
This section does not list common laboratory glassware (e.g., beakers and flasks).
Chromatography column -- 300 mm x 10 mm ID, fitted with a polytetrafluoroethylene (PTFE) stopcock.
NOTE: Columns equipped with fritted glass discs are difficult to clean once the column has been used to process highly contaminated extracts. Columns without frits may bepurchased, and a small pad of glass wool may be used to retain the adsorbent. Prewash the glass wool pad with 50 mL of acetone followed by 50 mL of elutionsolvent prior to packing the column with adsorbent.
Muffle furnace -- Capable of maintaining 400 EC.
Vials -- Glass, 10-mL and 25-mL capacity, fitted with polytetrafluoroethylene (PTFE)-lined screw caps or crimp tops.
Vacuum manifold -- VacElute Manifold SPS-24 (Analytichem International), Visiprep (Supelco, Inc.), or equivalent, consisting of glass vacuum basin, collection rack andfunnel, collection vials, replaceable stainless steel delivery tips, built-in vacuum bleed valve andgauge. The system is connected to a vacuum pump or water aspirator through a vacuum trapmade from a 500-mL sidearm flask fitted with a one-hole stopper and glass tubing. Themanifold is necessary for use of the cartridge cleanup protocol.
Top-loading balance -- Capable of weighing to 0.01 g.
Reagent grade chemicals must be used in all tests. Unless otherwise indicated, it is intended that all reagents conform to the specifications of the Committee on AnalyticalReagents of the American Chemical Society, where such specifications are available. Othergrades may be used, provided it is first ascertained that the reagent is of sufficiently high purityto permit its use without lessening the accuracy of the determination. Reagents should bestored in glass to prevent the leaching of contaminants from plastic containers.
Organic-free reagent water -- All references to water in this method refer to organic-free reagent water, as defined in Chapter One.
Granular Florisil® -- For column cleanup procedure. Florisil® is produced in four grades, two of which are appropriate for this procedure. The differences between grades areprimarily a function of the activation temperature, resulting in somewhat different chemicalcharacteristics among the grades. Florisil® Grade PR is activated at 675 EC and is most useful for pesticide residue analyses. Florisil® Grade A is activated at 650 EC and is generally used for other analytes. Whichever grade is used, store Florisil® in glass containers fitted withground-glass stoppers or foil-liner screw caps.
Lauric acid -- Reagent grade. Used for the standardization of the Florisil® activity. Weigh 10.00 g of lauric acid in a 500-mL volumetric flask. Add 50 mL of hexane to the flask todissolve the lauric acid. Swirl the flask gently until the lauric acid is dissolved, then dilute thesolution in the flask to 500 mL with additional hexane.
Phenolphthalein indicator -- Dissolve 1 g of phenolphthalein in ethanol and dilute to Sodium hydroxide, NaOH -- Weigh out 20 g of NaOH (pellets, reagent grade) in a 500-mL volumetric flask. Dissolve in organic-free reagent water and dilute to 500 mL to make a1 N solution. Dilute 25 mL of the 1 N NaOH to 500 mL with water in a second 500-mLvolumetric flask, yielding a 0.05N solution. The NaOH solution should be standardized againstlauric acid, as follows.
Weigh 100 to 200 mg of lauric acid to the nearest 1 mg in a 125-mL Erlenmeyer flask. Add 50 mL of ethanol to the flask and swirl to dissolve the lauric acid. Add 3 drops of phenolphthalein indicator to the flask, and titrate with the 0.05 N NaOH solution to a permanent endpoint (i.e., the indicator color does not disappearwhen the solution is allowed to stand for 1 min). Calculate the "strength" of the NaOH solution as the mg of lauric acid neutralized per mL of NaOH solution.
Deactivation of Florisil® -- for cleanup of phthalate esters. To prepare for use, place 100 ± 10 g of Florisil® into a 500-mL beaker and heat to 140 EC for approximately 16 hrs. After heating, transfer to a 500-mL reagent bottle. Tightly seal andcool to room temperature. When cool, add 3 ± 0.1 mL of organic-free reagent water. Mix thoroughly by shaking or rolling for 10 min and let stand for at least 2 hrs. Keep the bottlesealed tightly. Activation of Florisil® -- for all cleanups other than phthalate esters. It is advisable to treat both Florisil® Grade A and Grade PR prior to use to drive off anymoisture adsorbed during storage and handling. Heat the Florisil® in a glass containerloosely covered with aluminum foil in an oven at 130 EC overnight. Cool the Florisil® in a Florisil® from different batches or sources may vary in adsorptive capacity. To standardize the amount of Florisil® which is used, use the lauric acid value,described below. The procedure determines the adsorption from a hexane solution oflauric acid (mg) per g of Florisil®.
Weigh 2.000 g of Florisil® in a 25-mL, glass-stoppered, Erlenmeyer flask. Cover loosely with aluminum foil and heat overnight at 130 EC. Stopper the flask and cool to room temperature.
Add 20.0 mL of the lauric acid solution to the flask, stopper, Let the Florisil® settle and using a volumetric pipet, transfer 10.0 mL of supernatant liquid into a 125-mL Erlenmeyer flask. Avoid including anyFlorisil®.
Add 60 mL of ethanol and 3 drops of the phenolphthalein Titrate the solution in the flask with the 0.05N NaOH solution until a permanent end point is reached (i.e., the indicator color does not disappearwhen the solution is allowed to stand for 1 min).
The lauric acid value is calculated as follows: Lauric acid value ' 200 & (titration volume in mL of NaOH) (strength of NaOH) where the strength of the NaOH is measured in Sec. 7.6.3 as the mg of lauric acidneutralized per mL of NaOH solution.
Use the following equation to obtain an equivalent quantity of NOTE: This equation was written incorrectly in the previous version of this method.
Sodium sulfate (granular, anhydrous), Na SO -- Purify by heating at 400 EC for 4 hrs in a shallow tray, or by precleaning the sodium sulfate with methylene chloride. A method blank should be analyzed in order to demonstrate that there is no interference from the sodiumsulfate. Florisil® cartridges -- 40 µm particles, 60 D pores. The cartridges from which this method were developed consist of 6-mL serological-grade polypropylene tubes, with the 1 g ofFlorisil® held between two polyethylene or stainless steel frits with 20 µm pores. Cartridgescontaining 0.5 g and 2.0 g of Florisil® are available, however, the compound elution patternsshould be verified when cartridges containing other than 1 g of Florisil® are used. The choice of solvent will depend on the analytes of interest and no single solvent is universally applicable to all analyte groups. Whatever solvent system is employed includingthose specifically listed in this method, the analyst must demonstrate adequate performance forthe analytes of interest, at the levels of interest. At a minimum, such a demonstration willencompass the initial demonstration of proficiency described in Method 3500, using a cleanreference matrix. Method 8000 describes procedures that may be used to develop performancecriteria for such demonstrations as well as for matrix spike and laboratory control sampleresults. All solvents should be pesticide quality or equivalent. Solvents may be degassed prior to Diethyl ether, C H OC H -- Must be free of peroxides as indicated by test strips (EM Quant, or equivalent). Procedures for removal of peroxides are provided withthe test strips. After cleanup, 20 mL of ethyl alcohol preservative must be added to eachliter of ether. Petroleum ether (boiling range of 30 - 60 EC) Florisil® cartridge phenol check solution (for the organochlorine pesticide technique) -- Prepare a solution of 2,4,5-trichlorophenol in acetone at a concentration of 0.1µg/L. See the note in Sec. 7.12.
Florisil® cartridge pesticide check solution -- Prepare a solution containing the NOTE: The concentrations of these analytes were incorrectly listed in units of mg/L in the Chlorophenoxy acid herbicide check solution -- Prepare a solution containing 2,4,5-T methyl ester at 100 µg/L, pentachlorophenyl methyl ester at 50 µg/L, and piclorammethyl ester at 200 µg/L. See the note in Sec. 7.12.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE See the introductory material to Chapter Four, "Organic Analytes" and the specific Refer to Chapter One for additional guidance on quality assurance (QA) and quality control (QC) protocols. When inconsistencies exist between QC guidelines, method-specific QC criteria take precedence over both technique-specific criteria and those criteriagiven in Chapter One, and technique-specific QC criteria take precedence over the criteria inChapter One. Any effort involving the collection of analytical data should include developmentof a structured and systematic planning document, such as a Quality Assurance Project Plan(QAPP) or a Sampling and Analysis Plan (SAP), which translates project objectives andspecifications into directions for those that will implement the project and assess the results. Each laboratory should maintain a formal quality assurance program. The laboratory shouldalso maintain records to document the quality of the data generated. All data sheets andquality control data should be maintained for reference or inspection. Also refer to Method 3600 for cleanup QC procedures, and Method 8000 and the specific determinative method to be used for information on determinative QC procedures.
Initially, before processing any samples, the analyst should demonstrate that all parts of the equipment in contact with the sample and reagents are interference-free. This isaccomplished through the analysis of a method blank. As a continuing check, each timesamples are extracted, cleaned up, and analyzed, and when there is a change in reagents, amethod blank should be prepared and analyzed for the compounds of interest as a safeguardagainst chronic laboratory contamination. The analyst must demonstrate that the compounds of interest are being quantitatively recovered by the cleanup technique before applying this method to actualsamples. This test applies to both the column cleanup and cartridge cleanup procedures. Arecovery check needs to be performed using standards of the target analytes at knownconcentration. This test needs to be conducted on each batch of Florisil® following its The efficiency of each lot of the solid-phase extraction cartridges needs to be verified. Only lots of cartridges from which the spiked analytes are quantitativelyrecovered may be used to process the samples. A check should also be performed at least once on each individual lot of cartridges and at least once for every 300 cartridges ofa particular lot, whichever frequency is greater.
Organochlorine pesticides --To check each new lot of Florisil® cartridges before use, perform the following in duplicate: Combine 0.5 mL of the 2,4,5-trichlorophenol solution in Sec.
7.11, 1.0 mL of the pesticide solution in Sec. 7.12, and 0.5 mL of hexane in a vial. Condition the cartridge as described in Sec. 11.1 and then perform the cartridge cleanup starting with Sec. 11.7.
Elute the cartridge with 9 mL of acetone/hexane (10/90, v/v) only. Reduce the volume to 1.0 mL and analyze by Method 8081.
The lot of Florisil® cartridges is acceptable if all pesticides are recovered at 80 to 110%, if the recovery of trichlorophenol is less than 5%, and ifno peaks interfering with the target analytes are detected. Chlorophenoxy acid herbicides -- To check each new lot of granular Add 5 mL of the chlorophenoxy acid herbicide check solution (Sec. 7.13) to a Florisil® column packed and washed as in Sec. 11.13.2. Elute Fractions 1 and 2 as described in Secs. 11.13.3 and 11.13.4, collecting each in a separate flask. Elute the column with approximately 100 mL of diethyl ether and collect ten separate 10-mL fractions. Concentrate Fraction 1 and Fraction 2 separately and concentrate each of the ten 10-mL diethyl ether fractions to 5 mL.
Analyze each of the 12 eluates by GC/ECD and calculate the recovery of each analyte. Pentachlorophenyl methyl ether should be found inFraction 1. 2,4,5-T methyl ester (and the methyl esters of the other chlorophenoxyacids) should be found in Fraction 2. Determine the volume of diethyl ether that isnecessary to elute picloram methyl ester.
The lot of Florisil® is acceptable if the target analytes are quantitatively recovered and if the recovery of trichlorophenol is less than 5%. Nointerferences should be detected in any of these eluates.
Any method blanks, matrix spike samples, or replicate samples should be subjected to the same analytical procedures (Sec. 11.0) as those used on actual samples. Forsample extracts that are cleaned up using this method, the associated quality control samplesshould also be processed through this cleanup method.
There are no calibration or standardization steps directly associated with this cleanup Sec. 11.1 describes the procedures for assembling and conditioning the Florisil® cartridges. Sec. 11.2 describes general procedures for handling sample extracts prior tocleanup. Secs. 11.3 through 11.13 describe the column and cartridge procedures for phthalateesters (Secs. 11.3 and 11.4); nitrosamines (Sec. 11.5); organochlorine pesticides, haloethers,and organophosphorus pesticides (Secs. 11.6 and 11.7); nitroaromatics and isophorone (Sec.
11.8); chlorinated hydrocarbons (Secs. 11.9 and 11.10); aniline and aniline derivatives (Sec.
11.11); organophosphates (Sec. 11.12); and derivatized chlorophenoxy acid herbicides (Sec.
11.13).
The column chromatography procedures employ a larger amount of Florisil® than the cartridge procedures and, therefore, have a greater cleanup capacity. Samples that exhibitgreater degrees of interferences should be cleaned up using the column procedures. However,both techniques have limitations on the amount of interferences which can be removed.
If the interference is caused by high boiling materials, then Method 3640 should be employed prior to Florisil® cleanup. If the interference is caused by relatively polar compoundsin the same boiling range as the analytes of interest, then multiple column or cartridge cleanupsmay be necessary. For additional cleanup of organochlorine pesticides and PCBs, see Method3665. If crystals of sulfur are present in the extract, then Method 3660 should be employedprior to Florisil® cleanup.
Whenever Florisil® is used to fractionate groups of target compounds (rather than to simply remove potential interferants) it is critical that the specific fractionation scheme bevalidated using spiked solutions or spiked sample extracts that contain most or all of theanalytes of interest. This may be particularly important when the Florisil® cartridge techniquesare employed, because the differences between the various cartridge formats andmanufacturers may affect the fractionation patterns. In addition, it may be useful to archive anyfractions not originally intended for analysis, in the event that the fractionation scheme chosendoes not yield the intended results. Once the determinative analysis has been performed anddemonstrates that the fractionation has been successful, such archived fractions may bedisposed of in an appropriate manner. However, if the fractionation did not perform as intended,the analytes of interest may be contained in the archived fractions which may be analyzed orcombined with the other fraction(s) for reanalysis.
Following Florisil® cleanup, extracts may need further concentration and/or solvent exchange. Consult the appropriate determinative method and 3500 series extraction method fordetails.
Arrange the cartridges on the manifold in the closed-valve position.
Turn on the vacuum pump and set the vacuum to 10 in (254 mm) of Hg. Do not exceed the manufacturer's recommendation for manifold vacuum. Flow rates maybe controlled by opening and closing cartridge valves.
Condition the cartridges by adding 4 mL of hexane to each cartridge. Slowly open the cartridge valves to allow hexane to pass through the sorbent beds to thelower frits. Allow a few drops per cartridge to pass through the manifold to remove all airbubbles. Close the valves and allow the solvent to soak the entire sorbent bed for 5 mins. Do not turn off the vacuum.
Slowly open cartridge valves to allow the hexane to pass through the cartridges. Close the cartridge valves when there is still at least 1 mm of solvent abovethe sorbent bed. Do not allow the cartridges to become dry. If cartridges go dry, repeatthe conditioning step.
Most sample extracts have to be concentrated to a smaller volume prior to the use of Florisil® cleanup. The extract volume is a function of the analytical sensitivity necessary tomeet the project objectives. The extract volume will also affect the ability of the Florisil® toseparate target analytes from potential interferences, particularly for the cartridge procedures,where applying large extract volumes to the cartridges may cause poor results. As noted inSec. 11.0, consult the appropriate extraction and determinative methods for the details on finalextract volumes, extract concentration techniques, and solvent exchange procedures.
Reduce the sample extract volume to 2 mL prior to cleanup for: The extract solvent should be hexane for the phthalate esters, nitroaromatics, chlorinated hydrocarbons, and chlorophenoxy acid herbicides, and methylene chloride forthe nitrosamines and aniline and aniline derivatives.
Reduce the sample extract volume to 10 mL prior to cleanup for: The extract solvent should be hexane for these analytes. In most cases, given the sensitivity of the determinative methods, only 1 mL of the 10 mL extract needs to besubjected to the Florisil® cleanup procedure. The remaining 9 mL should be archived forlater use, if needed.
If the extract was in cold storage, allow it to reach room temperature. Inspect the extract visually to ensure that there are no particulates or phase separationsand that no evaporative loss has taken place. If crystals of sulfur are visible or if thepresence of sulfur is suspected, proceed with Method 3660.
Place approximately 10 g of deactivated Florisil® (Sec. 7.7.1) into a 10 mm ID chromatographic column. Tap the column to settle the Florisil® and addapproximately 1 cm of anhydrous sodium sulfate to the top.
Pre-elute the column with 40 mL of hexane. The rate for all elutions should be about 2 mL/min. Discard the eluate and, just prior to exposure of the sodiumsulfate layer to the air, quantitatively transfer the 2-mL sample extract onto the columnusing an additional 2 mL of hexane to complete the transfer. Just prior to exposure of the sodium sulfate layer to the air, add 40 mL of hexane and continue the elution of the column. Discard this hexane eluate.
Elute the column with 100 mL of ethyl ether/hexane (20/80, v/v) and collect this fraction in a flask (e.g., a 500-mL K-D flask equipped with a clean 10 mLconcentrator tube). Concentrate the eluate to the volume listed in the determinativemethod, using the techniques described in the appropriate 3500 series method. Nosolvent exchange is necessary. Compounds that elute in this fraction are: Using 1-g Florisil® cartridges, condition the cartridges with hexane as Transfer the extract (Sec. 11.2) to the cartridge. Open the cartridge valve to allow the extract to pass through the cartridge bed at approximately 2 mL/min.
When the entire extract has passed through the cartridges, but before the cartridges become dry, rinse the sample vials with an additional 0.5 mL of solvent, andadd the rinse to the cartridges to complete the quantitative transfer.
Close the cartridge valve and turn off the vacuum after the solvent has passed through, ensuring that the cartridge never gets dry.
Place a 5-mL vial or volumetric flask into the sample rack corresponding to the cartridge position. Attach a solvent-rinsed stainless steel solvent guide to themanifold cover and align it with the collection vial.
If the sample is suspected to contain organochlorine pesticides, elute the cartridge with methylene chloride/hexane (20/80, v/v). Turn on the vacuum pump andadjust the pump pressure to 10 in (254 mm) of Hg. Allow the solvent to soak the sorbentbed for 1 min or less. Slowly open the cartridge valve, and collect the eluate (this fractioncontains the organochlorine pesticides, and should be discarded).
Close the cartridge valve, replace collection vials, and add 10 mL of acetone/hexane (10/90, v/v) to the cartridge. Slowly open the cartridge valve and collectthe eluate into the collection vial. This fraction contains the phthalate esters, and shouldbe retained for analysis.
Concentrate the eluate to the volume listed in the determinative method, using the techniques described in the appropriate 3500 series method. Add a weight of activated Florisil® (nominally 22 g) predetermined by calibration (Sec. 7.7.3.7) into a 20 mm ID chromatographic column. Tap the column tosettle the Florisil® and add about 5 mm of anhydrous sodium sulfate to the top.
Pre-elute the column with 40 mL of ethyl ether/pentane (15/85, v/v). Discard the eluate and, just prior to exposure of the sodium sulfate layer to the air,quantitatively transfer the 2-mL sample extract (Sec. 11.2) onto the column using anadditional 2 mL of pentane to complete the transfer.
Just prior to the exposure of the sodium sulfate layer to the air, elute the column with 90 mL of ethyl ether/pentane (15/85, v/v). Discard the eluate. This fractionwill contain any diphenylamine present in the extract.
Elute the column with 100 mL of acetone/ethyl ether (5/95, v/v), collecting the eluate in a flask (e.g., a 500 mL K-D flask equipped with a clean 10 mL concentratortube). This fraction will contain all of the nitrosamines listed in the scope of the method.
Add 15 mL of methanol to the collected fraction, and concentrate this fraction to the volume listed in the determinative method, using the techniques describedin the appropriate 3500 series method. Column procedure for organochlorine pesticides, haloethers, and organophosphorus pesticides (see Table 6 for fractionation patterns of organophosphoruspesticides).
Add a weight of activated Florisil® (nominally 20 g), predetermined by calibration (Sec. 7.7.3.7), to a 20 mm ID chromatographic column. Settle the Florisil® bytapping the column. Add anhydrous sodium sulfate to the top of the Florisil® to form alayer 1 to 2 cm deep.
Pre-elute the column with 60 mL of hexane and discard the eluate. Just prior to exposure of the sodium sulfate to air, quantitatively transfer the 10-mL sampleextract (Sec. 11.2) onto the column, completing the transfer with two 1- to 2-mL rinses withhexane.
Place a flask (e.g., a 500-mL K-D flask equipped with a clean concentrator tube) under the chromatographic column. Drain the column into the flaskuntil the sodium sulfate layer is nearly exposed. Elute the column with 200 mL of ethylether/hexane (6/94, v/v) using a drip rate of about 5 mL/min. This is Fraction 1, and all ofthe haloethers are in this fraction. Remove the flask and set aside for later concentration. Elute the column again, using 200 mL of ethyl ether/hexane (15/85, v/v), into a second flask. This is Fraction 2.
Perform a third elution using 200 mL of diethyl ether/hexane (50/50, v/v), collecting the eluate in a third flask. This is Fraction 3.
Perform a final elution with 200 mL of 100% ethyl ether, collecting the eluate in a fourth flask. This is Fraction 4.
Concentrate the four eluates to the volume listed in the determinative method, using the techniques described in the appropriate 3500 series method. Cartridge procedure for organochlorine pesticides and PCBs Using 1-g Florisil® cartridges, condition the cartridges with hexane, as Transfer the 1 mL (or other appropriate volume) of the extract (Sec. 11.2) to the cartridge. Open the cartridge valve to allow the extract to pass through the cartridgebed at approximately 2 mL/min. When the entire extract has passed through the cartridge, but before the cartridge becomes dry, rinse the sample vial with an additional 0.5 mL of hexane, and addthe rinse to the cartridge to complete the quantitative transfer. Close the cartridge valve and turn off the vacuum after the solvent has passed through, ensuring that the cartridge never goes dry.
Place a 10-mL vial or volumetric flask into the sample rack corresponding to the cartridge position. Attach a solvent-rinsed stainless steel solvent guide to themanifold cover and align with the collection vial.
If there is no need to separate the organochlorine pesticides from the PCBs, then add 9 mL of acetone/hexane (10/90, v/v) to the cartridge. Turn on the vacuumpump and adjust the pump pressure to 10 in (254 mm) of Hg. Allow the solvent to soakthe sorbent bed for 1 min or less. Slowly open the cartridge valve and collect the eluateinto the collection vial. Go directly to Sec. 11.7.8.
The following procedures are used to separate the organochlorine Add 3 mL of hexane to the cartridge. Turn on the vacuum pump and adjust the pump pressure to 10 inches (254 mm) of Hg. Allow thesolvent to soak the sorbent bed for 1 min or less. Slowly open the cartridge valveand collect the eluate into the collection vial. This is Fraction 1 and it will containthe PCBs and a few of the organochlorine pesticides (see Table 5).
Close the cartridge valve, replace the collection vial, and add 5 mL of methylene chloride/hexane (26/74, v/v) to the cartridge. Slowly open thecartridge valve and collect the eluate into the collection vial. This is Fraction 2 andit will contain most of the pesticides.
Close the cartridge valve, replace collection vials, and add 5 mL of acetone/hexane (10/90, v/v) to the cartridge. Slowly open the cartridge valveand collect the eluate into the collection vial. This is Fraction 3 and it will containthe remaining pesticides.
As needed, perform a solvent exchange and adjust the final volume of the eluant to the volume listed in the determinative method, using the techniques described inthe appropriate 3500 series method.
Column procedure for nitroaromatics and isophorone Add a weight of activated Florisil® (nominally 10 g) predetermined by calibration (Sec. 7.7.3.7) into a 10 mm ID chromatographic column. Tap the column tosettle the Florisil® and add about 1 cm of anhydrous sodium sulfate to the top.
Pre-elute the column with methylene chloride/hexane (10/90, v/v) at about 2 mL/min. Discard the eluate and, just prior to exposure of the sodium sulfate layer to theair, quantitatively transfer the 2-mL sample extract (Sec. 11.2) onto the column using anadditional 2 mL of hexane to complete the transfer. Just prior to exposure of the sodium sulfate layer to the air, add 30 mL of methylene chloride/hexane (10/90, v/v) and continue the elution of the column. Discardthe eluate.
Elute the column with 90 mL of ethyl ether/pentane (15/85, v/v) and discard the eluate. This fraction will contain any diphenylamine present in the extract.
Elute the column with 100 mL of acetone/ethyl ether (5/95, v/v), and collect the eluate in a flask (e.g., a 500-mL K-D flask equipped with a 10-mL concentratortube). This fraction will contain all of the nitrosamines listed in the scope of the method.
Add 15 mL of methanol to the collected fraction, and concentrate to the volume listed in the determinative method, using the techniques described in theappropriate 3500 series method. Elute the column with 30 mL of acetone/methylene chloride (10/90, v/v), and collect the eluate in a flask (e.g., a 500-mL K-D flask equipped with a 10-mLconcentrator tube). Concentrate the collected fraction to the volume listed in thedeterminative method, using the techniques described in the appropriate 3500 seriesmethod, and exchanging the solvent to hexane. Compounds that elute in this fraction are: Column procedure for chlorinated hydrocarbons Add a weight of activated Florisil® (nominally 12 g) predetermined by calibration (Sec. 7.7.3.7) into a 10 mm ID chromatographic column. Tap the column tosettle the Florisil® and add about 1 to 2 cm of anhydrous sodium sulfate to the top.
Pre-elute the column with 100 mL of petroleum ether. Discard the eluate and, just prior to exposure of the sodium sulfate layer to the air, quantitatively transfer thesample extract (Sec. 11.2) to the column by decantation and subsequent petroleum etherwashings. Discard the eluate. Just prior to exposure of the sodium sulfate layer to the air, begin eluting the column with 200 mL of petroleum ether and collect the eluate in flask (e.g., a 500-mLK-D flask equipped with a 10-mL concentrator tube). This fraction should contain thefollowing chlorinated hydrocarbons: Concentrate the collected fraction to the volume listed in the determinative method, using the techniques described in the appropriate 3500 seriesmethod. 11.10 Cartridge procedure for chlorinated hydrocarbons 11.10.1 Using 1-g Florisil® cartridges, condition the cartridges with 5 mL of acetone/hexane (10/90, v/v) as described in Sec. 11.1.
11.10.2 Transfer the extract (Sec. 11.2) to the cartridge. Open the cartridge valve to allow the extract to pass through the cartridge bed at approximately 2 mL/min.
11.10.3 When the entire extract has passed through the cartridge, but before the cartridge becomes dry, rinse the sample vial with an additional 0.5 mL of acetone/hexane(10/90), and add the rinse to the cartridge to complete the quantitative transfer.
11.10.4 Close the cartridge valve and turn off the vacuum after the solvent has passed through, ensuring that the cartridge never gets dry.
11.10.5 Place a 5-mL vial or volumetric flask into the sample rack corresponding to the cartridge position. Attach a solvent-rinsed stainless steel solvent guide to themanifold cover and align it with the collection vial.
11.10.6 Add 10 mL of acetone/hexane (10/90, v/v) to the cartridge. Turn on the vacuum pump and adjust the pump pressure to 10 in (254 mm) of Hg. Allow the solvent tosoak the sorbent bed for 1 min or less. Slowly open the cartridge valve and collect theeluate into the collection vial. 11.10.7 Adjust the final volume of the eluant to the volume listed in the determinative method, using the techniques described in the appropriate 3500 seriesmethod.
11.11 Column procedure for aniline and aniline derivatives (see Table 8 for elution 11.11.1 Add a weight of activated Florisil® predetermined by calibration (Sec.
7.7.3.7) into a 20 mm ID chromatographic column. Tap the column to settle the Florisil®.
11.11.2 Pre-elute the column with 100 mL of 2-propanol/methylene chloride (5/95, v/v), followed by 100 mL of hexane/methylene chloride (50/50, v/v), followed by 100 mL ofhexane. Discard the eluate and leave a column of about 5 cm of hexane above theFlorisil®.
11.11.3 Quantitatively transfer the 2-mL sample extract (Sec. 11.2) onto 2.0 g of activated Florisil® in a 50-mL beaker, using a small volume of methylene chloride, and dryunder a gentle stream of nitrogen.
11.11.4 Place the dried Florisil® containing the sample extract onto the chromatographic column, and wash the beaker which contained the Florisil® with 75 mL ofhexane, adding this wash to the reservoir.
11.11.5 Elute the hexane from the column and discard. Stop the column flow just prior to the exposure of the Florisil® to air.
11.11.6 Elute the column with 50 mL of methylene chloride/hexane (50/50, v/v), using a drip rate of about 5 mL/min, and collect the eluate in a flask (e.g., a 500-mL K-Dflask equipped with a 10-mL concentrator tube). This is Fraction 1.
11.11.7 Elute the column with 50 mL of 2-propanol/hexane (5/95, v/v), and collect the eluate in a second flask. This is Fraction 2.
11.11.8 Elute the column a third time using 50 mL of methanol/hexane (5/95, v/v). Collect the eluate in a third flask. This is Fraction 3. Frequently, it will prove useful tocombine the three fractions prior to analysis. However, in some situations, analysis ofeach separate fraction may be necessary. Refer to Method 8131.
11.11.9 Concentrate the collected fractions to the volume listed in the determinative method, using the techniques described in the appropriate 3500 seriesmethod. 11.12 Column procedure for organophosphates 11.12.1 Add a weight of activated Florisil®, predetermined by calibration (Sec.
7.7.3.7), to a 20 mm ID chromatographic column. Settle the Florisil® by tapping thecolumn. Add anhydrous sodium sulfate to the top of the Florisil® to form a layer 1 to 2 cmdeep.
11.12.2 Pre-elute the column with 50-60 mL of hexane. Discard the eluate and just prior to exposure of the sulfate layer to air, quantitatively transfer the 10-mL sampleextract (Sec. 11.2) onto the column using a hexane wash to complete the transfer.
11.12.3 Just as the sample reaches the sodium sulfate, elute the column with 100 mL of diethyl ether/hexane (10/90, v/v). Discard the eluate.
11.12.4 Just prior to exposure of the sodium sulfate to air, elute the column with 200 mL of diethyl ether/hexane (30/70, v/v). This fraction contains all of the targetanalytes except for tris(2,3-dibromopropyl) phosphate.
11.12.5 Elute the column with 200 mL of diethyl ether/hexane (40/60, v/v). This fraction contains tris(2,3-dibromopropyl) phosphate.
11.12.6 Concentrate the collected fraction to the volume listed in the determinative method, using the techniques described in the appropriate 3500 seriesmethod. 11.13 Column procedure for derivatized chlorophenoxy acid herbicides 11.13.1 Add a weight of activated Florisil® (nominally 4 g) predetermined by calibration (Sec. 7.7.3.7) into a 20 mm ID chromatographic column. Tap the column tosettle the Florisil® and add approximately 5 mm of anhydrous sodium sulfate to the top.
11.13.2 Pre-elute the column with 15 mL of hexane. The rate for all elutions should be about 2 mL/min. Discard the eluate, and just prior to exposure of the sodiumsulfate to air, quantitatively transfer the 2-mL sample extract (Sec. 11.2) onto the column,using an additional 2 mL of hexane to complete the transfer.
11.13.3 Just prior to the exposure of the sodium sulfate layer to the air, elute the column with 35 mL of methylene chloride/hexane (20/80, v/v), collecting the eluate in aclean flask (e.g., a 500-mL K-D flask equipped with a concentrator tube). This is Fraction1, and will contain any pentachlorophenyl methyl ester that is present.
11.13.4 Elute the column with 60 mL of methylene chloride/acetonitrile/hexane (50/0.035/49.65, v/v/v), collecting the eluate in a second flask. This is Fraction 2.
11.13.5 If picloram is to be determined, perform a third elution with the volume of diethyl ether determined from the Florisil® check in Sec. 9.4.4, collecting this eluate in athird flask. This is Fraction 3, and will contain the picloram.
11.13.6 The three fractions may be combined for analysis. Concentrate the combined fractions to the volume listed in the determinative method, using the techniquesdescribed in the appropriate 3500 series method. There are no calculations explicitly associated with this cleanup procedure. See the appropriate determinative method for the calculation of final sample results.
Performance data and related information are provided in SW-846 methods only as examples and guidance. The data do not represent required performance criteria for users ofthe methods. Instead, performance criteria should be developed on a project-specific basis,and the laboratory should establish in-house QC performance criteria for the application of thismethod. These performance data are not intended to be and must not be used as absolute QCacceptance criteria for purposes of laboratory accreditation.
Table 1 provides example recoveries of phthalate esters obtained from the Florisil® column procedure. These data are provided for guidance purposes only.
Table 2 provides example recoveries of phthalate esters obtained from the Florisil® cartridge procedure. These data are provided for guidance purposes only.
Table 3 provides an example distribution of organochlorine pesticides and PCBs from the Florisil® column procedure. These data are provided for guidance purposes only.
Table 4 provides example recoveries of Aroclors from the Florisil® cartridge procedure. These data are provided for guidance purposes only.
Table 5 provides an example distribution of organochlorine pesticides from the Florisil® cartridge procedure, using 1-g cartridges. These data are provided for guidancepurposes only.
Table 6 provides an example distribution of organophosphorus pesticides from the Florisil® column procedure. These data are provided for guidance purposes only.
Table 7 provides example recoveries of chlorinated hydrocarbons obtained from the Florisil® cartridge procedure. These data are provided for guidance purposes only.
Table 8 provides example elution patterns for aniline compounds from the Florisil® column procedure. These data are provided for guidance purposes only.
Pollution prevention encompasses any technique that reduces or eliminates the quantity and/or toxicity of waste at the point of generation. Numerous opportunities for pollutionprevention exist in laboratory operations. The EPA has established a preferred hierarchy ofenvironmental management techniques that places pollution prevention as the managementoption of first choice. Whenever feasible, laboratory personnel should use pollution preventiontechniques to address their waste generation. When wastes cannot be feasibly reduced at thesource, the Agency recommends recycling as the next best option.
For information about pollution prevention that may be applicable to laboratories and research institutions consult Less is Better: Laboratory Chemical management for WasteReduction available from the American Chemical Society, Department of Government Relationsand Science Policy, 1155 16th Street, NW, Washington, DC, 20036, http://www.acs.org.
The Environmental Protection Agency requires that laboratory waste management practices be conducted consistent with all applicable rules and regulations. The Agency urgeslaboratories to protect the air, water, and land by minimizing and controlling all releases fromhoods and bench operations, complying with the letter and spirit of any sewer discharge permitsand regulations, and by complying with all solid and hazardous waste regulations, particularlythe hazardous waste identification rules and land disposal restrictions. For further informationon waste management, consult The Waste Management Manual for Laboratory Personnelavailable from the American Chemical Society at the address listed in Sec. 14.2.
A. J. Gordon and R. A. Ford, The Chemist's Companion: A Handbook of Practical Data,Techniques, and References, New York: John Wiley & Sons, Inc., pp. 372, 374, and 375,1972. Floridin of ITT System, Florisil: Properties, Application, Bibliography, Pittsburgh,Pennsylvania, 5M381DW.
P. A. Mills, "Variation of Florisil Activity; Simple Method for Measuring Absorbent Capacityand its use in Standardizing Florisil Columns," Journal of the Association of OfficialAnalytical Chemists, 51, 29, 1968.
U.S. Food and Drug Association, Pesticides Analytical Manual (Volume 1), July 1985.
V. Lopez-Avila, J. Milanes, N. S. Dodhiwala, and W.F. Beckert, "Cleanup ofEnvironmental Sample Extracts Using Florisil Solid-Phase Extraction Cartridges," J.
Chrom. Sci. 27, 209-215, 1989.
U.S. EPA, "Evaluation of Sample Extract Cleanup Using Solid-Phase ExtractionCartridges," Project Report, December 1989.
U.S. EPA, Method 650, "Aniline and Selected Substituted Derivatives." W. F. Beckert and V. Lopez-Avila, "Evaluation of SW-846 Method 8060 for PhthalateEsters," Proceedings of the Fifth Annual Waste Testing and Quality AssuranceSymposium, 1989, pp. 144-156.
U.S. EPA Method 608, Organochlorine Pesticides and PCBs, 40 CFR 136, October 26,1984.
17.0 TABLES, DIAGRAMS, FLOW CHARTS, AND VALIDATION DATA The following pages contain the tables referenced by this method.
EXAMPLE AVERAGE RECOVERIES OF 16 PHTHALATE ESTERS FROM THE a Average recovery from two determinations, data from Reference 8.
EXAMPLE AVERAGE RECOVERIES OF 16 PHTHALATE ESTERS a Average recovery from two determinations, data from Reference 6.
EXAMPLE DISTRIBUTION OF ORGANOCHLORINE PESTICIDES AND PCBs Fraction 1 -- 200 mL of 6% ethyl ether in hexaneFraction 2 -- 200 mL of 15% ethyl ether in hexaneFraction 3 -- 200 mL of 50% ethyl ether in hexane EXAMPLE AVERAGE RECOVERIES OF AROCLORS FROM Florisil® CARTRIDGES EXAMPLE ELUTION PATTERNS AND RECOVERIES OF ORGANOCHLORINE PESTICIDES a1-g Florisil® cartridges spiked with 0.5 µg of each compound.
bThese two compounds coelute on the DB-5 capillary column.
Fraction 1 -- 3 mL of hexaneFraction 2 -- 5 mL of methylene chloride/hexane (26/74, v/v)Fraction 3 -- 5 mL of acetone/hexane (10/90, v/v) EXAMPLE DISTRIBUTION OF ORGANOPHOSPHORUS PESTICIDES Fraction 1 -- 200 mL of 6% ethyl ether in hexaneFraction 2 -- 200 mL of 15% ethyl ether in hexaneFraction 3 -- 200 mL of 50% ethyl ether in hexaneFraction 4 -- 200 mL of 100% ethyl ether = Recovered (no percent recovery data provided by U.S. FDA) EXAMPLE PERCENT RECOVERIES AND ELUTION PATTERNS FOR 22 CHLORINATED HYDROCARBONS FROM 1-g Florisil® CARTRIDGES a aFlorisil® cartridges (Supelco, Inc.) were conditioned with 4 mL of hexane. Five replicateexperiments were performed.
The cartridges were spiked with 1.0 µg per cartridge for hexachloroethane,hexachlorobutadiene, hexachloropentadiene, pentachlorobenzne, and hexachlorobenzene. The trichlorobenzenes, tetrachlorobenzenes, benzal chloride, benzotrichloride, and the BHCswere spiked at 10 µg per cartridge. The dichlorobenzenes and benzyl chloride were spiked at100 µg per cartridge, and 2-chloronaphthalene was spiked at 200 µg per cartridge.
The cartridges were eluted with 5 mL of acetone/hexane (10/90, v/v).
EXAMPLE DISTRIBUTION OF ANILINES IN Florisil® CLEANUP FRACTIONS Fraction 1 -- 50% methylene chloride in hexaneFraction 2 -- 5% 2-propanol in hexaneFraction 3 -- 5% methanol in hexane

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