METHOD FOR SEPARATING AND QUANTIFYING SATURATED AND UNSATURATED DIALKYL KETONES

Abstract

To provide a method for separating and quantifying saturated and unsaturated dialkyl ketones. (1) A method for separating and quantifying saturated and unsaturated dialkyl ketones, the method including quantifying a dialkyl ketone-containing sample by a gas chromatograph using a polar column, except a nonpolar column and a slightly polar column, the polar column having a polarity value of 440 or greater. (2) The method according to (1), wherein the polar column is a polar column with medium polarity or higher polarity, the polar column having a polarity value of 700 or greater. (3) The method of (2), wherein the polar column is a polar column with high polarity or higher polarity, the polar column having a polarity value of 1550 or greater. (4) The method according to (3), wherein a liquid phase of the polar column with high polarity or higher polarity is an ionic liquid column. (5) A method for producing a chemical transesterified oil and/or fat, including separating and quantitatively analyzing saturated and unsaturated dialkyl ketones in an oil and/or fat, using the method described in any one of (1) to (4).

Claims

1. A method for separating and quantifying saturated and unsaturated dialkyl ketones, the method comprising quantifying a dialkyl ketone-containing sample by a gas chromatograph using a polar column, except a nonpolar column and a slightly polar column, the polar column having a polarity value of 440 or greater.

2. The method according to claim 1, wherein the polar column is a polar column with medium polarity or higher polarity, which has a polarity value of 700 or greater.

3. The method according to claim 2, wherein the polar column is a polar column with high polarity or higher polarity, which has a polarity value of 1550 or greater.

4. The method according to claim 3, wherein a liquid phase of the polar column with high polarity or higher polarity is an ionic liquid column.

5. A method for producing a chemical transesterified oil and/or fat, comprising separating and quantitatively analyzing saturated and unsaturated dialkyl ketones in an oil and/or fat, using the method described in claim 1.

Description

DESCRIPTION OF EMBODIMENTS

[0033] The present invention is directed to a method for separating and quantifying saturated and unsaturated dialkyl ketones, including quantifying a dialkyl ketone-containing sample by a gas chromatograph using a polar column, except a nonpolar column and a slightly polar column, the polar column having a polarity value of 440 or greater. The present invention will be described in detail below.

Dialkyl Ketone-Containing Sample

[0034] In the method of the present invention, various samples such as oils and/or fats used in foods or food raw materials or extracts of waterproof paper in contact with foods can be used as the dialkyl ketone-containing sample.

[0035] Among these samples, the method can be preferably used in analysis of edible oils and/or fats, particularly chemical transesterified oils and/or fats containing saturated and unsaturated dialkyl ketones.

[0036] It is also possible to extract and purify hydrophobic fractions from foods using chemical transesterified oils and/or fats or the like, for example, water-in-oil emulsions such as confectioneries, chocolates and margarines, oil-in-water emulsions such as fillings, whip creams and dressings, and beverages by an ordinary method, and analyze them.

Polar Column Except Nonpolar Column and Slightly Polar Column

[0037] The method of the present invention is characterized by using a polar column, except a nonpolar column and a slightly polar column, having a polarity value of 440 or greater, when quantifying a dialkyl ketone-containing sample by a gas chromatograph. The use of a polar column having a polarity value of 440 or greater enables separation and quantification of saturated and unsaturated dialkyl ketones.

[0038] Examples of the nonpolar column and the slightly polar column include columns in which the liquid phase is 100% dimethylpolysiloxane, 5% diphenyl-dimethylpolysilphenylenesiloxane, 5% diphenyl-95% dimethylpolysiloxane, and 14% diphenyl-86% dimethylpolysiloxane, and specific examples thereof can include DB-1, HP-1, Rtx-1, CP-Sil5CB, SPB-1, InertCap1 MS, InertCap1HP-5 ms, Rxi-5 ms, Equity-5, Rtx-5 MS, DB-5 ms, Rxi-5SilMS, VF-5 ms, SLB-5 ms, InertCap5 MS/Sil, InertCap5 MS/NP, InertCap5, DB-5, HP-5, Rtx-5, CP-Si18CB, SPB-5, and DB-XLB.

Polarity Value

[0039] In the present invention, the polarity of the column is medium polarity or higher polarity, and the polar column is characterized by having a polarity value of 440 or greater, and preferably has a polarity value of preferably 700 or greater, and more preferably 1550 or greater. The polarity value is a value indicating the total sum of McReynolds numbers of five kinds of compounds (benzene, n-butanol, 2-pentanone, nitropropane, and pyridine) in the column. The McReynolds number represents a difference between a value of Kovats Index on a squalane stationary phase column and a value of Kovats Index in a liquid phase, for a compound.

[0040] Examples of the medium polar column having a polarity value of from about 440 to about 1550 include a column in which the liquid phase is 20% diphenyl-80% dimethylpolysiloxane, a column in which the liquid phase is 35% diphenyl-65% dimethylpolysiloxane, a column in which the liquid phase is 50% diphenyl-50% dimethylpolysiloxane, a column in which the liquid phase is 6% cyanopropylphenyl-94% dimethylsiloxane, a column in which the liquid phase is 14% cyanopropylphenyl-86% dimethylsiloxane, a column in which the liquid phase is 50% diphenyl-50% dimethylsilphenylenesiloxane, and a column in which the liquid phase is 50% trifluoropropyl-50% methylpolysiloxane, and specific examples thereof can include InertCap35 MS, InertCap25, DB-35msUI, VF-35 ms, Rxi-35silMS, InertCap624, DB-624, HP-VOC, Rtx-624, VF-624 ms, InertCap624 MS, DB-624, HP-VOC, Rtx-624, Rxi-624SilMS, VF-624 ms, InertCap1301, DB-1301, HP-1301, Rtx-1301, VF-1301 ms, InertCap25, DB-35 ms, DB-35, HP-35 ms, HP-35, Rtx-35, VF-35 ms, InertCap35, VF-1701 ms, InertCap1701 MS, InertCap1701, DB-1701, Rtx-1701, VF-1701 ms, SPB-1701, InertCap17 MS/Sil, DB-17 MS, VF-17 ms, Rxi-17silMS, InertCap17 MS, DB-17 ms, Rxi-17SilMS, VF-17 ms, InertCap17, DB-17, HP-50, Rtx-50, CP-Sil24CB, SPB-50, InertCap210, DB-210, Rtx-200, and VF-200 ms.

[0041] Among them, examples of the medium polar column having a polarity value of 700 or greater include a column in which the liquid phase is 14% cyanopropylphenyl-86% dimethylsiloxane, a column in which the liquid phase is 35% diphenyl-65% dimethylpolysiloxane, a column in which the liquid phase is 50% diphenyl-50% dimethylpolysiloxane, a column in which the liquid phase is 50% diphenyl-50% dimethylsilphenylenesiloxane, and a column in which the liquid phase is 50% trifluoropropyl-50% methylpolysiloxane.

[0042] Further, in the method of the present invention, the polarity of the column is most preferably high polarity or higher polarity, and the polarity value is preferably 1550 or greater.

[0043] Examples of the highly polar column having a polarity value of 1550 or greater include a column in which the liquid phase is polyethylene glycol, a column in which the liquid phase is nitroterephthalic acid-modified polyethylene glycol, a column in which the liquid phase is 50% cyanopropylmethyl-50% phenyl-methylpolysiloxane, a column in which the liquid phase is 50% cyanopropylphenyl-50% dimethylsiloxane, a column in which the liquid phase is nitrile silicone, a column in which the liquid phase is 70% Cyanopropyl-Silphenylene siloxane, a column in which the liquid phase is poly(biscyanopropyl siloxane, proprietary phase, poly (80% biscyanopropyl/20% cyanopropylphenyl siloxane) phase, a column in which the liquid phase is 1,12-Di(tripropylphosphonium)dodecane bis(trifluoromethylsulfonyl)imide, a column in which the liquid phase is 1,12-Di(tripropylphosphonium)dodecane bis(trifluoromethylsulfonyl)imide, a column in which the liquid phase is 1,12-Di(tripropylphosphonium)dodecane bis(trifluoromethylsulfonyl)imide trifluoromethylsulfonate, a column in which the liquid phase is Tri(tripropylphosphoniumhexanamido)triethylamine bis(trifluoromethylsulfonyl)imide, a column in which the liquid phase is 1,12-Di(2,3-dimethylimidazolium)dodecane bis(trifluoromethylsulfonyl)imide, a column in which the liquid phase is 1,9-Di(3-vinylimidazolium)nonane bis(trifluoromethylsulfonyl)imide, and a column in which the liquid phase is 1,5-Di(2,3-dimethylimidazolium)pentane bis(trifluoromethylsulfonyl)imide, and specific examples thereof can include InertCap Pure-WAX, DB-WAX, HP-INNOWax, Rtx-Wax, Stabilwax, InertCap WAX-HT, DB-WAXetr, SolGel-WAX, InertCap FFAP, DB-FFAP, HP-FFAP, CP-WAX 58(FFAP)CB, HR-SS-10, PAG, SPB-225, InertCap 225, DB-225, Rtx-225, CP-Sil 43CB, SP-2330, TC-70, SP-2330, SP-2331, SP-2560, SLB-IL59, SLB-IL60, SLB-IL61, SLB-IL82, SLB-IL100, and SLB-IL111.

[0044] In the present invention, when the highly polar column is further most preferably an ionic liquid column, the saturated and unsaturated dialkyl ketones in the sample can be well separated from each other and more accurately quantified.

[0045] Examples of the ionic liquid column include a column in which the liquid phase is 1,12-Di(tripropylphosphonium)dodecane bis(trifluoromethylsulfonyl)imide, a column in which the liquid phase is 1,12-Di(tripropylphosphonium)dodecane bis(trifluoromethylsulfonyl)imide, a column in which the liquid phase is 1,12-Di(tripropylphosphonium)dodecane bis(trifluoromethylsulfonyl)imide trifluoromethylsulfonate, a column in which the liquid phase is Tri(tripropylphosphoniumhexanamido)triethylamine bis(trifluoromethylsulfonyl)imide, a column in which the liquid phase is 1,12-Di(2,3-dimethylimidazolium)dodecane bis(trifluoromethylsulfonyl)imide, a column in which the liquid phase is 1,9-Di(3-vinylimidazolium)nonane bis(trifluoromethylsulfonyl)imide, and a column in which the liquid phase is 1,5-Di(2,3-dimethylimidazolium)pentane bis(trifluoromethylsulfonyl)imide, and specific examples thereof can include SLB-IL59, SLB-IL60, SLB-IL61, SLB-IL82, SLB-IL100, and SLB-IL111.

Gas Chromatograph

[0046] In the method of the present invention, a gas chromatograph is used, and a commonly used gas chromatograph can be used as the gas chromatograph. Measurement conditions can be appropriately set so that saturated and unsaturated dialkyl ketones can be separated and quantified using the column described above.

[0047] For a detector and the like used in the gas chromatograph, commonly used ones can be used, and examples of the detector include a mass spectrometer (MS) and a flame ionization detector (FID).

EXAMPLES

[0048] The present invention will be described in more detail below by way of examples, but is not limited to these examples.

[0049] In describing the details of the present invention, the following columns were used for comparison: [0050] (Comparative Example 1) HP-1 MS UI (length: 30 m, internal diameter: 0.25 mm ID, membrane thickness: 0.25 ?m, nonpolar column, polarity value: about 130, liquid phase: 100% dimethylpolysiloxane/available from Agilent) [0051] (Comparative Example 2) HP-5 MS UI (length: 30 m, internal diameter: 0.25 mm ID, membrane thickness: 0.25 ?m, slightly polar column, polarity value: about 252, liquid phase: 5% diphenyl-95% dimethylpolysiloxane/available from Agilent) [0052] (Comparative Example 3) DB-XLB (length: 30 m, internal diameter: 0.25 mm ID, membrane thickness: 0.25 ?m, slightly polar column, polarity value: about 300, liquid phase: 14% diphenyl-86% dimethylpolysiloxane/available from Agilent) [0053] (Example 1) InertCap 1701 (length: 30 m, internal diameter: 0 25 mm ID, membrane thickness: 0.25 ?m, medium polar column, polarity value: about 700, liquid phase: 14% cyanopropylphenyl-86% dimethylsiloxane/available from GL Science) [0054] (Example 2) DB-17 ms (length: 30 m, internal diameter: 0.25 mm ID, membrane thickness: 0.25 ?m, medium polar column, polarity value: about 948, liquid phase: 50% diphenyl-50% dimethylpolysiloxane/available from Agilent) [0055] (Example 3) DB-WAX (length: 30 m, internal diameter: 0.25 mm ID, membrane thickness: 0.25 ?m, highly polar column, polarity value: about 2324, liquid phase: polyethylene glycol/available from Agilent) [0056] (Example 4) HR-SS-10 (length: 25 m, internal diameter: 0.25 mm ID, highly polar column, polarity value: about 2500, liquid phase: nitrile silicone/available from Shinwa Chemical Industries Ltd.) [0057] (Example 5) SLB-IL60 (length: 30 m, internal diameter: 0.25 mm ID, thickness: 0.2 ?m, highly polar column, polarity value: about 2622, liquid phase: 1,12-Di(tripropylphosphonium)dodecane bis(trifluoromethylsulfonyl)imide/available from Supelco)

(Analysis Example 1) Separation and Analysis of Unsaturated DAKs

Adjustment of Sample

[0058] For nine unsaturated DAK synthetic products (C11-CO-C17:1, C11-CO-C17:2, C13-CO-C17:1, C13-CO-C17:2, C15-CO-C17:1, C15-CO-C17:2, C17-CO-C17:1, C17:1-CO-C17:1, C17:1-CO-C17:2), each of the unsaturated DAK synthetic products was dissolved in a mixed solution of toluene and hexane so as to attain a concentration of 125 ppm, and the solution was subjected to GC.

[0059] DAKs are expressed so as to represent their chemical structures. For example, DAKs produced from two molecules of steric acid (C18) is expressed as C17-CO-C17 (18-pentatriacontanone), and DAKs produced from two molecules of oleic acid (C18:1) is expressed as C17:1-CO-C17:1 (pentatriacontane-9,26-dien-18-one). This notation is used hereinafter.

Analysis Conditions for GC

[0060] GC system: Agilent GC 7890B

Column:

[0061] (Comparative Example 1) HP-1 MS UI (30 m?0.25 mm, 0.25 ?m) [0062] (Comparative Example 2) HP-5 MS UI (30 m?0.25 mm, 0.25 ?m) [0063] (Comparative Example 3) DB-XLB (30 m?0.25 mm, 0.25 ?m) [0064] (Example 1) InertCap 1701 (30 m?0.25 mm, 0.25 ?m) [0065] (Example 5) SLB-IL60 (30 m?0.25 mm, 0.2 ?m) [0066] Injection port: 300? C., split mode, split ratio=25:1 [0067] Carrier gas: helium gas constant linear velocity mode, 38 cm/s [0068] Oven: 80? C. (retained for 1 min).fwdarw.200? C. (not retained, 25? C./min).fwdarw.290? C. (retained for 30 min, 5? C./min) [0069] FID temperature: 340? C. [0070] Injection volume: 1 ?L

[0071] The separation of unsaturated DAKs was confirmed based on the number of peaks on the chromatogram obtained in the analysis. The results are listed in Table 1.

TABLE-US-00001 TABLE 1 Separation of Column Polarity unsaturated DAKs Comparative HP-1MS UI Nonpolarity Was not able to Example 1 be separated. Comparative HP-5MS UI Slight Was not able to Example 2 polarity be separated. Comparative DB-XLB Slight Was not able to Example 3 polarity be separated. Example 1 InertCap 1701 Medium Separated polarity Example 5 SLB-IL60 High polarity Separated

[0072] In the case of using a nonpolar column (HP-1 MS UI) or a slightly polar column (HP-5 MS UI, DB-XLB) used in analysis of DAKs in the related art, nine unsaturated DAKs were not able to be separated, but, in the case of using a medium polar column (InertCap 1701) or a highly polar column (SLB-IL60), unsaturated DAKs were able to be separated.

(Analysis Example 2) Separation and Analysis of DAKs

Adjustment of Sample

[0073] For seventeen DAK synthetic products (C10-CO-C10, C11-CO-C11, C11-CO-C13, C11-CO-C15, C11-CO-C17, C11-CO-C17:1, C11-CO-C17:2, C15-CO-C15, C13-CO-C17:1, C13-CO-C17:2, C15-CO-C17, C15-CO-C17:1, C15-CO-C17:2, C17-CO-C17, C17-CO-C17:1, C17:1-CO-C17:1, C17:1-CO-C17:2), each of the DAK synthetic products was dissolved in a mixed solution of toluene and hexane so as to attain a concentration of 118 ppm, and the solution was subjected to GC.

Analysis Conditions for GC

[0074] GC system: Agilent GC 7890B

Column:

[0075] (Example 2) DB-17 ms (30 m?0.25 mm, 0.25 ?m) [0076] (Example 3) DB-WAX (30 m?0.25 mm, 0.25 ?m) [0077] (Example 4) HR-SS-10 (25 m?0.25 mm) [0078] (Example 5) SLB-IL60 (30 m?0.25 mm, 0.2 ?m) [0079] Injection port: 300? C., split mode, split ratio=25:1 [0080] Carrier gas: helium gas constant linear velocity mode, 38 cm/s

Oven:

[0081] DB-17 ms, SLB-IL60 [0082] 80? C. (retained for 1 min).fwdarw.180? C. (not retained, 25? C./min).fwdarw.290? C. (retained for 30 min, 5? C./min)

DB-WAX

[0083] 80? C. (retained for 1 min).fwdarw.180? C. (not retained, 25? C./min).fwdarw.245? C. (retained for 30 min, 5? C./min)

HR-SS-10

[0084] 80? C. (retained for 1 min).fwdarw.180? C. (not retained, 25? C./min).fwdarw.220? C. (retained for 30 min, 5? C./min) [0085] FID temperature: 340? C. [0086] Injection volume: 1 ?L.

[0087] The separation of the 17 DAK mixtures was confirmed on various polar columns. The results are listed in Table 2.

Table 2

[0088] As shown by the results, it was possible to separate and analyze saturated and unsaturated DAKs by using a medium polar column and a highly polar column.

(Analysis Example 3) Quantitative Analysis of DAKs Using Highly Polar Column SLB-IL60

Example 5

Adjustment of Sample

[0089] For sixteen DAK synthetic products (C11-CO-C11, C11-CO-C13, C11-CO-C15, C11-CO-C17, C11-CO-C17:1, C11-CO-C17:2, C15-CO-C15, C13-CO-C17:1, C13-CO-C17:2, C15-CO-C17, C15-CO-C17:1, C15-CO-C17:2, C17-CO-C17, C17-CO-C17:1, C17:1-CO-C17:1, C17:1-CO-C17:2), each of the DAK synthetic products was dissolved in a mixed solution of toluene and hexane so as to attain a concentration of from 0.2 to 250 ppm. In total, six types of solutions in concentration were subjected to GC, respectively.

Analysis Conditions for GC

[0090] GC system: Agilent GC 7890B [0091] Column: SLB-IL60 (30 m?0.25 mm, 0.2 ?m) [0092] Injection port: 300? C., split mode, split ratio=25:1 [0093] Carrier gas: helium gas constant linear velocity mode, 38 cm/s [0094] Oven: 80? C. (retained for 1 min).fwdarw.180? C. (not retained, 25? C./min).fwdarw.290? C. (retained for 30 min, 5? C./min) [0095] FID temperature: 340? C. [0096] Injection volume: 1 ?L

Processing of Analysis Data

[0097] OpenLab CDS software (version 2.5) available from Agilent was used in processing of the resulting chromatogram. That is, a calibration curve of the peak area provided by peak picking and the injection concentration was created, and a square value (R2) of a correlation coefficient R and slope and intercept of the calibration curve were calculated. A detection limit (LOD) and a lower limit of quantification (LOQ) were calculated as concentrations at which the SN ratio was 3 and 10, respectively. The results are listed in Table 3.

TABLE-US-00002 TABLE 3 RT Inter- LOD LOQ DAKs (min) R2 Slope cept (ppm) (ppm) C10-C0- 12.645 C10 C11-C0- 14.699 0.9998 0.5605 0.6609 0.14 ? 0.03 0.45 ? 0.09 C11 C11-C0- 16.732 0.9997 0.6955 0.6651 0.13 ? 0.02 0.42 ? 0.08 C13 C11-C0- 18.734 0.9997 0.5107 0.7081 0.15 ? 0.02 0.51 ? 0.05 C15 C11-C0- 20.612 0.9997 0.6662 0.6834 0.16 ? 0.01 0.53 ? 0.01 C17 C11-C0- 20.868 0.9997 0.5122 0.6860 0.18 ? 0.01 0.59 ? 0.03 C17:1 C11-C0- 21.336 0.9997 0.5113 0.6279 0.18 ? 0.03 0.61 ? 0.10 C17:2 C15-C0- 22.417 0.9997 0.4690 0.6261 0.21 ? 0.02 0.69 ? 0.05 C15 C13-C0- 22.651 0.9997 0.6078 0.6973 0.15 ? 0.01 0.50 ? 0.05 C17:1 C13-C0- 23.162 0.9997 0.4300 0.6605 0.23 ? 0.01 0.75 ? 0.04 C17:2 C15-C0- 24.096 0.9998 0.5673 0.4850 0.14 ? 0.01 0.47 ? 0.04 C17 C15-C0- 24.38 0.9998 0.4918 0.5827 0.19 ? 0.13 0.64 ? 0.10 C17:1 C15-C0- 24.843 0.9998 0.5367 0.4746 0.20 ? 0.02 0.65 ? 0.08 C17:2 C17-C0- 25.719 0.9998 0.5189 0.5639 0.21 ? 0.03 0.71 ? 0.10 C17 C17-C0- 25.974 0.9999 0.5819 0.3801 0.23 ? 0.02 0.75 ? 0.06 C17:1 C17:1-C0- 26.277 0.9998 0.4907 0.5076 0.40 ? 0.14 1.34 ? 0.47 C17:1 C17:1-C0- 26.739 0.9998 0.5363 0.3832 0.45 ? 0.03 1.54 ? 0.11 C17:2 *Values of LOD and LOQ were mcans ? S. D. (n = 3)

[0098] As shown in Table 3, a highly correlated calibration curve (R2>0.999) was able to be created, and a low LOD value (>0.46 ppm) and a low LOQ value (>1.54 ppm) were calculated, demonstrating that this method is a technique having high quantitativity.

(Example 4) Addition Recovery Test

Adjustment of Sample

[0099] For sixteen DAK synthetic products (C11-CO-C11, C11-CO-C13, C11-CO-C15, C11-CO-C17, C11-CO-C17:1, C11-CO-C17:2, C15-CO-C15, C13-CO-C17:1, C13-CO-C17:2, C15-CO-C17, C15-CO-C17:1, C15-CO-C17:2, C17-CO-C17, C17-CO-C17:1, C17:1-CO-C17:1, C17:1-CO-C17:2) each of the DAK synthetic products was dissolved in a mixed solution of toluene and hexane, and the solution was added to palm refined oil free of DAKs (available from Fuji Oil Co., Ltd.) so as to attain a final concentration of 50 mg/kg oil. The adjusted samples were used.

[0100] Before being subjected to GC, the sample was subjected to the following pretreatment for extraction and purification.

Pretreatment

[0101] To 2 g of the sample, the internal standard substance (C10-CO-C10) was added so as to attain a final concentration of 100 ppm, and 12 mL of ethanol and 2 mL of a 50% KOH aqueous solution were added. The mixture was heated at about 80? C. for 1 hour, and subjected to a saponification reaction. After cooling, 10 mL of distilled water was added, extraction was performed three times with petroleum ether, and the recovered petroleum ether fraction was washed three times with 10% ethanol-water. The petroleum ether layer was dehydrated with sodium sulfate. Then, the solvent of the filtrate was distilled off, followed by re-dissolution with hexane, the whole amount was passed through a silica gel column (Supelco, Discovery DSC-Si SPE Tube bed wt. 2 g, volume 12 mL) conditioned with hexane, and the DAKs were eluted with 15 mL of hexane/ether (95/5 v/v). After the solvent of the resulting fraction was distilled off, the residue was dissolved in a mixed solution of toluene and hexane, and the solution was subjected to GC.

Analysis Conditions for GC

[0102] GC system: Agilent GC 7890B [0103] Column: SLB-IL60 (Example 4) (30 m?0.25 mm, 0.2 ?m) [0104] Injection port: 300? C., split mode, split ratio=25:1 [0105] Carrier gas: helium gas constant linear velocity mode, 38 cm/s [0106] Oven: 80? C. (retained for 1 min).fwdarw.180? C. (not retained, 25? C./min).fwdarw.290? C. (retained for 30 min, 5? C./min) [0107] FID temperature: 340? C. [0108] Injection volume: 1 ?L

[0109] After the analysis result of the sample was corrected with the internal standard, the amounts of the DAKs were quantified using the calibration curve created in Example 3, and a recovery rate was calculated in percentage (%) from comparison with theoretical values of the amounts added. The results are listed in Table 4.

TABLE-US-00003 TABLE 4 DAKs Recovery rate (%) C11-CO-C11 102.2 ? 0.4 C11-CO-C13 103.0 ? 0.4 C11-CO-C15 101.5 ? 0.4 C11-CO-C17 102.9 ? 0.5 C11-CO-C17:1 102.9 ? 0.6 C11-CO-C17:2 103.4 ? 0.8 C15-CO-C15 103.5 ? 0.8 C13-CO-C17:1 102.7 ? 0.4 C13-CO-C17:2 102.9 ? 0.2 C15-CO-C17 101.3 ? 0.9 C15-CO-C17:1 103.9 ? 1.2 C15-CO-C17:2 101.9 ? 0.5 C17-CO-C17 102.3 ? 0.7 C17-CO-C17:1 101.5 ? 0.6 C17:1-CO-C17:1 104.6 ? 0.9 C17:1-CO-C17:2 103.9 ? 0.6

[0110] As a result, high recovery rates of from 101% to 104% were provided, indicating that the developed method has both high quantitativity and practicability.