Method for detecting fatty acid methyl ester and fatty acid ethyl ester in biological fuel oil

Abstract

The invention discloses a detection method and application of fatty acid methyl ester and fatty acid ethyl ester in biological fuel oil, the detection method comprises the following steps: S1, dissolving the biological fuel oil with a solvent to obtain a solution, carrying out solid-phase extraction, and collecting an eluent as a solution to be detected; and S2, carrying out gas chromatography-mass spectrometry analysis on the solution to be detected in the step S1 by using an external standard method and adopting a direct sample injection mode, and calculating to obtain the contents of fatty acid methyl ester and fatty acid ethyl ester in the biological fuel oil. According to the method, the biological fuel oil is dissolved by adopting a specific solvent, then solid-phase extraction is carried out by adopting a specific solid-phase extraction column, a specific eluting agent and a specific eluent, and analysis is carried out by utilizing gas chromatography-mass spectrometry of a specific gas chromatography stationary phase. Qualitative and quantitative detection of the fatty acid methyl ester and the fatty acid ethyl ester in the biological fuel oil is achieved, and the method is high in sensitivity, accuracy, precision and reproducibility.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of marine fuel oil detection, and more particularly to a method for detecting fatty acid methyl esters and fatty acid ethyl esters in biofuel oil BACKGROUND With the urgent demand for carbon emission reduction in the global shipping industry, marine biofuel oil as a low-carbon substitute for traditional fossil fuels has become one of the core technical paths to achieve the International Maritime Organization (IMO) greenhouse gas emission reduction strategy by 2050. Biodiesel is a typical green and environmentally friendly alternative fuel, and its core component is fatty acid methyl ester (FAME) and fatty acid ethyl ester (FAEE) extracted from animal and vegetable oils through ester exchange reaction. This type of biofuel, due to its biodegradable properties, extremely low sulfur content, and sustainable regeneration, etc., has shown significant environmental protection characteristics, and has been vigorously promoted in Europe, the United States and China. With the tightening of environmental protection policies in various countries, the proportion of biodiesel in fuel oil is gradually increasing, for example, the T / CIQA88-2024 standard clearly stipulates that the FAME content in B20-B50 fuel oil needs to be monitored. On the other hand, the addition of biodiesel also brings a series of technical challenges. Fuel containing fatty acid methyl ester and fatty acid ethyl ester is also prone to produce strong irritant volatile organic compounds such as propylene aldehyde during low-temperature combustion, further increasing the risk of respiratory diseases in the contact group, and may also cause a series of technical problems such as decreased oxidation stability of oil, microbial reproduction, and fuel filter clogging. Therefore, accurately determining the content of fatty acid methyl ester and fatty acid ethyl ester compounds in fuel oil is of great significance for the quality monitoring and scientific research of marine biofuel oil, and even for the safety of shipping and equipment, human health and port ecological protection.

[0002] FAME and FAEE, although similar in chemical structure (both formed by ester compounds of fatty acid chains and alcohol groups), have significant differences in physicochemical properties. FAEE has a lower freezing point and higher oxidation stability than FAME, which makes FAEE have better use value. However, due to the current manufacturing cost reasons in the FAME and FAEE industry, fatty acid esters are generally added FAME, and relevant detection standards such as NB / SH / T 0916-2015 clearly stipulate that they are only applicable to the determination of FAME. Once the fuel oil contains FAEE, the detection result will have a significant negative deviation. This deviation is particularly prominent in the detection of mixed biodiesel, which may cause quality misjudgment and regulatory loopholes.

[0003] The detection of ester compounds in current fuel oils faces the dual challenges of insufficient method specificity and serious matrix interference. The EU EN 14078 standard shows the innovative needs of the detection system brought by biofuel blending. The 2003 original standard can only detect middle distillate oil in the concentration range of 1.7%-22.7%, and the 2014 revised version has made two breakthroughs: the detection range has been greatly widened to 0.05%-50% and divided into four calibration intervals; and the technical details use diesel oil without fatty acid methyl ester (FAME) instead of cyclohexane as the dilution solvent, and according to the concentration gradient, 0.1 mm or 0.5 mm sample cell is selected. However, the current standard system still has a significant technical gap: it has not solved the cross-interference problem of fatty acid ethyl ester (FAEE), and it has not covered the detection of high-viscosity oil such as marine heavy fuel oil. It is worth noting that the American ASTM D8144-18 standard innovatively uses solid phase extraction-gas chromatography to realize the simultaneous determination of aromatics and FAME, but it also omits the detection requirements of FAEE. This technical gap has not been filled in the Chinese-led ASTM D8144-18 standard and the revised version of GB / T 23801, resulting in a lack of authoritative detection basis for cross-border bonded marine fuel oil quality disputes.

[0004] Currently, the existing detection methods mainly include infrared spectroscopy, gas chromatography (GC), high performance liquid chromatography (HPLC), gas chromatography tandem mass spectrometry (GC-MS / MS), and gas chromatography-mass spectrometry. Infrared spectroscopy has the advantages of simple operation, rapidness and economy, but it has limitations in FAEE specificity detection, complex matrix applicability and anti-interference ability; gas chromatography can realize component separation but is limited by FAEE co-elution, and is not suitable for heavy fuel oil containing a large amount of unseparated impurities; HPLC has good selectivity, but its separation ability for FAEEs homologues is limited; GC-MS / MS technology solves the problem of distinguishing FAME and FAEE through specific ion recognition, and realizes trace detection of FAME / FAEE through MRM mode, but on the one hand, the equipment cost is high, and on the other hand, the accuracy is low when detecting samples with high FAME / FAEE content (such as biofuel oil).

[0005] GC-MS (gas chromatography-mass spectrometry) has the characteristics of low-cost equipment, and is a detection technology possessed by conventional laboratories. Currently, there is no prior art of GC-MS method for detecting FAEE in biofuel oil, and there is no prior art of GC-MS method for simultaneously detecting FAME and FAEE contents in biofuel oil. Especially, since the mass spectrometry of GC-MS has weak qualitative ability for FAME / FAEE, it needs to rely more on gas chromatography to separate and qualify FAME, FAEE and FAME and FAEE. Moreover, the types of FAME and FAEE in biofuel oil may be up to 30 or more, and the properties and structures of such fatty acid esters are similar. It is quite difficult to simultaneously realize 30 or more FAME and FAEE through gas chromatography. SUMMARY

[0006] In order to solve the problem that the prior art cannot simultaneously realize the detection of FAME and FAEE or needs to rely on high-cost detection equipment, the present application provides a detection method for fatty acid methyl ester and fatty acid ethyl ester in biofuel oil. The detection method can realize high-selectivity qualitative and quantitative analysis of FAME and FAEE, has high sensitivity, precision and accuracy, and can meet the accurate quantitative demand of fatty acid ester compounds in marine fuel oil.

[0007] Another object of the present application is to provide the application of the above detection method in marine fuel oil detection.

[0008] The above objects of the present application are realized by the following technical solutions. A detection method for fatty acid methyl ester and fatty acid ethyl ester in biofuel oil, comprising the following steps: S1. Solvent is used to dissolve biofuel oil to obtain a dissolution solution, solid phase extraction is performed, and eluate is collected as a to-be-tested solution; S2. External standard method is used to perform gas chromatography-mass spectrometry analysis on the to-be-tested solution of step S1 by direct injection, and the contents of fatty acid methyl ester and fatty acid ethyl ester in biofuel oil are calculated; In step S1, the solvent is a mixed solvent of n-hexane and dichloromethane; the stationary phase of the solid phase extraction column for solid phase extraction is a mixture of palladium salt, silver salt, aluminum oxide and silica gel; the eluent for solid phase extraction is cyclohexane, and the eluent is a mixed solvent of dichloromethane and ethyl acetate; In step S2, the stationary phase of the chromatographic column used in the gas chromatographic analysis is polyethylene glycol, and the temperature rising degree of the gas chromatography is: temperature rising program: initial temperature 120℃, holding for 1 min, then rising to 200℃ at 20℃ / min, holding for 5 min, then rising to 260℃ at 10℃ / min, holding for 20 min.

[0009] It should be noted that: The specific solvent is used to dissolve the biofuel oil, then the specific solid-phase extraction column, the specific eluent and the specific eluent are used for solid-phase extraction, and then the specific gas chromatography stationary phase of gas chromatography-mass spectrometry is used for analysis, so that the qualitative and quantitative detection of fatty acid methyl ester and fatty acid ethyl ester in the biofuel oil is realized, and the sensitivity, accuracy, precision and reproducibility are high. Among them, the present application does not depend on expensive equipment such as GC-MS / MS technology, but only through the cooperation of the stationary phase of the gas chromatography in the gas chromatography-mass spectrometry and the specific temperature rising degree, 35 kinds of fatty acid methyl ester (including methyl octanoate, methyl decanoate, methyl dodecanoate, methyl tetradecanoate, methyl hexadecanoate, methyl palmitoleate, methyl octadecanoate, methyl oleate, methyl linoleate, methyl linolenate, methyl eicosanoate, methyl docosanoate, methyl erucic acid, methyl tetracosanoate) and fatty acid ethyl ester (including methyl tetradecanoate, methyl hexadecanoate, methyl palmitoleate, methyl heptadecanoate, methyl octadecanoate, methyl oleate, methyl linoleate, methyl linolenate, methyl eicosanoate, 11-cis-eicosan-1-ol, methyl eicosatrienoate (cis-11, 14, 17), methyl arachidonic acid, methyl eicosapentaenoic acid, methyl heneicosanoic acid, methyl docosanoic acid, methyl erucic acid, methyl docosapentaenoate, methyl docosahexaenoic acid, methyl heneicosanoic acid, methyl tetracosanoic acid, methyl tetracosene acid) can be baseline separated, so that the detection method can be popularized in most laboratories. In addition, the detection method of the present application also has the characteristics of simple operation.

[0010] Preferably, the biofuel oil is a marine biofuel oil.

[0011] Preferably, the total content of FAME and FAEE in the biofuel oil is ≤50 wt%.

[0012] The detection method of the present application can not only detect trace FAME / FAEE samples, but also detect large amount of FAME / FAEE samples.

[0013] More preferably, the total content of FAME and FAEE in the biofuel oil is 20-30 wt%.

[0014] Preferably, before the dissolving in step S1, the method further comprises a step of performing vacuum distillation on the biofuel oil and collecting light components.

[0015] The vacuum distillation treatment can improve the recovery rate of the target substance, thereby improving the accuracy.

[0016] Specifically, step S1 comprises: performing vacuum distillation on the biofuel oil and collecting light components; dissolving the light components with a solvent to obtain a dissolving solution; and performing solid phase extraction and collecting an eluent as the to-be-tested solution.

[0017] More preferably, the temperature of the vacuum distillation is 260-270°C, and the pressure is 4-8 bar.

[0018] Preferably, the volume ratio of the dissolving solution to the eluent in step S1 is 1:(1.8-3).

[0019] Preferably, the volume ratio of the dissolving solution to the eluent in step S1 is 1:(2.2-3.5).

[0020] Preferably, in the mixed solvent of n-hexane and dichloromethane, the volume ratio of n-hexane to dichloromethane is (3.8-4.2):1.

[0021] Preferably, in the mixed solvent of dichloromethane and ethyl acetate, the volume ratio of dichloromethane to ethyl acetate is (3.8-4.2):1.

[0022] Preferably, after the collection of the eluent in step S1, the method further comprises a step of concentrating and constant volume.

[0023] More preferably, the method of concentrating and constant volume is nitrogen blow concentration.

[0024] Preferably, the mass ratio of the palladium salt, the silver salt, the aluminum oxide and the silica gel is 0.25:(0.2-0.3):(40-60):(18-22).

[0025] Preferably, the preparation method of the silica gel solid phase extraction column in step S1 is: The palladium salt, the silver salt, the aluminum oxide and the silica gel are mixed to obtain a mixture; and the mixture is filled into a container to obtain the solid phase extraction column.

[0026] More preferably, the palladium salt, the silver salt, the aluminum oxide and the silica gel are mixed with an organic solvent.

[0027] Further preferably, the organic solvent is acetone.

[0028] Further preferably, the specific process of the mixing is: first mixing the palladium salt, the silver salt and the organic solvent, then adding the aluminum oxide to mix, drying to obtain a mixed powder; and then mixing the mixed powder with the silica gel.

[0029] Further preferably, the drying temperature is 75-85℃, and the drying time is 1.5-2.5h.

[0030] More preferably, the palladium salt is palladium acetate.

[0031] More preferably, the silver salt is silver acetate.

[0032] More preferably, the aluminum oxide is dried aluminum oxide.

[0033] More preferably, before the palladium salt-silver salt-aluminum oxide-silica gel mixture is filled into the container, the process further comprises the step of laying a bottom padding on the bottom of the container.

[0034] Further preferably, the padding is glass wool.

[0035] More preferably, after the palladium salt-silver salt-aluminum oxide-silica gel mixture is filled into the container, the process further comprises the steps of: discharging the solvent, then laying a top padding on the top of the palladium salt-silver salt-aluminum oxide-silica gel mixture, and then activating.

[0036] Further preferably, the method of discharging the solvent is extrusion.

[0037] Further preferably, the top padding is glass fiber.

[0038] Preferably, before the solid phase extraction, the process further comprises the step of activating the solid phase extraction column.

[0039] Further preferably, the activation process is: adding a non-polar solvent into the container; and the non-polar solvent is at least one of n-hexane and n-heptane.

[0040] Preferably, the carrier gas for the gas chromatography analysis in step S2 is at least one of helium, neon and argon.

[0041] Preferably, the gas chromatography analysis in step S2 is performed under the following conditions: carrier gas: high-purity helium, column flow rate: 1 mL / min; injection mode: splitless injection, septum purge flow rate: 20 mL / min; injection port temperature: 260℃; injection volume: 1 μL; transfer line temperature: 260℃.

[0042] Preferably, the ion source for the mass spectrometry analysis in step S2 is an electron impact ion source.

[0043] Preferably, the detection mode for the mass spectrometry analysis in step S2 is selected ion mode.

[0044] More preferably, the mass spectrometry analysis in step S2 is performed under the following conditions: Ion source: Electron impact ion source (EI); ionization energy: 70 eV; ion source temperature: 230 °C; quadrupole mass analyzer temperature: 150 °C; detection mode: selected ion mode (SIM); solvent delay: 4 min.

[0045] Preferably, the fatty acid methyl ester is at least one of methyl octanoate, methyl decanoate, methyl dodecanoate, methyl tetradecanoate, methyl hexadecanoate, methyl palmitoleate, methyl octadecanoate, methyl oleate, methyl linoleate, methyl linolenate, methyl eicosanoate, methyl docosanoate, methyl erucate, methyl tetracosanoate.

[0046] Preferably, the fatty acid ethyl ester is at least one of ethyl tetradecanoate, ethyl hexadecanoate, ethyl palmitoleate, ethyl heptadecanoate, ethyl octadecanoate, ethyl oleate, ethyl linoleate, ethyl linolenate, ethyl eicosanoate, ethyl 11-cis-eicosenoate, ethyl eicosatrienoate (cis-11,14,17), ethyl arachidonate, ethyl eicosapentaenoate, ethyl heneicosanoate, ethyl docosanoate, ethyl erucate, ethyl docosapentaenoate, ethyl docosahexaenoate, ethyl tricosanoate, ethyl tetracosanoate, ethyl tetracosenoate.

[0047] Compared with the prior art, the present application has the following advantages: The application adopts specific solvents to dissolve biofuel oil, then adopts specific solid-phase extraction column, specific eluent and specific eluent to perform solid-phase extraction, and then uses specific gas chromatography stationary phase of gas chromatography-mass spectrometry to analyze, so as to realize qualitative and quantitative detection of fatty acid methyl ester and fatty acid ethyl ester in biofuel oil, and the sensitivity, accuracy, precision and reproducibility are high. Among them, the application does not need to rely on expensive equipment of GC-MS / MS technology, but only through the cooperation of the stationary phase of gas chromatography in gas chromatography-mass spectrometry and the specific temperature rising degree, 35 kinds of fatty acid methyl ester (including methyl octanoate, methyl decanoate, methyl dodecanoate, methyl tetradecanoate, methyl hexadecanoate, methyl palmitoleate, methyl octadecanoate, methyl oleate, methyl linoleate, methyl linolenate, methyl eicosanoate, methyl docosanoate, methyl erucate, methyl tetracosanoate) and fatty acid ethyl ester (including ethyl tetradecanoate, ethyl hexadecanoate, ethyl palmitoleate, ethyl heptadecanoate, ethyl octadecanoate, ethyl oleate, ethyl linoleate, ethyl linolenate, ethyl eicosanoate, ethyl 11-cis-eicosan-1-enoate, ethyl eicosatrienoate (cis-11, 14, 17), ethyl arachidonate, ethyl eicosapentaenoate, ethyl heneicosanoate, ethyl docosanoate, ethyl erucate, ethyl docosapentaenoate, ethyl docosahexaenoate, ethyl heneicosanoate, ethyl tetracosanoate, ethyl tetracosenoate) can be baseline separated, so that the detection method can be popularized in most laboratories. BRIEF DESCRIPTION OF DRAWINGS

[0048] Figure 1 It is a typical selected ion total ion chromatogram of 35 kinds of fatty acid methyl ester and fatty acid ethyl ester compounds.

[0049] Figure 2 It is a graph of the influence of equilibrium temperature on the analysis results of several fatty acid methyl ester compounds in headspace sampling mode.

[0050] Figure 3 It is a graph of the influence of reduced pressure distillation temperature on the recovery rate of two kinds of fatty acid methyl ester compounds. DETAILED DESCRIPTION

[0051] The application will be further described below in combination with the drawings and specific examples in the specification, but the examples do not limit the application in any form. Unless otherwise specified, the reagents, methods and equipment used in the application are conventional reagents, methods and equipment in the technical field.

[0052] Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0053] Some reagents selected by the embodiments of the application are described as follows: Fatty acid methyl ester (FAME) mix: Dr. Ehrenstorfer GmbH, Germany, purity > 99.7%, Fatty acid methyl ester mix is an equimolar mixture of the following compounds: Methyl octoate, Methyl decanoate, Methyl laurate, Methyl myristate, Methyl palmitate, Methyl palmitoleate, Methyl stearate, Methyl oleate, Methyl linoleate, Methyl linolenate, Methyl arachidate, Methyl behenate, Methyl erucate, Methyl lignocerate; Fatty acid ethyl esters (FAEE) mix: NU-CHEK, USA, purity > 98%, fatty acid ethyl esters mix is an equal mixture of the following compounds: Ethyl myristate, Ethyl palmitate, Ethyl palmitoleate, Ethyl heptadecanoate, Ethyl stearate, Ethyl oleate, Ethyl linoleate, Ethyl linolenate, Ethyl arachidate, Ethyl 11-eicosenoate, Ethyl 11,14,17-eicosatrienoate, Ethyl arachidonate, Ethyl eicosapentaenoate, Ethyl heneicosanoate, Ethyl behenate, Ethyl erucate, Ethyl docosapentaenoate, Ethyl docosahexaenoate, Ethyl tricosanoate, Ethyl lignocerate, Ethyl nervonate; Standard working solution (FAME standard solution, FAEE standard solution), the preparation method is as follows: respectively take 10 mg FAME mixed standard and 10 mg FAEE mixed standard, add into 10 mL volumetric flask, dilute with n-hexane to constant volume, obtain FAME / FAEE standard stock solution with concentration of 1000 mg / L. Accurately transfer 1000 μL FAME / FAEE standard stock solution into two 10 mL volumetric flasks, constant volume with methanol, obtain FAME / FAEE standard intermediate solution with concentration of 100 mg / L. Accurately transfer 200 μL, 500 μL, 1000 μL of 1000 mg / L FAME / FAEE standard intermediate solution into 3 10 mL volumetric flasks respectively; accurately transfer 250 μL, 500 μL, 1000 μL of 100 mg / L FAME / FAEE standard stock solution into 3 10 mL volumetric flasks respectively, constant volume with n-hexane / dichloromethane (4 / 1, V / V) mixed solution as solvent, prepare FAME / FAEE standard working solution with FAME and FAEE concentration of 2.0 mg / L, 5.0 mg / L, 10 mg / L, 25 mg / L, 50 mg / L, 100 mg / L respectively.

[0054] Palladium-silver-doped silica gel solid phase extraction column: self-made, the preparation method is as follows: Dissolve about 0.25 g palladium acetate powder with 10 mL acetone, after complete dissolution, add 0.25 g silver acetate powder, after dissolution, add about 50 g roasted alumina, ultrasonic treatment and continuously stir to mix uniformly, stir to form a viscous slurry. Stand for 24 h, place it in a constant temperature oven, first dry at 80℃ for 2 h, after cooling, obtain mixed powder and store in a dry brown bottle. Take 10.0 g of the above mixed powder and mix with 20.0 g roasted silica gel for standby. Take a 6 mL syringe, put appropriate amount of glass wool at the bottom, then add 5 mL n-hexane, then take 3.00 g of the above palladium acetate-silver acetate-alumina-silica gel mixture and add into the syringe, gently knock to compact the filler. Extrude to discharge the remaining n-hexane, add another layer of glass fiber membrane on the top of the syringe column filler, re-add 5 mL n-hexane for activation and balance, and discharge the n-hexane for standby.

[0055] Example 1 The embodiment provides a method for detecting fatty acid methyl ester and fatty acid ethyl ester in biofuel oil, comprising the following steps: I. Pretreatment of sample First, 1.0 g of biofuel oil was taken in a distillation apparatus, and vacuum distillation was carried out at a temperature of 260°C and a pressure of 6 bar to collect the light components. After ultrasonic dissolution of the light components with n-hexane / dichloromethane (4 / 1, V / V) and constant volume to 10 mL, a dissolved solution was obtained. 1.0 mL of the dissolved solution was added to an activated and doped palladium-silver silica gel solid-phase extraction column, and after the dissolved solution was completely absorbed, 2.0 mL of n-hexane was added for elution, the eluent was discarded, and 2.5 mL of n-hexane / dichloromethane (4:1, V / V ) mixed solvent was added for elution, and the eluent was collected and concentrated to 1.0 mL by nitrogen blowing, filtered, and the measured solution was obtained for machine analysis.

[0056] II. Gas chromatography-mass spectrometry analysis The external standard method was used, and the measured solution and standard working solution were analyzed by gas chromatography-mass spectrometry in a direct injection mode, and the contents of fatty acid methyl esters and fatty acid ethyl esters in the biofuel oil were calculated; Gas chromatography-mass spectrometry instrument: Agilent 8890A-5977C gas chromatograph-mass spectrometer, American Agilent Company; Gas chromatography analysis conditions: chromatographic column: HP-INNOWAX capillary column, specifications (60 m x 0.320 mm x 0.50 µm); carrier gas: high-purity helium, column flow rate 1 mL / min; injection mode: splitless injection, septum purge flow: 20 mL / min; injection port temperature 260°C; injection volume 1 µL; temperature program: initial temperature 120°C, hold for 1 min, increase to 200°C at 20°C / min, hold for 5 min, then increase to 260°C at 10°C / min, hold for 20 min; transfer line temperature: 260°C.

[0057] Mass spectrometry analysis conditions: ion source: electron impact ion source (EI); ionization energy: 70 eV; ion source temperature: 230°C; quadrupole mass analyzer temperature: 150°C; detection mode: selected ion mode (SIM); solvent delay: 4 min.

[0058] The quantitative and qualitative ion parameters of FAME and FAEE compounds are shown in Table 1; the typical selected ion total ion chromatogram of 35 kinds of FAME and FAEE compounds is shown in Figure 1 .

[0059] Table 1 Quantitative and qualitative ion parameters of 35 kinds of FAME and FAEE compounds

[0060] Example 2 Methodology verification (1) Matrix effect evaluation: In this study, considering that the complex matrix environment of the actual fuel sample may interfere with the precision and accuracy of the analysis method, it is necessary to evaluate the matrix effect of the method in depth. Therefore, according to the method of Example 1, the blank matrix solutions of No. 5 fuel oil, No. 6 fuel oil, No. 7 fuel oil, marine fuel oil and transformer oil were prepared by processing the blank matrix samples. The same concentration of target fatty acid ester standard addition solution as the pure solvent standard curve 1-10 mg / L was added to the blank matrix extract, mixed, and then injected and analyzed under the same conditions as the pure solvent standard solution. The matrix effect was evaluated by calculating the ratio of the peak area of the blank matrix extract to the peak area of the pure solvent standard curve. The matrix effect evaluation results of 35 kinds of fatty acid ester (FAME & FAEE) compounds are shown in Table 2. The results show that in the evaluated fuel oil, the ME% range is between -10-10%, indicating that there is no significant matrix effect in the detection of target fatty acid esters under the analysis conditions provided in this example.

[0061] (2) Linear range and lower limit of quantification of the method The signal-to-noise ratio (S / N) of 3 times was used as the method detection limit (LOD), and the standard deviation (SMDL) of 7 repeated tests was multiplied by the t-distribution coefficient (t(n-1, 1-α=0.99)) to verify the effectiveness of the detection limit. At the same time, the signal-to-noise ratio (S / N) of 10 times was used as the lowest limit of quantification (LOQ). The correlation coefficient and lower limit of quantification of 35 kinds of fatty acid esters (FAME & FAEE) compounds are shown in Table 2. The results show that the method detection limit of 35 kinds of fatty acid esters (FAME & FAEE) compounds ranges from 0.1 to 1.2 mg / kg, and the quantification limit ranges from 0.3 to 4.0 mg / kg. These compounds all show good linear relationship in the concentration range of 0.05-2.50 mg / L, and the correlation coefficient is greater than 0.9955.

[0062] Table 2 Method quantification limit and matrix effect evaluation

[0063] (3) Method recovery and precision: In this study, 10 mg / L of 35 fatty acid ester (FAME & FAEE) compound mixed standard intermediate liquid was used for method validation. The experiment used a three-stage gradient standard design: 100 μL, 500 μL of FAME & FAEE standard intermediate liquid and 250 μL, 300 μL of FAME & FAEE standard stock solution were accurately transferred into the blank marine biofuel oil matrix, respectively, and then diluted to 10 mL to form the standard addition system. Finally, four concentration gradients of 1.0 mg / L, 5.0 mg / L, 25 mg / L, and 30 mg / L were obtained. Each concentration level was determined in 6 replicates, and the relative standard deviation RSD and recovery rate are shown in Table 3. The results show that the matrix standard recovery rate of the target compound is 82%~116%, and the relative standard deviation (RSD) is ≤5.5%, all parameters meet the quality control standards of trace analysis.

[0064] Table 3 Recovery and relative standard deviation of fatty acid ester (FAME & FAEE) compounds

[0065] Example 3 Optimization of conditions 1. Selection of sample extraction and purification method Biofuel oil is a complex mixture of hydrocarbons, containing a large amount of non-hydrocarbon compounds, gum and asphaltene components in the matrix. These substances not only pollute the analysis equipment, but also may interfere with the accuracy of the analysis results. Therefore, three pretreatment methods of liquid-liquid extraction, solid-phase extraction and microwave-assisted extraction were compared. In addition to the sample extraction and purification method, the other contents of the test method are the same as Example 1.

[0066] The liquid-liquid extraction method uses a mixture of n-hexane and dichloromethane as the solvent system. Although the operation process is simple, only extraction and filtration are required before analysis, this method has obvious limitations: on the one hand, a large amount of interfering impurities will be co-extracted, on the other hand, the recovery rate of the target compound is often low.

[0067] Microwave-assisted extraction technology uses microwave energy to promote solvent molecules to oscillate at high speed, significantly improving the ability of the solvent to penetrate the sample matrix, thereby accelerating the dissolution rate of ester compounds. However, the high temperature conditions of this method may cause the degradation of heat-sensitive esters, so it is not suitable.

[0068] Marine biofuel oil is completely dissolved by mixed solvent, and the extraction liquid often contains original interfering substances of oil products, such as various asphaltenes, resins, isomeric alkanes, polar compounds and dispersants, etc. These interfering substances may cause errors in subsequent gas chromatography-mass spectrometry analysis, and therefore must be purified and separated. Comparative tests were carried out by using C18-H carbon octadecyl column, AL-N neutral alumina column, Florisil-PR Florisil column, Silica silica gel column and silica gel solid phase extraction column doped with palladium-silver for comparison, and cyclohexane was used as eluent to wash out non-polar oligomers and saturated hydrocarbons from the separation column, and then dichloromethane / methanol mixed solution was used for elution. The results show that the matrix interference of C18-H carbon octadecyl column is serious, a large amount of non-polar and weakly polar hydrocarbons in fuel oil are also strongly retained by C18 column, and it is difficult to achieve selective separation and the purification effect is poor; the retention capacity of AL-N neutral alumina column for target fatty acid methyl ester is insufficient, and it is difficult to achieve effective separation and enrichment, and the activation and use conditions of alumina need to be strictly controlled, which is a challenge to the reproducibility; the adsorption capacity of Florisil-PR Florisil column is stronger than that of alumina, but its strong adsorption property may cause fatty acid esters to be excessively retained, elution is difficult, and the recovery efficiency is low; the Silica silica gel column has suitable retention capacity for fatty acid ester oil, which can effectively exclude the interference of main non-polar hydrocarbon matrix, but cannot exclude the interference of polar matrix such as aromatic hydrocarbon; the silica gel solid phase extraction column doped with palladium-silver has good purification capacity, the highest recovery rate and better anti-interference capacity of complex matrix, which may be due to the weak coordination between the carbonyl group of the target fatty acid and the palladium-silver metal ions on the silica gel, so that the target substance can be adsorbed on the stationary phase and eluted out.

[0069] Therefore, after comprehensive comparison of the above methods, it is finally determined to use the silica gel solid phase extraction column doped with palladium-silver for sample pretreatment by solid phase extraction, which has good purification effect, simple operation, can be directly sampled for analysis, and can meet the accurate determination requirements of target compounds in biofuel oil.

[0070] 2. Selection of sample extraction solvent Positive samples containing FAME and FAEE compounds were tested by using n-hexane, n-hexane / dichloromethane (4 / 1, V / V ), dichloromethane, dichloromethane / ethyl acetate (5 / 1, V / V ), dichloromethane / methanol (5 / 1, V / V ), ethyl acetate, acetone, methanol and other polar solvents from weak to strong for dissolution test. Except for the sample extraction solvent, the other contents of the test method are the same as in Example 1.

[0071] The experimental results show that the non-polar n-hexane cannot completely dissolve the sample, has low extraction efficiency for high-polarity esters, and is easy to cause the target to be wrapped by the precipitated gel and polar substances, affecting the accuracy of detection; the medium-polarity dichloromethane can completely dissolve the sample, but may also dissolve pigments or polymers, increasing the difficulty of subsequent purification; ethyl acetate, acetone and methanol are strong polar solvents, which have good extraction effect on FAME and FAEE compounds, but also dissolve polar substances such as ketones and phospholipid salts, which is not conducive to subsequent purification; the dichloromethane mixed solvent containing ethyl acetate and methanol has strong polarity, and the target is easy to be washed out too fast in the subsequent SPE column purification and concentration process, causing test error; the n-hexane mixed solvent containing dichloromethane effectively makes up for the defect of non-polarity of n-hexane, improves the coverage range of medium-polarity FAME / FAEE, and improves the extraction efficiency of polar esters, so the n-hexane / dichloromethane (4 / 1, V / V ) mixed solution is finally selected as the extraction solvent for marine biofuel oil samples.

[0072] 3. Selection of elution and elution conditions For the extraction of fatty acid esters in fuel oil, this study establishes an optimized pretreatment method by systematically comparing the extraction efficiency of different solvents. First, non-polar solvents are used for impurity elution: four solvents (n-pentane, n-hexane, cyclohexane and dichloromethane) with volume gradient 1.0-3.0 mL are tested for the purification effect of blank spiked samples. Except for the eluent, the other contents of the test method are the same as in Example 1. The results show that n-pentane has strong volatility and is easy to evaporate during the elution process, making it difficult to control the flow rate of the eluent; dichloromethane has poor selectivity, causing a large loss of target substances during the elution process, which in turn leads to a low recovery rate; n-hexane has good elution effect on non-polar impurities (such as saturated straight-chain or branched-chain alkanes) due to its non-polar nature, but has poor elution effect on polar impurities (such as aromatic hydrocarbons).

[0073] Cyclohexane has an appropriate boiling point and selectivity, and becomes the best eluent: it can effectively remove both non-polar impurities and polar impurities (removal rate > 95%), and can also retain the target fatty acid esters, and 2.0 mL of the eluent can achieve the desired purification effect.

[0074] In the elution stage, the elution efficiency of four mixed solvent systems on fatty acid esters was investigated: n-hexane / dichloromethane (4:1), dichloromethane / ethyl acetate (4:1), dichloromethane / ethanol (4:1), and dichloromethane / methanol (4:1). Except for the eluent, the other contents of the test method were the same as in Example 1. It was found that the dichloromethane / ethyl acetate (4:1) mixed solvent with moderate polarity performed best: when the amount was 2.5 mL, the recovery rate of C16-C18 fatty acid methyl esters reached more than 92%, and the solvent had good compatibility and did not cause phase separation. In contrast, the n-hexane / dichloromethane system was not completely eluted (recovery rate <70%), while the alcohol-containing systems (dichloromethane / ethanol (4:1) and dichloromethane / methanol (4:1)) had high recovery rates, but since the alcohol-containing solvents were not miscible with the cyclohexane eluent, the residual eluent in the extraction column was eluted into the eluent, which easily caused the eluent to separate into layers and not be homogeneous, thereby affecting the determination of the target substance and leading to low reproducibility of the test results. Therefore, 2.5 mL of dichloromethane / ethyl acetate (4:1) was finally selected as the elution solvent, and the method had good reproducibility and recovery rate.

[0075] 4. Injection mode selection Considering that the substrate of marine biofuel oil is relatively complex and contains a large amount of non-hydrocarbons, gum, asphaltene, etc., which may cause pollution to the analysis equipment or affect the analysis results, two ways of headspace direct injection and dilution direct injection were compared. The fatty acid methyl ester number positive samples were taken, and the headspace injection at 120°C and the direct injection after dilution were used. The results showed that the calculated fatty acid methyl ester content after the sample was heated in the headspace bottle and injected was only 10%-80% of the fatty acid methyl ester content when directly injected, as shown in Table 4; the recovery rate of the low carbon number fatty acid methyl esters was good, the recovery rate of C16 and above fatty acid methyl esters was only about 40-60%, and the greater the viscosity, the less the fatty acid methyl ester content obtained by headspace injection, and when the temperature exceeded 150°C, the content of unsaturated fatty acid methyl esters was less with the increase of temperature, as shown in Table 5. This may be related to the viscosity of the sample and the special nature of phenols in the fuel oil substrate. In a heated environment, compounds containing unsaturated groups may undergo polymerization or hydrolysis reactions, resulting in a decrease in concentration. Therefore, the dilution direct injection mode was selected. Figure 2

[0076] Table 4 Comparison of headspace injection and direct injection

[0077] 5. Vacuum distillation (vacuum distillation) ​Test 500.0 g (accurate to 0.01 g) of marine biofuel oil product in a 500 mL distillation apparatus, investigate the vacuum temperature of 200-300 ℃ and record the mass of evaporation, the extraction efficiency of fatty acid methyl ester is shown in Figure 3 It can be seen that as the vacuum temperature increases from 200 to 260 ℃, the extraction efficiency of the target product is significantly improved, and the recovery rate is basically unchanged at a temperature of 260 ℃. When the temperature continues to rise, the rate slows down, and the extraction recovery rate is basically unchanged, reaching equilibrium, and other high-temperature evaporation increases. Impurities increase the purification in the later stage, so 260 ℃ is selected as the vacuum distillation extraction temperature for the experiment, and the recovery rate is compared with the recovery rate without vacuum distillation treatment. The comparison results are shown in Table 5. Figure 3

[0078] Table 5 Comparison of recovery rates with and without vacuum distillation treatment

[0079] Overall, the recovery rates of both vacuum distillation and non-vacuum distillation are good, and the recovery rates of most target products after vacuum distillation are better. This may be due to the fact that during non-vacuum distillation, asphaltene and resin in marine biofuel oil are easily dissolved and constant volume, and the target product may be partially wrapped, resulting in lower recovery rate. Therefore, distillation cutting is selected as the pretreatment step of the method, and the distillation cutting temperature is 260-270 ℃.

[0080] 6. Selection of chromatographic column Several temperature-applicable and polar chromatographic columns such as conventional DB-5MS UI (30 m x 0.25 mm x 0.25 µm), DB-35MS UI (30 m x 0.25 mm x 0.25 µm), DB-5MS (60 m x 0.25 mm x 0.25 µm), HP-INNOWAX (60 m x 0.25 mm x 0.25 µm), and HP-INNOWAX (60 m x 0.32 mm x 0.50 µm) were selected for comparison. Except for the different chromatographic columns, the other contents of the test method were the same as in Example 1.

[0081] ​The separation effects of DB-5MS UI, DB-35MS UI and DB-5MS are all poor, especially the separation effects of methyl octadecanoate, methyl oleate and methyl linolenate are poor, and the above compounds cannot be qualitatively determined by gas chromatography. The above compounds can be well separated by selecting HP-INNOWAX. The results show that the target substances can be well separated on HP-INNOWAX (60m*0.25mm*0.25µm) and HP-INNOWAX (60m*0.32mm*0.50µm) chromatographic columns, and the HP-INNOWAX (60m*0.32mm*0.50µm) chromatographic column with a thicker film can make the homologous substances completely separated, therefore, the method selects HP-INNOWAX as the analysis chromatographic column, because it has comprehensive advantages in the target substance retention index (k'>2.0) and separation efficiency (R>1.8), and can meet the experimental requirements.

[0082] Obviously, the above embodiments of the present application are only examples for clearly illustrating the present application, and are not intended to limit the embodiments of the present application. Based on the above description, other different forms of changes or variations can be made by those skilled in the art. All the embodiments do not need to be exhausted here. Any modification, equivalent replacement and improvement made within the spirit and principle of the present application shall be included in the protection scope of the claims of the present application.

Claims

1. A method for detecting fatty acid methyl esters and fatty acid ethyl esters in biofuel oil, characterized in that, Includes the following steps: S1. Dissolve the biofuel oil in a solvent to obtain a solution, perform solid-phase extraction, and collect the eluent as the test solution; S2. Using the external standard method and direct injection, the solution to be tested described in step S1 is analyzed by gas chromatography-mass spectrometry, and the contents of fatty acid methyl esters and fatty acid ethyl esters in biofuel oil are calculated. In step S1, the solvent is a mixture of n-hexane and dichloromethane; the stationary phase of the solid-phase extraction column is a mixture of palladium salt, silver salt, alumina and silica gel; the eluent for solid-phase extraction is cyclohexane, and the eluent is a mixture of dichloromethane and ethyl acetate. In step S2, the stationary phase of the chromatographic column used in the gas chromatography analysis is polyethylene glycol, and the temperature rise of the gas chromatography is as follows: initial temperature 120℃, hold for 1 min, then rise to 200℃ at 20℃ / min, hold for 5 min, then rise to 260℃ at 10℃ / min, hold for 20 min.

2. The detection method according to claim 1, characterized in that, The mass ratio of palladium salt, silver salt, alumina and silica gel is 0.25:(0.2~0.3):(40~60):(18~22).

3. The detection method according to claim 1, characterized in that, The solid-phase extraction column is prepared by: Palladium salt, silver salt, alumina and silica gel are mixed to obtain a mixture; the mixture is then filled into a container to obtain a solid phase extraction column.

4. The detection method according to claim 1, characterized in that, Before dissolution as described in step S1, the process also includes vacuum distillation of the biofuel oil to collect light components.

5. The detection method according to claim 1, characterized in that, In the mixed solvent of dichloromethane and ethyl acetate, the volume ratio of dichloromethane to ethyl acetate is (3.8~4.2):

1.

6. The detection method according to claim 1, characterized in that, After collecting the eluent, the process further includes the steps of concentration, volume adjustment, and filtration.

7. The detection method according to claim 1, characterized in that, The conditions for gas chromatography analysis in step S2 are as follows: carrier gas: high-purity helium, column flow rate: 1 mL / min; injection mode: splitless injection, septum purge flow rate: 20 mL / min; injection port temperature: 260 ℃; injection volume: 1 μL; transfer line temperature: 260 ℃.

8. The detection method according to claim 1, characterized in that, The mass spectrometry detection mode of the gas chromatography-mass spectrometry analysis described in step S2 is selected ion mode.

9. The detection method according to claim 1, characterized in that, The fatty acid methyl ester is at least one of methyl octanoate, methyl decanoate, methyl dodecanoate, methyl tetradecanoate, methyl hexadecanoate, methyl palmitate, methyl octadecanoate, methyl oleate, methyl linoleate, methyl linolenic acid, methyl eicosanoate, methyl docosanoate, methyl erucic acid, and methyl tetracosanoate.

10. The detection method according to claim 1, characterized in that, The fatty acid ethyl ester is at least one of ethyl tetradecanoate, ethyl hexadecanoate, ethyl palmitate, ethyl heptadecanoate, ethyl octadecanoate, ethyl oleate, ethyl linoleate, ethyl linolenic acid, ethyl eicosanoate, ethyl 11-cis-eicosodeenoate, ethyl eicostrienoate (cis-11,14,17), ethyl arachidonicate, ethyl eicosapodeenoate, ethyl dodecanoate, ethyl erucic acid, ethyl dodecanosapodeenoate, ethyl dodecanohexaenoate, ethyl tridecanoate, ethyl tetradecanoate, and ethyl tetradecenoate.

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