Method for determining 14C in tubular combuste-CO2 direct absorption biological sample

Abstract

The invention relates to the field of element content determination, in particular to a method for determining 14C in a tubular combuste-CO2 direct absorption biological sample, which comprises the following steps: pretreating a sample; a proper amount of sample powder is weighed and placed in a tubular combustion furnace to be combusted, and the optimal combustion program of sample combustion is obtained; weighing the sample powder according to the optimal combustion procedure, putting the sample powder into a tubular combustion furnace, taking a liquid flash counting bottle, adding Carbo-Sorb E absorption liquid, accurately weighing, putting the liquid flash counting bottle into ice water, connecting with a gas outflow part of the tubular combustion furnace, combusting the sample, and weighing the counting bottle again after the sample is completely combusted; adding scintillation liquid into the counting bottle which completely absorbs CO2, oscillating and uniformly mixing, placing the counting bottle in a liquid scintillation counter for dark adaptation, and measuring the counting rate of the sample; and calculating the 14C activity concentration in the sample according to the sample counting rate. The determination method provided by the invention can accurately, quickly and efficiently determine the 14C in the biological sample, and provides a better choice for detecting the 14C in the biological sample.

Description

Technical Field

[0001] The present invention relates to the field of element content determination, and in particular to a method for determining the content of an element in a biological sample by direct absorption of CO2 using a tubular combustion method. 14 Method for determination of C. Background Art

[0002] Carbon-14 (Carbon-14, 14 C) is a radioactive isotope of carbon with an energy range of 0 ~ 156.5 keV and a half-life of 5730 years. 14 C is a natural radionuclide and also an artificial radionuclide. 14 Once C is produced, it is rapidly oxidized to 14 CO, which is then oxidized to 14 CO2, 14 CO2 is transported to the lower part of the atmosphere, absorbed by the hydrosphere, and incorporated into biological matter through photosynthesis, becoming part of the global carbon cycle. 14 C is a low-toxic radionuclide. Due to its low beta particle energy and weak penetrating ability, the external radiation effect can be ignored. However, due to its long half-life, high isotope exchange rate, and easy absorption and assimilation by biological organisms, 14 C can easily enter the food chain through the carbon cycle, causing continuous internal radiation hazards to human tissues and organs. 14 The long-term cumulative radiation dose of C cannot be ignored, as it can cause biological effects such as DNA breakage and gene mutation, which can have a potential impact on health. 14 The monitoring of C has important scientific significance and public health value.

[0003] Currently, biological samples 14 The separation of carbon (C) primarily relies on the combustion method. This method completely converts carbon (C) in biological samples into CO2 through an oxidation reaction under normal or high pressure conditions. Depending on the specific operation method, combustion methods can be further divided into two main types: oxygen bomb combustion and tube furnace combustion. The oxygen bomb combustion method offers advantages such as fast combustion speed, high carbon recovery rate, and simple operation. However, it suffers from poor safety and potential risks such as explosion caused by improper operation. The tube furnace combustion method also presents some challenges in the determination of carbon-14 in biological samples. For example, the generated CO2 must be converted to calcium carbonate using reagents such as NaOH, CaCl2, and NH4Cl for measurement. The preparation steps are cumbersome and time-consuming, only one sample can be separated at a time, and the sample separation time is long (13.5 hours).

[0004] Based on LSC measurements 14The sample preparation methods include calcium carbonate suspension method, direct absorption method and benzene synthesis method. The key step is to convert carbon in the sample into CO2 by acidification of inorganic compounds or oxidation of organic compounds (wet oxidation method or combustion method). The three preparation methods have advantages and disadvantages. The calcium carbonate suspension method is simple to operate, but has the following disadvantages: (1) when NH4Cl is used to adjust pH, the pH cannot be too low, otherwise Ca(OH)2 will be formed in addition to CaCO3 precipitation, and even CO2 will be lost, which will adversely affect the quality of CaCO3 powder and subsequent analysis; (2) because the added CaCl2 is a saturated solution, CaCO3 and Ca(OH)2 will be produced at the same time, which will cause quenching interference when measured by LSC, thereby causing errors; (3) when the sample source is prepared, CaCO3 powder and scintillation liquid are prepared to form a milky white gel, which is easy to be non-uniform and to settle, resulting in low counting efficiency and poor repeatability of counting; (4) the impurities of CaCO3 are the main factors causing quenching, so this preparation method is not suitable for samples with more impurities. Compared with the direct absorption method, the benzene synthesis method has higher sensitivity, but the conversion process is time-consuming and labor-intensive, the cost is high, and the waste produced is toxic.

[0005] In comparison, the direct absorption method is simple, safe and significantly reduces the cost, so the direct absorption method is the preferred preparation method. However, the current direct absorption method also has limitations: before operation, the sample needs to be oxidized and combusted (about 13.5 h), and then 250 mL of NaOH solution (5-15 g of sample) is used to absorb and separate the generated CO2; then ammonium chloride is added to adjust the pH value to 10-11.5, and then saturated calcium chloride solution is titrated until no white precipitate is generated (about 1-2 h). After filtering the white precipitate to convert it into calcium carbonate precipitate powder, 2 mol / L HCl solution (about 50-100 mL) is used to titrate the calcium carbonate powder, and the generated CO2 is captured in 9 mL of Carbo-Sorb E absorption liquid (about 0.5-1 h). The whole process takes about 13 h or more, and a large amount of reagents such as NaOH, CaCl2, NH4Cl and HCl are consumed, which does not meet the concept of green sample preparation; in addition, the method can only absorb and prepare one sample at a time, which is low in efficiency; finally, when the sample amount is small, the measurement accuracy will be affected, and heat will be generated during the absorption of CO2, which will reduce the capture and recovery rate of CO2.

[0006] Therefore, how to make the sample fully combust in a short time, and how to make the recovery rate of CO2 high, maximize the sample analysis amount, improve the counting efficiency and reduce the detection limit, are the main problems that need to be solved. SUMMARY

[0007] The present application aims to provide a tubular combustion-CO2 direct absorption method for determining the carbon content in biological samples. 14The determination method of C is an efficient and reliable online oxidation combustion, preparation and measurement analysis method to quickly analyze the content of C in different types of biological samples. 14 C, solve the above technical problems.

[0008] The technical solution of the present invention is as follows:

[0009] A tubular combustion-CO2 direct absorption method for biological samples 14 Methods for determining C include:

[0010] Sample pretreatment: freeze-dry the edible part and grind it into powder for later use;

[0011] Weigh an appropriate amount of sample powder and place it in a tubular combustion furnace for combustion to obtain the optimal combustion program for the sample, including sample combustion amount, heating rate, holding time, and gas flow rate;

[0012] According to the optimal combustion procedure, weigh the sample powder and place it in a tubular combustion furnace. Take a liquid scintillation counting bottle, add Carbo-Sorb E absorption liquid and accurately weigh it. Then, place it in ice water and connect it to the gas outlet of the tubular combustion furnace. Ventilate and purge the air in the device and burn the sample. After the sample is completely burned, wipe the moisture outside the counting bottle and weigh it again.

[0013] Add scintillation liquid to the counting bottle after CO2 absorption, shake and mix evenly, place in the liquid scintillation counter for dark adaptation, select 14 C measurement mode, measuring sample counting rate;

[0014] Calculate the sample count rate based on the sample 14 C activity concentration.

[0015] In a preferred embodiment of the present invention, the sample pretreatment, wherein the edible portion is freeze-dried and then ground into powder for later use, comprises:

[0016] Wipe the surface moisture of the edible part of the sample dry, cut into small pieces and grind evenly, weigh 1.50-5.00 g of the grinded sample onto the sample plate of the moisture analyzer, spread it evenly and measure its moisture content;

[0017] The remaining samples were canned and frozen in a -20°C refrigerator for 2 to 3 days, then placed in a vacuum freeze dryer for 4 to 5 days. After the samples were dried, they were ground into powder using a grinder. 1.00 to 2.00 mg of sample powder was weighed and its carbon content was determined by an elemental analyzer. The remaining sample powder was set aside.

[0018] In a preferred embodiment of the present invention, the process of weighing an appropriate amount of sample powder and placing it in a tubular combustion furnace to obtain an optimal combustion program for the sample includes:

[0019] A quantitative sample powder is weighed and placed in the sample area of ​​the tubular combustion furnace. A liquid scintillation counting bottle is taken, and Carbo-Sorb E absorption liquid is added and accurately weighed. The liquid is then placed in ice water and connected to the gas outflow of the tubular combustion furnace. An O2 / N2 mixed gas is introduced to the air in the scavenging device. Subsequently, under the catalysis of Pt-Al2O3, oxidative combustion is carried out according to the set combustion program to generate and collect CO2.

[0020] After the sample is completely burned, wipe the moisture outside the counting bottle and weigh it again. The C recovery rate is calculated based on the difference in mass before and after the counting bottle.

[0021] The effects of sample combustion volume, heating rate, holding time, and gas flow rate on C recovery were analyzed to determine the optimal combustion procedure for sample combustion.

[0022] In a preferred embodiment of the present invention, the set combustion program includes: the sample combustion amount is 2.00 g, 2.50 g, 3.00 g, 3.50 g, 4.00 g, 4.50 g, and 5.00 g, respectively; the heating rates are 5°C / min, 6°C / min, 7°C / min, 8°C / min, 9°C / min, and 10°C / min from 50°C to 500°C, respectively; the holding times are 0, 5, 15, and 30 min at 200°C, 300°C, and 400°C, respectively, and then increased to 800°C at a rate of 10°C / min; the gas flow rates are 0.10 L / min, 0.15 L / min, 0.20 L / min, 0.25 L / min, 0.30 L / min, 0.35 L / min, 0.40 L / min, and 0.45 L / min, respectively.

[0023] In the preferred embodiment of the present invention, the optimal combustion procedure for the sample combustion includes: the sample combustion amount is 2.50 g, the heating rate is 5 ° C / min from 50 ° C to 500 ° C, the holding time is 5 min, and the gas flow rate is 0.30 L / min.

[0024] In a preferred embodiment of this invention, 9 mL of Carbo-Sorb E absorption solution was added to a 20 mL liquid scintillation counting bottle and the mixture was aerated for 5 to 10 minutes.

[0025] In the preferred embodiment of this invention, 9 mL of Permafluor E was added to the counting bottle after CO2 absorption. + The scintillation fluid was mixed evenly and placed in a liquid scintillation counter to dark adapt for 12 h. 14 In C measurement mode, the energy window range is 0-156 keV and the measurement time is 300 min.

[0026] In the preferred embodiment of the present invention, the calculation of the number of samples in the sample according to the sample counting rate is as follows: 14 C activity concentrations include:

[0027] (1) Calculate LSC counting efficiency:

[0028]

[0029] in: For the instrument 14 Counting efficiency of C, %; for 14 C standard sample counting rate, min -1 ; for 14 C Background sample counting rate, min -1 ; To be added to the standard sample 14 Radioactivity of C, Bq;

[0030] (2) Calculate the sample 14 C activity concentration:

[0031]

[0032] in: For the sample 14 C activity concentration, Bq / kg fresh weight; for 14 C Sample counting rate, min -1 ; for 14 C Background sample counting rate, min -1 ; is the carbon content of the biological dry sample, %; is the moisture content of the sample, %; is the mass of carbon dioxide absorbed in the direct absorption method, g; For the instrument 14 Counting efficiency of C, %; is the mass fraction of carbon in carbon dioxide, %.

[0033] In the preferred embodiment of the present invention, the calculation of the number of samples in the sample according to the sample counting rate is as follows: 14 C activity concentration also includes:

[0034] (3) Calculation of minimum detectable activity:

[0035]

[0036] in: is the minimum detectable activity, Bq / g carbon; is the sample counting time, min.

[0037] In the preferred embodiment of this invention, the counting efficiency The minimum detectable activity is 87%. It is 0.01 Bq / g carbon.

[0038] In the preferred solution of this embodiment, uncertainty assessment is further included, specifically:

[0039] Combined uncertainty :

[0040]

[0041] in: is the combined uncertainty; ) is the uncertainty of instrument counting; (m) is the uncertainty of the mass of absorbed carbon dioxide; ( ) is the uncertainty of the instrument counting efficiency; (Y) is the uncertainty of the oxidation combustion recovery rate;

[0042] Relative standard uncertainty :

[0043]

[0044] in: is the relative standard uncertainty; For the sample 14 C activity concentration, Bq / kg fresh weight;

[0045] :

[0046]

[0047] in: To expand uncertainty; is 2.

[0048] The technical effects and advantages of the present invention: The present invention establishes an accurate, rapid and efficient tubular combustion-CO2 direct absorption combined analysis method for determining the content of CO2 in biological samples. 14 C activity concentration. Specifically, it can be better understood from at least the following aspects:

[0049] (1) The analytical method provided by the present invention achieves the best results under the following conditions by adopting an optimized online oxidation combustion-absorption program: sample combustion amount 2.50 g, heating rate 5℃ / min, holding time 5 min, gas flow rate 0.3L / min. The entire combustion-absorption process only takes 130 min, and the C recovery rate is 85.63%. This allows the sample to be fully burned in a relatively short time and a high C recovery rate can be obtained.

[0050] (2) The analytical method provided by the present invention is the first innovative method to combine a six-channel tubular combustion furnace to directly "absorb CO2 online" and prepare six samples at the same time, and then directly mix them with scintillation liquid and measure them using LSC. It is applicable to different types of seafood. 14 C analysis can maximize the sample analysis volume and achieve efficient multi-sample analysis in a short time (130 min). 14 The simultaneous analysis of C reduces the use of chemical reagents and conforms to the concept of green analysis.

[0051] (3) The analytical method provided by the present invention improves the counting efficiency (87%) and significantly reduces the detection limit (MDA is 0.01 Bq / g (carbon)). Compared with the calcium carbonate suspension method, the relative error range is within a reasonable range (less than 10%). It has good precision and accuracy and is suitable for the detection of different biological samples in emergency situations. 14 Detection of C.

[0052] (4) The analysis method provided by the present invention determines the overall 14 The biggest contributor to the uncertainty of C determination is the uncertainty of instrument counting, while the uncertainty of the mass of absorbed carbon dioxide can be ignored. Therefore, improving the counting efficiency and reducing the background level of the procedure to increase the sample counting rate is an effective way to reduce the counting uncertainty.

[0053] Other features and advantages of the present invention will be set forth in the description which follows, and in part will be obvious from the description, or may be learned by practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0054] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following is a brief introduction to the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0055] Figure 1 A schematic flow chart of a determination method provided in an embodiment of the present invention;

[0056] Figure 2 A principle diagram of a tubular combustion-CO2 direct absorption provided for an embodiment of the present application is shown in the figure;

[0057] Figure 3A A C recovery rate result diagram of different insulation time lengths provided for an embodiment of the present application (insulation time length of 30 min) is shown in the figure;

[0058] Figure 3B A C recovery rate result diagram of different insulation time lengths provided for an embodiment of the present application (insulation time length of 15 min) is shown in the figure;

[0059] Figure 3C A C recovery rate result diagram of different insulation time lengths provided for an embodiment of the present application (insulation time length of 5 min) is shown in the figure;

[0060] Figure 3D A C recovery rate result diagram of different insulation time lengths provided for an embodiment of the present application (insulation time length of 0 min) is shown in the figure;

[0061] Figure 4 A picture of a mixed scintillation liquid before being measured by an instrument provided for an embodiment of the present application is shown in the figure;

[0062] Figure 5 A fishbone diagram of uncertainty evaluation provided for an embodiment of the present application is shown in the figure. DETAILED DESCRIPTION

[0063] The technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the drawings in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments of the present application. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor fall within the scope of protection of the present application.

[0064] It should be noted that the structures, proportions, sizes, etc. shown in the drawings attached to the present specification are only used to cooperate with the content disclosed in the present specification, to enable those skilled in the art to understand and read, and are not used to limit the limiting conditions of the embodiments of the present application, so they do not have technical significance. Any modification of structure, change of proportion relationship or adjustment of size, without affecting the effects and purposes that can be achieved by the present application, should still fall within the scope of the technical content disclosed by the present application. At the same time, the terms such as "up", "down", "left", "right", "middle", "one", "first" and the like in the present specification are only for the convenience of clear understanding and description, and are not used to limit the scope of the embodiments of the present application. The change or adjustment of relative relationship, without substantially changing the technical content, is also regarded as the scope of the embodiments of the present application.

[0065] The present invention provides a tubular combustion-CO2 direct absorption biological sample 14 The determination method of C, such as Figure 1 The flowchart shown includes: the first step, sample pretreatment, taking the edible part, freeze-drying it, and then grinding it into powder for later use; the second step, weighing an appropriate amount of sample powder and placing it in a tubular combustion furnace for combustion to obtain the optimal combustion program for sample combustion, including the sample combustion amount, heating rate, insulation time and gas flow rate; the third step, weighing the sample powder according to the optimal combustion program and placing it in a tubular combustion furnace, taking a liquid scintillation counting bottle, adding Carbo-Sorb E absorption liquid and accurately weighing it, then placing it in ice water, connecting it to the gas outflow of the tubular combustion furnace, ventilating and removing the air in the device, and burning the sample. After the sample is completely burned, wipe off the moisture outside the counting bottle and weigh the counting bottle again; the fourth step, adding scintillation liquid to the counting bottle that has absorbed CO2, shaking and mixing it evenly, and placing it in a liquid scintillation counter for dark adaptation, selecting 14 C measurement mode, measure the sample counting rate; the fifth step is to calculate the sample count rate according to the sample counting rate. 14 The present invention establishes an accurate, rapid and efficient tubular combustion-CO2 direct absorption combined analysis method to determine the activity concentration of C in biological samples. 14 C activity concentration, which can achieve efficient multi-sample concentration in a short time (130 min). 14 The simultaneous analysis of C reduces the use of chemical reagents and conforms to the concept of green analysis. Compared with the calcium carbonate suspension method, the relative error range is within a reasonable range (less than 10%), with good precision and accuracy, and is suitable for different biological samples in emergency situations. 14 Detection of C.

[0066] In order to better understand the technical solutions of the present invention, specific embodiments are provided below.

[0067] 1. Main reagents and instruments

[0068] Carbo-Sorb E, Permafluor E + (PerkinElmer), Na2 14 CO3 standard solution (1069 Bq / g, China Institute of Metrology), Ca 14CO3 standard material (19.6 Bq / g, National Institute of Metrology, China), moisture analyzer (CYS-1.2, Shenzhen Finan Instrument Co., Ltd.), vacuum freeze dryer (4.5 L, -105 ℃, Labonco Corporation, U.S.A.), high-throughput tissue grinder (NB-YMY-48A, Nanbei Instrument Co., Ltd.), tube furnace oxidation system (Pyrolyser 6 Trio, Raddec International Ltd), elemental analyzer (Flash Smart, Thermoscientific), multi-parameter benchtop analyzer (S475, Mettler-Toledo Instruments (Shanghai) Co., Ltd.), low potassium glass counting bottle (PerkinElmer), balance (ME 204, Mettler-Toledo Instruments (Shanghai) Co., Ltd.), ultrapure water machine (UPT-I-5T, Sichuan Youpu Ultraclean Technology Co., Ltd.), low-background liquid scintillation spectrometer (Quantulus GCT 6220, PerkinElmer).

[0069] 2. Sample collection

[0070] Representative seafoods, including sea miscellaneous fish, sea shrimp, clam, sea crab and seaweed, were collected from local residents within 30 km of the Qinshan Nuclear Power Plant, and 3 kinds of terrestrial foods, including pomelo, peanut and duck egg, were also collected. Each sample was at least 1 kg. The specific sampling information is shown in Table 1.

[0071] Table 1. Sample collection information

[0072]

[0073] 3. Sample pretreatment

[0074] The samples were pretreated, and the edible parts were freeze-dried and ground into powder for use.

[0075] Specifically, after sample collection, the sea miscellaneous fish was scaled, gilled and gutted, the sea shrimp was shelled and had its head removed, the clam was washed to remove surface mud and had its shell removed to take the soft body part, the sea crab was shelled, and the seaweed was washed with ultrapure water to remove surface mud and mucus; the pomelo was peeled, the peanut and duck egg were shelled; after removing the inedible parts, the surface moisture of all samples was wiped dry, and the samples were cut into small pieces and stirred evenly with a stirring machine. 1.50 ~ 5.00 g of the stirred sample was weighed into a moisture analyzer sample pan and spread evenly to determine its moisture content. The remaining samples were placed in a -20 ℃ refrigerator for 2 ~ 3 days, then placed in a vacuum freeze dryer (-105 ℃, 0.1 mbar) for 4 ~ 5 days. After drying, the samples were ground into powder using a grinder, and 1.00 ~ 2.00 mg of the sample powder was weighed into an elemental analyzer to determine its carbon content. The remaining sample powder was reserved for use.

[0076] Here in the sample pretreatment section together with the determination of moisture content, carbon content, in subsequent calculation 14 C activity concentration will be used.

[0077] 4, determine the best combustion program

[0078] The appropriate amount of sample powder is placed in the tube furnace for combustion, and the best combustion program of sample combustion is studied, including sample combustion amount, heating rate, holding time and gas flow rate.

[0079] As shown in Figure 2 The tube furnace in this embodiment adopts a six-channel tube furnace, which is provided with a sample area and a catalytic area. The upstream is connected to the liquid scintillation counting bottle through the silica gel tube, and the downstream is connected to the liquid scintillation counting bottle through the silica gel tube.

[0080] Specifically, first take a 20 mL liquid scintillation counting bottle, add 9 mL Carbo-Sorb E absorption liquid and accurately weigh, then put it into ice water, connect with the gas outlet of the tube furnace, and ventilate for 5 ~ 10 min to remove the air in the device; Then take a quantitative sample powder and place it in the sample area of the tube furnace, and introduce O2 / N2 mixed gas (O2:N2=78:22), and under the catalysis of Pt-Al2O3, based on the set combustion program, the sample is oxidized and combusted to generate CO2, which is collected in the form of ions in the alkali absorption liquid (i.e. Carbo-Sorb E absorption liquid) by reacting with the alkali absorption liquid; After the sample is completely combusted, wipe off the water on the outside of the counting bottle, weigh the counting bottle again, and calculate the C recovery rate by the mass difference before and after the counting bottle; Analyze the influence of sample combustion amount, heating rate, holding time and gas flow rate on C recovery rate to determine the best combustion program of sample combustion.

[0081] In this embodiment, sea bream is taken as the research object, and the best oxidation combustion condition is screened. The sample combustion amount is 2.00 g, 2.50 g, 3.00 g, 3.50 g, 4.00 g, 4.50 g, 5.00 g, the heating rate is 5 ℃ / min, 6 ℃ / min, 7 ℃ / min, 8 ℃ / min, 9 ℃ / min and 10 ℃ / min, the holding time is 0, 5, 15 and 30 min at 200 ℃, 300 ℃ and 400 ℃, and then the temperature is raised to 800 ℃ at a rate of 10 ℃ / min, and the gas flow rate is 0.10 L / min, 0.15 L / min, 0.20 L / min, 0.25 L / min, 0.30 L / min, 0.35 L / min, 0.40 L / min and 0.45 L / min.

[0082] 4.1 Effect of sample combustion amount on recovery rate

[0083] The sample sampling amount, i.e. the combustion amount, was set to different gradients in this example, and the C recovery rate was compared, as shown in Table 2. When the combustion amount was ≤ 2.50 g, the C recovery rate was maintained at above 85%; with the increase of the sample amount, the C recovery rate showed a decreasing trend, and when the combustion amount was > 4.00 g, the C recovery rate tended to be stable, being 58.11% ~ 62.01%. During the reaction process, with the increase of the sample amount, the biological dry sample was not completely carbonized and ashed, and impurities were generated into the Carbo-Sorb E absorbent, which changed from transparent to yellow or even black, resulting in a quenching effect, which affected the repeatability and accuracy of the sample measurement results. Therefore, in order to make the sample fully react and obtain a higher C recovery rate, the sample combustion amount was selected to be 2.50 g in this example.

[0084] Table 2 C recovery rate results of different sample combustion amounts

[0085]

[0086] 4.2 Effect of heating rate on recovery rate

[0087] During the oxidation and combustion process of the biological sample 14 C will undergo carbonization and ashing processes during the oxidation and combustion process: below 400 ℃, it is mainly the carbonization process, and above 400 ℃, it is mainly the ashing process. If the carbonization process has a too fast heating rate, the possibility of deflagration and other situations will be higher, resulting in a lower C oxidation and combustion recovery rate. The change of the C recovery rate at different heating rates was analyzed in this example, as shown in Table 3. Although the oxidation and combustion time was shortened with the increase of the heating rate (from 206 min at 5 ℃ / min to 163 min at 10 ℃ / min), from the C oxidation and combustion recovery rate results, the recovery rate was the highest (94.35%) at 5 ℃ / min, and gradually decreased with the increase of the heating rate, and the recovery rate was the lowest (67.77%) at 10 ℃ / min. Therefore, the heating rate of the tube furnace from 50 ℃ to 500 ℃ was 5 ℃ / min in this example.

[0088] Table 3 C recovery rate results at different heating rates

[0089] 4.3 Effect of holding time on recovery rate

[0090] During the sample combustion process, the holding time of the sample has a crucial influence on the complete carbonization of the sample. The change of the C recovery rate at different holding times during the carbonization process was analyzed in this example, as shown in Table 4. When the holding time was 5 min, the C recovery rate was the highest (94.35%), and gradually decreased with the increase of the holding time, and when the holding time was 15 min, the C recovery rate was the lowest (67.77%). Therefore, the holding time of the sample during the carbonization process was 5 min in this example. Figure 3A 、 3BAs the holding time in the carbonization process was shortened (from 30 min to 0 min), the oxidation combustion time was also shortened (from 206 min to 106 min). When the holding time ranged from 5 min to 30 min, the C recovery rate ranged from 82.35% to 85.63%, and when there was no holding time, the C recovery rate was the lowest, only 79.39%. In this study, a holding time of 5 min was selected to achieve rapid oxidation combustion.

[0091] 4.4 Effect of gas flow rate on recovery rate

[0092] This example sets different gradients of gas flow rate to compare the C recovery rate, and the results are shown in Table 4. Under the condition of controlling other factors, the oxidation combustion recovery rate of C generally increases with the increase of gas flow rate. When the gas flow rate ranges from 0.10 mL / min to 0.35 mL / min, the recovery rate gradually increases, when the gas flow rate is 0.10 L / min, the oxidation combustion recovery rate of C is only 58.58%, when the gas flow rate is 0.30 L / min, the recovery rate can be as high as 90.78%, and when the gas flow rate is > 0.35 mL / min, the C recovery rate will decrease. Therefore, the gas flow rate selected in this example is 0.30 L / min.

[0093] Table 4 Oxidation combustion collection results of different gas flow rates

[0094]

[0095] 5. Combustion and absorption (sample source preparation)

[0096] According to the optimal combustion program, the sample powder was weighed and placed in the sample boat of the tube furnace, a liquid scintillation counting bottle was taken, Carbo-Sorb E absorption liquid was added and accurately weighed, then it was placed in ice water, connected with the gas outlet of the tube furnace, the air in the air purging device was removed, the sample was combusted, after the sample was completely combusted, the water on the outside of the counting bottle was wiped dry, and the counting bottle was weighed again.

[0097] Continuing to refer to Figure 2In this embodiment, according to the optimal combustion procedure, 2.50 g of sample powder was weighed and placed in a tubular combustion furnace. 9 mL of Carbo-Sorb E absorption liquid was added to a 20 mL liquid scintillation counting bottle and accurately weighed. The bottle was then placed in ice water (to prevent the absorption liquid from volatilizing and improve the CO2 absorption efficiency) and connected to the gas outflow of the tubular combustion furnace. An O2 / N2 mixed gas was introduced for 5 to 10 minutes to purge the air in the device. Subsequently, the sample was burned under Pt-Al2O3 catalysis (the catalyst makes the reaction more complete) according to the optimal combustion procedure, that is, the temperature of the tubular combustion furnace was increased from 50°C to 500°C at a rate of 5°C / min and kept at this temperature for 5 minutes. The gas flow rate was 0.30 L / min. After the sample was completely burned, the moisture outside the counting bottle was wiped dry and the counting bottle was weighed again.

[0098] The present invention also compared and studied the sample source preparation method using a calcium carbonate suspension method. Specifically, the absorption solution in the alkaline absorption bottle was transferred to a 500 mL conical flask, ammonium chloride was added, and the pH was adjusted to between 10.5 and 11.5. Then, a saturated CaCl2 solution was added and titrated until no white precipitate formed. The white precipitate was filtered, washed three times with 20 mL of deionized water and three times with anhydrous ethanol, and then dried in a 105°C oven to constant weight. After cooling, it was ground and used for later use.

[0099] 6. Sample measurement

[0100] Add scintillation liquid to the counting bottle after CO2 absorption, shake and mix evenly, place in the liquid scintillation counter for dark adaptation, select 14 C measurement mode, measuring sample counting rate;

[0101] Specifically, Figure 4 As shown, add 9 mL of Permafluor E directly into the CO2 counting bottle after absorption. + Scintillation fluid was mixed evenly and placed in a liquid scintillation counter for 12 hours of dark adaptation. 14 C measurement mode, energy window range: 0–156 keV. Measurement duration: 300 min.

[0102] The specific measurement method using calcium carbonate suspension method is as follows: weigh 2.0 g of CaCO3 powder sample, add 4 mL of deionized water and 14 mL of homemade scintillation fluid (made by using p-xylene instead of solvent, 1,4-bis(5-phenyloxazole)benzene instead of second scintillator, and mixing 2,5-diphenoxy and Triton X-100), shake and mix evenly, and place in liquid scintillation counter for dark adaptation for 12 hours. 14 In C measurement mode, the energy window range is 0 ~ 156 keV and the measurement time is 300 min.

[0103] 7.14 C activity concentration calculation

[0104] (1) Calculate LSC counting efficiency:

[0105]

[0106] Where: is the counting efficiency of the instrument for 14 C, %; is the counting rate of the 14 C standard sample, min -1 ; is the counting rate of the 14 C background sample, min -1 ; is the radioactivity of 14 C added to the standard sample, Bq;

[0107] (2) Calculate the 14 C activity concentration in the sample:

[0108] Using the direct absorption method:

[0109]

[0110] Where: is the 14 C activity concentration in the sample, Bq / kg (fresh weight); is the counting rate of the 14 C sample, min -1 , is the counting rate of the 14 C background sample, min -1 , both measured by a liquid scintillation counter; is the carbon content of the biological dry sample, %; is the water content of the sample, %, which has been measured during the previous sample pretreatment process; is the mass of absorbed carbon dioxide in the direct absorption method, g, which is obtained by weighing after combustion and absorption; is the counting efficiency of the instrument for 14 C, %; is the mass fraction of carbon in carbon dioxide, %;

[0111] Calcium carbonate suspension method:

[0112]

[0113] Where: is the 14 C activity concentration in the sample, Bq / kg (fresh weight); is the counting rate of the 14 C sample, min -1; For 14 C Background sample count rate, min -1 ; For 14 C Mass of CaCO3 measured, g For counting efficiency of instrument to 14 C, %; For mass fraction of carbon in calcium carbonate, %.

[0114] (3) Calculate the minimum detectable activity:

[0115] Direct absorption method:

[0116]

[0117] Where: Minimum detectable activity, Bq / g (carbon); Counting time of sample, min.

[0118] Calcium carbonate suspension method:

[0119]

[0120] Where: Minimum detectable activity, Bq / g (carbon); Background sample count rate, min -1 .

[0121] 8. Uncertainty evaluation

[0122] According to the calculation formula of 14 C, the main sources of uncertainty affecting the results include the uncertainty of instrument counting ), the uncertainty of absorbed carbon dioxide mass (m), the uncertainty of instrument counting efficiency ( ), and the uncertainty of oxidation combustion recovery rate (Y), as shown in Figure 5 , the final uncertainty is the relative standard uncertainty and the expanded uncertainty after synthesizing each component. The main contribution of instrument counting uncertainty is the statistical error of counting rate.

[0123] Synthetic uncertainty :

[0124]

[0125] Where: Uncertainty of instrument counting; Uncertainty of absorbed carbon dioxide mass (m); ( ) the uncertainty in the instrument count efficiency; (Y) the uncertainty in the oxidation combustion recovery; wherein

[0126] Uncertainty in instrument count :

[0127]

[0128] wherein: the uncertainty in the instrument count; the sample counting time, min -1 ; the background sample counting time, min -1 .

[0129] Uncertainty in the mass of carbon dioxide absorbed (m):

[0130]

[0131] wherein: (m) the uncertainty in the mass of carbon dioxide absorbed; the balance bias error; the repeatability bias; the mass of carbon dioxide absorbed, g.

[0132] Uncertainty in the instrument count efficiency ( ):

[0133]

[0134] wherein: ( ) the uncertainty in the instrument count efficiency; 0.010 is 14 the relative uncertainty of the C standard solution; the 14 C measurement repeatability uncertainty.

[0135] Uncertainty in the oxidation combustion recovery (Y) can be obtained by calculating the relative standard deviation of multiple repeated combustion recovery results.

[0136] Relative standard uncertainty :

[0137]

[0138] wherein: the relative standard uncertainty.

[0139] Expanded uncertainty :

[0140]

[0141] in: To expand uncertainty; is 2.

[0142] 9. Methodological Validation

[0143] 9.1 Precision Verification

[0144] In order to evaluate and verify the repeatability and applicability of the analytical method, the present invention respectively analyzed the content of 3 different types of food (pomelo, peanut and duck egg). 14 The C concentration was measured repeatedly, and the relative standard deviation (RSD) of the test results of different foods was calculated to quantitatively evaluate the precision of the method in different food matrices.

[0145] The results of processing and analyzing pomelo, peanuts and duck eggs are shown in Table 5. 14 The C activity concentrations were 21.29 Bq / kg (fresh weight), 146.18 Bq / kg (dry weight) and 57.05 Bq / kg (fresh weight), respectively. The RSD range of multiple repeated detection results was 3.69% ~ 7.46%, which was less than 10%, far lower than the requirements of the "Technical Specifications for Radiation Environment Monitoring HJ61-2021" (relative standard deviation not more than 30%), indicating that the method has good parallelism.

[0146] Pomelo and duck eggs 14 The MDA of C is 0.01 Bq / g (carbon) and 1.36 ~ 3.71 Bq / kg (fresh weight), respectively. 14 The MDA of C was 0.01 Bq / g (carbon) and 8.84 ~ 9.20 Bq / kg (dry weight), respectively. 14 The differences in C activity concentration and MDA may be related to the carbon content and material composition characteristics of the samples.

[0147] Table 5 Food 14 C test results parallel test results

[0148]

[0149] Note: a Peanut samples were dry samples, while pomelo and duck eggs were fresh samples. b The quality is based on carbon mass. c Quality is based on fresh weight.

[0150] 9.2 Accuracy Verification

[0151] In order to evaluate and verify the reliability of the analytical method, the present invention respectively analyzed the 5 different types of seafood (marine fish, shrimp, razor clams, crabs and seaweed) 14 The C concentration was measured repeatedly, and the standard deviation (SD) of the test results of different foods was calculated to quantitatively evaluate the accuracy of the method in different food matrices.

[0152] The background count of the direct absorption method established in the present invention is 1.75 min -1 , the counting efficiency is 87.00%, the MDA is 0.01Bq / g (carbon); and the background counting of the calcium carbonate suspension method is 1.91 min -1 , a counting efficiency of 75.00%, and an MDA of 0.03 Bq / g (carbon). The results are shown in Table 6. The direct absorption method developed in this invention, after sample purification, provides low LSC measurement background and high counting efficiency, approximately 12% higher than the calcium carbonate suspension method. This method also offers a low MDA. Compared to existing studies, the counting efficiency is higher (56.30% to 65.00%).

[0153] Table 6 Comparison of measurement results between direct absorption method and calcium carbonate suspension method

[0154]

[0155] The direct absorption method was used to measure the concentration of seafood around Qinshan Nuclear Power Station. 14 The C activity concentration ranged from 5.06 to 22.61 Bq / kg (fresh weight), which is lower than the standard (CAC / GL5-2006) issued by the Food and Agriculture Organization of the United Nations (FAO) / World Health Organization (WHO), which is the C activity concentration in food. 14 The guidance level of C is 1000 Bq / kg (infant food) and 10,000 Bq / kg (foods other than infant food). Seaweed has the lowest activity concentration, while razor clams have the highest activity concentration, as shown in Table 7. 14 The C activity concentration ranged from 5.56 to 23.08 Bq / kg (fresh weight). The radioactivity levels in food measured by the two methods were consistent, with relative errors ranging from 1.50% to 9.99%, both less than 10%, demonstrating the high accuracy of the direct absorption method established in this study.

[0156] In addition, the results of the direct absorption method and the calcium carbonate suspension method were consistent, and the relative error range was within a reasonable range, which further verified the reliability and feasibility of the direct absorption method. Compared with the calcium carbonate suspension method, the direct absorption method has higher calculation efficiency and is simple to operate, and can be used as a 14 An effective method for determining C activity.

[0157] Table 7: Foods in the vicinity of the Qinshan Nuclear Power Station 14 C activity concentration

[0158]

[0159] 10. Uncertainty results

[0160] The uncertainty analysis results are shown in Table 8. The uncertainty of instrument counting is the overall 14 The largest contributor to the uncertainty of the C determination is the uncertainty of the combustion recovery rate and the uncertainty of the counting efficiency, while the uncertainty associated with the weighing of the absorbed carbon dioxide mass is negligible. Therefore, increasing the sample counting rate by improving the counting efficiency and reducing the procedural background level is an effective way to reduce the counting uncertainty.

[0161] Table 8 Samples 14 Relative uncertainty components and combined uncertainty of C

[0162]

[0163] In summary, the present invention has established an accurate, rapid and efficient tubular combustion-CO2 direct absorption analysis method to determine the 14 For the first time, the innovative six-channel tubular combustion furnace is combined to directly "absorb CO2 online" while burning to prepare six samples at the same time, and then directly mix them with scintillation liquid and measure them with LSC, achieving efficient multi-sample concentration in a short time (130 minutes). 14 C, reducing the use of chemical reagents, in line with the concept of green analysis, and establishing 14 The C determination analysis method significantly reduced the MDA (0.01 Bq / g (carbon)) and improved the counting efficiency (87%). Compared with the calcium carbonate suspension method, the relative error range was within a reasonable range (less than 10%). It has good precision and accuracy and is suitable for emergency situations in different biological samples. 14 Detection of C.

[0164] Finally, it should be noted that the above only describes the preferred embodiments of the present application and is not intended to limit the present application. Although the present application has been described in detail with reference to the foregoing embodiments, those skilled in the art will appreciate that the technical solutions described in the foregoing embodiments can be modified or some technical features thereof can be replaced by equivalent features, and any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A tubular combustion-CO2 direct absorption method for biological samples 14 The method for measuring C is characterized in that include: Sample pretreatment: freeze-dry the edible part and grind it into powder for later use; Weigh an appropriate amount of sample powder and place it in a tubular combustion furnace for combustion to obtain the optimal combustion program for the sample, including sample combustion amount, heating rate, holding time, and gas flow rate; According to the optimal combustion procedure, weigh the sample powder and place it in a tubular combustion furnace. Take a liquid scintillation counting bottle, add Carbo-Sorb E absorption liquid and accurately weigh it. Then put it into ice water and connect it to the gas outflow of the tubular combustion furnace. Ventilate and purge the air in the device to burn the sample. After the sample is completely burned, wipe the moisture outside the counting bottle and weigh it again. Add scintillation liquid to the counting bottle after CO2 absorption, shake and mix evenly, place in the liquid scintillation counter for dark adaptation, select 14 C measurement mode, measuring sample counting rate; Calculate the sample count rate based on the sample count rate 14 C activity concentration.

2. The measuring method according to claim 1, wherein The sample pretreatment, taking the edible part and freeze-drying it and then grinding it into powder for later use, includes: Wipe the surface moisture of the edible part of the sample dry, cut into small pieces and grind evenly, weigh 1.50-5.00 g of the grinded sample onto the sample plate of the moisture analyzer, spread it evenly and measure its moisture content; The remaining samples were canned and frozen in a -20°C refrigerator for 2 to 3 days, then placed in a vacuum freeze dryer for 4 to 5 days. After the samples were dried, they were ground into powder using a grinder. 1.00 to 2.00 mg of sample powder was weighed and its carbon content was determined by an elemental analyzer. The remaining sample powder was set aside.

3. The measuring method according to claim 1, wherein The method of weighing an appropriate amount of sample powder and placing it in a tubular combustion furnace to obtain an optimal combustion program for the sample combustion includes: A quantitative sample powder is weighed and placed in the sample area of ​​the tubular combustion furnace. A liquid scintillation counting bottle is taken, and Carbo-Sorb E absorption liquid is added and accurately weighed. The liquid is then placed in ice water and connected to the gas outflow of the tubular combustion furnace. An O2 / N2 mixed gas is introduced to the air in the scavenging device. Subsequently, under the catalysis of Pt-Al2O3, oxidative combustion is carried out according to the set combustion program to generate and collect CO2. After the sample is completely burned, wipe the moisture outside the counting bottle and weigh it again. The C recovery rate is calculated based on the difference in mass before and after the counting bottle. The effects of sample combustion volume, heating rate, holding time, and gas flow rate on C recovery were analyzed to determine the optimal combustion procedure for sample combustion.

4. The measuring method according to claim 3, wherein The set combustion program includes: the sample combustion amounts are 2.00 g, 2.50 g, 3.00 g, 3.50 g, 4.00 g, 4.50 g, and 5.00 g, respectively; the heating rates are 5 ℃ / min, 6 ℃ / min, 7 ℃ / min, 8 ℃ / min, 9 ℃ / min, and 10 ℃ / min from 50 ℃ to 500 ℃, respectively; the holding times are 0, 5, 15, and 30 min at 200 ℃, 300 ℃, and 400 ℃, respectively, and then increased to 800 ℃ at a rate of 10 ℃ / min; the gas flow rates are 0.10 L / min, 0.15 L / min, 0.20 L / min, 0.25 L / min, 0.30 L / min, 0.35 L / min, 0.40 L / min, and 0.45 L / min, respectively.

5. The measuring method according to claim 4, wherein The optimal combustion procedure for the sample combustion includes: the sample combustion amount is 2.50 g, the temperature is increased from 50 °C to 500 °C at a rate of 5 °C / min, the holding time is 5 min, and the gas flow rate is 0.30 L / min.

6. The measuring method according to claim 1, wherein Take a 20 mL liquid scintillation counting bottle, add 9 mL Carbo-Sorb E absorption solution, and aerate for 5-10 minutes; Add 9 mL of Permafluor E to the counting bottle after CO2 absorption. + The scintillation fluid was mixed evenly and placed in a liquid scintillation counter to dark adapt for 12 h. 14 In C measurement mode, the energy window range is 0-156 keV and the measurement time is 300 min.

7. The measuring method according to claim 1, wherein The calculation of the sample count rate according to the sample count rate 14 C activity concentrations include: (1) Calculate LSC counting efficiency: ； in: For the instrument 14 Counting efficiency of C, %; for 14 C standard sample counting rate, min-1; for 14 C background sample counting rate, min-1; To be added to the standard sample 14 Radioactivity of C, Bq; (2) Calculate the sample 14 C activity concentration: ； in: For the sample 14 C activity concentration, Bq / kg fresh weight; for 14 C Sample counting rate, min -1 ; for 14 C Background sample counting rate, min -1 ; is the carbon content of the biological dry sample, %; is the moisture content of the sample, %; is the mass of carbon dioxide absorbed in the direct absorption method, g; For the instrument 14 Counting efficiency of C, %; is the mass fraction of carbon in carbon dioxide, %.

8. The measuring method according to claim 7, wherein The calculation of the sample count rate according to the sample count rate 14 C activity concentration also includes: (3) Calculation of minimum detectable activity: ； in: is the minimum detectable activity, Bq / g carbon; is the sample counting time, min.

9. The measuring method according to claim 8, characterized in that The counting efficiency The minimum detectable activity is 87%. It is 0.01 Bq / g carbon.

10. The measuring method according to any one of claims 1 to 9, characterized in that It also includes uncertainty assessment, specifically: Combined uncertainty : ； in: is the combined uncertainty; ) is the uncertainty of instrument counting; (m) is the uncertainty of the mass of absorbed carbon dioxide; ( ) is the uncertainty of the instrument counting efficiency; (Y) is the uncertainty of the oxidation combustion recovery rate; Relative standard uncertainty : ； in: is the relative standard uncertainty; is the 14C activity concentration in the sample, Bq / kg fresh weight; ： ； in: To expand uncertainty; is 2.

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