1、Designation: D 3650 93 (Reapproved 2006)Standard Test Method forComparison of Waterborne Petroleum Oils ByFluorescence Analysis1This standard is issued under the fixed designation D 3650; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revis
2、ion, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon (e) indicates an editorial change since the last revision or reapproval.1. Scope*1.1 This test method covers the comparison of waterbornepetroleum oils with oils from possible sources
3、by means offluorescence spectroscopy (1).2Useful references for this testmethod include: (2) and (3) for fluorescence analysis in generaland (4), (5), and (6) for oil spill identification includingfluorescence.1.2 This test method is applicable to crude or refinedpetroleum products, for any sample o
4、f neat oil, waterborne oil,or sample of oil-soaked material. Unless the samples arecollected soon after the spill occurs, it is not recommended thatvolatile fuels such as gasoline, kerosine, and No. 1 fuel oils beanalyzed by this test method, because their fluorescencesignatures change rapidly with
5、weathering. Some No. 2 fueloils and light crude oils may only be identifiable up to 2 daysweathering, or less, depending on the severity of weathering. Ingeneral, samples weathered up to 1 week may be identified,although longer periods of weathering may be tolerated forheavy residual oils, oil weath
6、ered under Arctic conditions, oroil that has been protected from weathering by collecting in athick layer.1.3 This standard does not purport to address all of thesafety concerns, if any, associated with its use. It is theresponsibility of the user of this standard to establish appro-priate safety an
7、d health practices and determine the applica-bility of regulatory limitations prior to use.2. Referenced Documents2.1 ASTM Standards:3D 1129 Terminology Relating to WaterD 1193 Specification for Reagent WaterD 3325 Practice for Preservation of Waterborne OilSamplesD 3326 Practice for Preparation of
8、Samples for Identifica-tion of Waterborne OilsD 3415 Practice for Identification of Waterborne OilsD 4489 Practices for Sampling of Waterborne OilsE 131 Terminology Relating to Molecular SpectroscopyE 275 Practice for Describing and Measuring Performanceof Ultraviolet, Visible, and Near-Infrared Spe
9、ctrophotom-etersE 520 Practice for Describing Photomultiplier Detectors inEmission and Absorption Spectrometry3. Terminology3.1 DefinitionsFor definitions of terms used in this testmethod refer to Terminology D 1129, Practice D 3415, andTerminology E 131.4. Summary of Test Method4.1 This test method
10、 consists of fluorescence analyses ofdilute solutions of oil in spectroquality cyclohexane. In mostcases the emission spectra, with excitation at 254 nm, over thespectral range from 280 to 500 nm, are adequate for matching.4.2 Identification of the sample is made by direct visualcomparison of the sa
11、mples spectrum with the spectra frompossible source samples.NOTE 1When weathering has occurred, it may be necessary toconsider known weathering trends when matching spectra (Fig. 1 and Fig.2).5. Significance and Use5.1 This test method is useful for rapid identification ofwaterborne petroleum oil sa
12、mples as well as oil samplesobtained from fuel or storage tanks, or from sand, vegetation,or other substrates. This test method is applicable to weatheredand unweathered neat oil samples.5.2 The unknown oil is identified through the comparison ofthe fluorescence spectrum of the oil with the spectra
13、(obtainedat similar instrumental settings on the same instrument) of1This test method is under the jurisdiction of ASTM Committee D19 on Waterand is the direct responsibility of Subcommittee D19.06 on Methods forAnalysis forOrganic Substances in Water.Current edition approved Feb. 15, 2006. Publishe
14、d February 2006. Originallyapproved in 1978. Last previous edition approved in 1999 as D3650 93 (1999).2The boldface numbers in parentheses refer to the references at the end of thistest method.3For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Customer Service at s
15、erviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.1*A Summary of Changes section appears at the end of this standard.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959,
16、United States.possible source samples. A match of the entire spectrumbetween the unknown and possible source sample indicates acommon source.6. Interferences6.1 The fluorescence spectrum will be distorted if an oilsample has been contaminated by an appreciable amount, forexample, 1 % of common chemi
17、cal impurities such as otheroils that are fluorescent on excitation at 254 nm.NOTE 2Storage of samples in improper containers (for example,plastics) may result in contamination. This interference can be eliminatedby observing proper procedures for collection and preservation ofsamples. Refer to Prac
18、tice D 3325.NOTE 3“Spectroquality” cyclohexane may not have a low enoughfluorescence solvent blank. Lots vary in the content of fluorescentimpurities, which may increase with storage time even if the bottle isunopened.6.2 Oil residues may build up in fluorescence cells particu-larly after prolonged
19、usage with heavy oils. In such a case,follow the procedure using nitric acid for cleaning glassware(10.1.3).6.3 Possible interferences from Raman or RayleighTyndallscattering are not observed in the emission scan rangesselected.7. Apparatus7.1 Fluorescence Spectrophotometer (or Spectro-fluorometer)A
20、n instrument recording in the spectral range of220 nm to at least 600 nm for both excitation and emissionresponses and capable of meeting the specifications stated inTable 1.7.2 Excitation SourceA high-pressure xenon lamp (a150-W xenon lamp has proven acceptable). Other continuumsources, such as deu
21、terium or high-pressure xenon-mercury,which have sufficient intensity in the ultraviolet region, couldbe used as excitation sources.NOTE 4Line sources such as a low-pressure mercury lamp may alsobe used for excitation at 254 nm, if the flexibility of using arbitraryexcitation wavelengths or excitati
22、on spectra is not desired and if sourceintensity is adequate.7.3 Fluorescence CellsStandard cells, made fromfluorescence-free fused silica with a pathlength of 10 mm anda height of 45 mm.7.4 Recorder or ComputerStrip chart or X-Y recorder,with a response time less than 1 s for full-scale deflection,
23、 or acomputer capable of digitizing the data at a rate of 1 data pointper nanometre.7.5 Cell-Filling DeviceDisposable Pasteur capillary pi-pet.7.6 Volumetric FlasksLow-actinic glass, ground-glassstoppered volumetric flasks (100-mL).7.7 Micropipet, 10 to 50-L capacity.7.8 Analytical Balance, with a p
24、recision of at least 60.1 mg.7.9 Weighing Pans, 5 to 7-mm diameter, 18 mm deep, madeof aluminum or equivalent.7.10 Test Tubes, disposable 15-mL glass test tubes.7.11 Micropipet, or microsyringe, 9-L capacity; with anaccuracy of 1 % and reproducibility of 0.1 % of pipet capacity.7.12 Micropipet, 200-
25、L capacity with disposable tips; withan accuracy of 1 % and reproducibility of 0.1 % of pipettorcapacity.FIG. 1 Fluorescence Spectra for a Typical No. 2 Fuel Oil(Unweathered and Weathered One Day)FIG. 2 Fluorescence Spectra for a Typical No. 6 Fuel Oil(Unweathered and Weathered One Day)TABLE 1 Speci
26、fications for Fluorescence SpectrophotometersWavelength ReproducibilityExcitation monochromator better than6 2nmEmission monochromator better than 62nmGratings (Typical Values)Excitation monochromator minimum of 600 lines/mm blazed at300 nmAEmission monochromator minimum of 600 lines/mm blazed at300
27、 nm or 500 nmAPhotomultiplier TubeBEither S-20Cor S-5DResponseEResolutionExcitation monochromator better than 2 nmEmission monchromator better than 2 nmTime Constantnot to exceed one secondAOr designed to have a good efficiency in this spectral region.BSee Practice E 520.CPhotomultiplier tubes such
28、as Hamamatsu R-446-UR.DPhotomultiplier tubes such as RCA 1P28 or Hamamatsu R-106.EOr equivalent having a good spectral response in the spectral region from 280to 600 nm.D 3650 93 (2006)27.13 Solvent Dispenser, adjustable to deliver 10 mL.7.14 Vortex Mixer.8. Reagents and Materials8.1 Purity of Reage
29、ntsSpectroquality grade reagentsshould be used in all instances unless otherwise stated. It isintended that all reagents shall conform to the specifications ofthe Committee onAnalytical Reagents of theAmerican Chemi-cal Society, where such specifications are available.48.2 Purity of Water References
30、 to water shall be under-stood to mean Type IV reagent water conforming to Specifi-cation D 1193. However, since fluorescent organic impuritiesin the water may constitute an interference, the purity of thewater should be checked by running a water blank using thesame instrument conditions as for the
31、 solvent blank.8.3 Acetone (CH3COCH3).8.4 Nitric Acid (sp gr 1.42)Concentrated nitric acid(HNO3).8.5 Cyclohexane, spectroquality grade, with a fluorescencesolvent blank less than 2 % of the intensity of the major peakof the sample fluorescence generated with the same instrumen-tal settings over the
32、emission range used. Cyclohexane isdispensed throughout the procedure from a 500-mL TFE-fluorocarbon wash bottle. For prolonged storage, cyclohexaneshould be stored only in glass. Check the suitability of thesolvent by running a solvent blank. The solvent blank can alsobe used to check for scatter.N
33、OTE 5Cyclohexane can be reused, if necessary, after one or moredistillations in an all-glass still. The distilled cyclohexane must have nodetectable fluorescence (2 %) in the 280 to 500-nm region of thespectrum when excited at 254 nm.NOTE 6Methylcyclohexane can also be used as a solvent, instead ofc
34、yclohexane. This is useful, particularly if the solution is needed forlow-temperature luminescence measurements as well.8.6 Aluminum Foil.9. Sampling and Sample Preparation9.1 Collect a representative sample as directed in PracticeD 4489.9.2 Preserve samples in containers as specified in PracticeD 3
35、325. However, to avoid dewaxing, do not cool samplesbelow 5C.9.3 Preparation of Oil Samples, as described in PracticesD 3326. Avoid the use of deasphalting procedures, if possible.Spectroquality cyclohexane is the preferred solvent for samplepreparation for fluorescence.9.4 Preparation of Solutions
36、for Fluorescence AnalysisEither of the following techniques for diluting the prepared oilsample with cyclohexane may be used:9.4.1 Weighing TechniqueTo prepare oil solutions at aconcentration of approximately 20 g/mL, weigh out 0.0016 60.0001 g of oil (equivalent weight for each sample) onto aclean
37、aluminum weighing pan using a micropipet. Transferweighed oil sample into a clean 100 mL, low-actinic glassvolumetric flask by creasing the aluminum pan and washingthe oil directly into the volumetric flask using spectroqualitycyclohexane dispensed from a TFE-fluorocarbon wash bottle.Dilute the solu
38、tion up to volume (100 mL) and shake vigor-ously several times and allow the prepared solution to stand for30 min and shake again prior to performing the analysis toensure that all oil dissolves. Occasionally, depending onfluorescence yield of the oil tested and instrumentation used, itmay be necess
39、ary to use 100 ppm concentration to get adequatefluorescence intensity. In these cases, weigh out 0.0078 60.0001 g of oil and proceed as above.NOTE 7It is preferable that the prepared solution be used the sameday. Do not use solutions that have been standing for periods in excess of6 h unless they h
40、ave been refrigerated. In no case use solutions more than2 days old.9.4.2 Volume TechniqueAllow the prepared oil sample tocome to room temperature and shake until they are homoge-neous. Transfer 9 L of the oil to a 15-mL disposable glass testtube with a micropipet or microsyringe and add 10 mL ofspe
41、ctroquality cyclohexane with a solvent dispenser. Place acap of aluminum foil over the top of the test tube and vortex forapproximately 30 s. With a micropipet, transfer 200 L of thissolution to a second 15-mL test tube and then add 10 mL ofcyclohexane. Place a cap of aluminum foil over the top of t
42、hesecond test tube and vortex for approximately 30 s. Prepare allsamples in this manner.NOTE 8If a micropipet with disposable plunger and tips is used,potential cross contamination is avoided. Otherwise, careful cleaningfollowing the procedures specified in 10.1 is required.10. Preparation of Appara
43、tus10.1 Cleaning Glassware:10.1.1 Clean all glassware used in this procedure in thefollowing manner: first rinse volumetric flasks and cells threetimes with spectroquality cyclohexane. Prior to the use ofglassware and cells throughout this procedure, rinse again withspectroquality cyclohexane.10.1.2
44、 If there is water present, rinse the glassware threetimes with spectroquality acetone, and then three times withcyclohexane as in 10.1.1. Use detergents only if they have beenchecked for low fluorescence. If laboratory detergent solutionsare used, repeated rinsing with Type IV reagent (see 8.3) wat
45、erwill be required.10.1.3 When working with heavy oils, a cleaning procedureusing organic solvents may not be sufficient. Heavy oils buildup a residue on cells that solvent cleaning will not remove. Ifthe solvent blank shows significant impurities, a residual filmon the cell, rather than an impure s
46、olvent, may be the cause.Soak the cells in undiluted nitric acid for 1 h. Observe propersafety precautions by using adequate eye and hand protection.Rinse the cells repeatedly with Type IV reagent water, and thenproceed as in 10.1.2.10.2 Calibration of Spectrophotometer:10.2.1 Adjust and calibrate t
47、he spectrophotometer (that is,the emission and excitation monochromators) using a low-pressure mercury lamp (or similar line source). Refer toPractice E 275 for the approved calibration method.4“Reagent Chemicals, American Chemical Society Specifications,” Am. Chemi-cal Soc., Washington, DC. For sug
48、gestions on the testing of reagents not listed bythe American Chemical Society, see “Analar Standards for Laboratory Chemicals,”BDH Ltd., Poole, Dorset, U.K., and the “United States Pharmacopeia.”D 3650 93 (2006)311. Procedure for Recording Fluorescence EmissionSpectrum11.1 Fill a clean fluorescence
49、 cell with oil solution using aPasteur pipet or similar techniques, taking care not to contami-nate the outside of the cell. Gently wipe the outside of the cellwith lens paper (nonsilicone-treated) and place the cell in thesample compartment.11.2 Set the excitation monochromator at 254 nm.11.3 Set the slit width of the excitation monochromatorbetween 5 and 35 nm depending on the intensity required. -nmslit width is recommended. Set the emission slit width to amaximum of 2.5 nm and a minimum of 1 nm.NOTE 9Commercial instruments have a wide variation in
