Phytolith extraction device and method

Abstract

The application relates to a phytolith extraction device and belongs to the technical field of plant extraction. The device comprises a charring device and a microwave digestion instrument. The charring device comprises a crucible body, a crucible cover and a temperature control electric furnace. A porous isolation plate parallel to the bottom surface of the crucible body is arranged in the cavity of the crucible body, and a gas inlet and a temperature control probe inlet are arranged on the side wall of the crucible body. The charring device and the microwave digestion instrument are combined, a ventilation temperature control charring-microwave digestion combined method is developed, the method realizes rapid extraction of sufficient phytolith from a plant body with low phytolith content, reduces the energy consumption of experiments and the waste of medicines in the experimental process, and increases the safety of the experimental process.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of plant extraction, and particularly relates to a phytolith extraction device and a method for extracting phytoliths using the same. BACKGROUND

[0002] Phytoliths are solid amorphous silica-containing particles formed by the precipitation of amorphous silicon (SiO2•nH2O) in the cells, cell walls, and intercellular spaces of plants after silicon in the form of soluble monosilicic acid (H4SiO4 or Si(OH)4) is absorbed by plant roots from the soil. The main components of phytoliths are SiO2 (75% - 95%), organic carbon (0.1% - 6%), water (3% - 12%), and a small amount of inorganic elements such as Na, K, Ca, Fe, Al, Ti, etc. In addition, the size of phytoliths varies greatly among different plants, generally ranging from 20 to 200 µm, with the smallest being about 2 µm and the largest being up to 2000 µm. The color of phytoliths under transmitted light ranges from colorless to black, with the most common being light red. Phytoliths have strong resistance to decomposition, corrosion, and high temperature, and can accumulate in soil and rocks and remain for thousands or even tens of thousands of years.

[0003] As a long-term stable form of carbon sequestration, phytolith carbon is an important part of forest carbon sink and cannot be ignored. In the process of phytolith carbon determination, the extraction of phytolith and the determination of organic carbon content in phytolith are the two most important steps in the determination of phytolith carbon. At present, the method for determining the organic carbon in phytolith has been mature, and through alkali dissolution spectrophotometry, the extremely small amount of organic carbon wrapped by phytolith can be detected. However, the content of phytolith in different plant types, different plant parts and different growth stages is different. For example, the phytolith of Gramineae plants is rich in the epidermal cells of leaves, and a small amount is in the palisade parenchyma cells and vessels. The phytolith of woody plants is mainly distributed in the vascular bundle, epidermal hair, base of epidermal hair and epidermal cells of leaves. However, the content of phytolith in most plants is extremely low, and it is very difficult to extract phytolith from these plants with extremely low content. The reason is that the commonly used method for extracting phytolith is microwave digestion method, which needs to control the sample amount to be less than 0.5g in a single digestion. Too much sample not only leads to incomplete digestion, but also causes safety accidents. Therefore, this method can only be used for plant samples with high phytolith content. For the case that the content of phytolith in most plants is not high, the solution is to repeat the microwave digestion for dozens of times or even hundreds of times, and then combine the digestion liquid to collect enough phytolith samples for testing. This extraction method has the following defects: (1) the operation process is extremely tedious and time-consuming; (2) a large amount of extraction reagents are consumed, and the equipment is greatly wasted; (3) the repeated extraction of a sample for dozens of times or even hundreds of times causes great experimental error, and the result is unreliable. SUMMARY

[0004] In view of the above defects in the background art, the purpose of the present application is to provide a method for quickly extracting sufficient amount of phytolith from plants with low phytolith content and to provide a device for extracting phytolith. The present application adopts the following technical solutions to achieve the purpose:

[0005] The phytolith extraction device comprises a microwave digestion instrument, characterized in that the device further comprises an ashing device, the ashing device comprises a crucible body, a crucible cover matched with the crucible body, and a temperature control electric furnace for heating the crucible body; a plurality of porous isolation plates parallel to the bottom surface of the crucible body are arranged in the cavity of the crucible body, a gas inlet and a temperature control probe inlet are respectively arranged on the side wall of the crucible body, and the gas inlet is arranged below the porous isolation plate; the temperature control electric furnace is provided with a temperature controller, and the temperature controller is matched with a temperature control probe; the temperature control probe extends into the cavity of the crucible body through the temperature control probe inlet; the device further comprises an air flow regulating valve, an air inlet pipe connected to the air outlet end of the air flow regulating valve, and a first pipe and a second pipe connected to the air inlet end of the air flow regulating valve; the air flowing through the first pipe and the carbon dioxide flowing through the second pipe are mixed in the air flow regulating valve and then flow into the cavity of the crucible body at a set flow rate through the air inlet pipe.

[0006] The phytolith extraction device, characterized in that the temperature control probe inlet is arranged above the porous isolation plate.

[0007] The phytolith extraction device, characterized in that a step is arranged on the inner wall of the lower part of the crucible body, and the size of the porous isolation plate matches the step.

[0008] The phytolith extraction device, characterized in that the crucible cover is provided with a gas permeable hole and a cover handle.

[0009] The phytolith extraction device, characterized in that the diameter of the holes on the porous isolation plate is 1-2 mm, preferably 1.70 mm.

[0010] The phytolith extraction device, characterized in that the crucible body is a circular truncated cone, the bottom surface has a circular diameter of 5.0-10.0 cm, the height of the circular truncated cone is 5.0-10.0 cm, and the volume is 100-785 cm 3 .

[0011] The phytolith extraction method, characterized in that the method is a combination of temperature control and microwave digestion, and specifically includes the following steps:

[0012] Step 1: Dry the collected plant samples in a forced air drying oven, crush them through a 10-mesh sieve, and store them for later use;

[0013] Step 2: Weigh the plant samples and place them in a clean container;

[0014] Step 3: Prepare the phytolith extraction device described above;

[0015] Step 4: Place the crucible body on the temperature control electric furnace and burn for 20 minutes, then cool it in a desiccator and weigh it. Weigh the crucible body and place it on the temperature control electric furnace;

[0016] Step 5: Introduce air and carbon dioxide mixed gas into the crucible body, and control the flow rate to be 1.5 L / min. As a preferred embodiment, the air and carbon dioxide are mixed in a volume ratio of 5:1-10:1;

[0017] Step 6: Add the plant samples containing phytoliths to be extracted into the crucible body, and the single addition amount should not exceed 100 g. Cover the crucible cover 2 and turn on the temperature control electric furnace to start heating;

[0018] Step 7: Turn on the temperature controller and set the critical heating temperature to 350℃. The temperature control probe measures the temperature in the crucible body in real time. When the temperature controller receives a temperature lower than the critical temperature from the temperature control probe, the temperature control electric furnace starts heating. When the temperature is higher than the critical temperature, the temperature control electric furnace stops heating, until the plant samples are completely smoldered, and the remaining combustion residues are obtained.

[0019] Step 8: Close the airflow regulating valve, turn off the temperature control furnace and temperature controller, and place the crucible containing the combustion residue in the desiccator to cool and weigh it again;

[0020] Step 9: Weigh 0.5000±0.0010g of the combustion residue into a microwave digestion tube, add 4.0mL of concentrated nitric acid and 0.5mL of concentrated hydrochloric acid, let stand for 20min, then cover the microwave digestion tube and place it in a microwave digestion apparatus for digestion. After digestion, digestion solution and phytolith sample are obtained. Preferably, the digestion parameters are: 1200W, 130℃, 15min.

[0021] Step 10: Transfer all the digestion solution and phytolith sample from the microwave digestion tube to a 50mL centrifuge tube, dilute with distilled water, centrifuge at 3000 rpm for 5 min, and discard the supernatant.

[0022] Step 11: Add water to repeatedly wash and centrifuge the phytolith sample until the pH of the supernatant is 6.5-7.5. After discarding the supernatant, place the extracted phytolith sample along with the centrifuge tube in an oven at 65°C to dry, and obtain the phytolith.

[0023] The beneficial effects of this invention are as follows: This invention provides a phytolith extraction device, consisting of an ashing device and a microwave digester. The microwave digester utilizes existing technology, while this invention designs an ashing device. By combining the ashing device and the microwave digester, a combined method of aeration-temperature controlled ashing-microwave digestion is developed. Specifically, the ashing process employs aeration-temperature controlled method, followed by microwave digestion for phytolith extraction. This method can accurately, rapidly, and effectively extract plant phytoliths, regardless of the phytolith content in the plant sample. Compared to traditional microwave digestion methods, this method reduces experimental energy consumption, increases experimental safety, and lowers the limits of phytolith extraction and detection in plant samples, making accurate determination of phytolith carbon content in plants with low phytolith content possible. Furthermore, a sufficient amount of phytoliths can be extracted in a single operation, reducing the cumbersome nature of multiple extractions in traditional methods and minimizing waste of reagents and wear and tear on equipment during the experiment. Attached Figure Description

[0024] Figure 1 This is a schematic diagram of the ashing device of the present invention;

[0025] Figure 2 This is a schematic diagram of the structure of the crucible body and the crucible lid of the present invention.

[0026] Figure 3 This is a schematic diagram of the crucible body of the present invention without the porous isolation plate assembled;

[0027] Figure 4 This is a schematic diagram of the ashing device of the present invention in use;

[0028] Figure 5 This is a schematic diagram of the structure of device two in Example 3;

[0029] Figure 6 Linearity graphs showing the results of the combined ventilation-temperature controlled ashing-microwave digestion method and the microwave digestion method;

[0030] In the diagram: 1-Crucible body; 1-1-Porous isolation plate; 1-2-Gas inlet; 1-3-Temperature control probe inlet; 1-4-Step; 2-Crucible lid; 2-1-Vent hole; 2-2-Lid handle; 3-Temperature-controlled electric furnace; 4-Temperature controller; 5-Temperature control probe; 6-Inlet duct; 7-Airflow regulating valve; 8-First duct; 9-Second duct; 10-Compressed air cylinder; 11-Compressed carbon dioxide cylinder. Detailed Implementation

[0031] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments.

[0032] Example 1: Design of Phytolith Extraction Device

[0033] This invention develops and designs a silica extraction device that combines an ashing device and a microwave digester. The microwave digester is existing technology. The structure of the ashing device is described below. Figure 1 The ashing device is an improvement on existing crucibles used to hold plant material. The crucible mainly consists of a crucible body 1 and a crucible lid 2 that matches the crucible body 1 (see...). Figure 1 , Figure 2 The crucible is constructed as follows: the crucible body 1 is preferably truncated cone-shaped, and the crucible lid 2 is provided with a vent hole 2-1 and a lid handle 2-2; unlike existing crucibles, the crucible body 1 is provided with a porous partition plate 1-1, which has several evenly distributed circular through holes, preferably 5-10 holes / cm. 2 The principle of setting the aperture is to allow air to circulate freely but to prevent most of the plant samples from falling from the porous isolation plate 1-1 to the bottom surface of the crucible body 1. Preferably, the aperture of the holes on the porous isolation plate is set to 1.0-2.0 mm, preferably 1.7 mm; the porous isolation plate 1-1 is parallel to the bottom surface of the crucible body 1.

[0034] Ashing is the process by which plant samples undergo smoldering. Completely ashed plant samples are often grayish-white powder. When using a regular crucible to ashed plant samples, the sample amount should be controlled to within 20g. Otherwise, if an excessive amount of sample is processed at once, the crushed plant sample will easily accumulate due to the lack of air circulation inside the crucible. The surface will burn violently while the internal combustion will be incomplete and uneven, which can easily lead to oxygen-deficient carbonization. Samples carbonized in oxygen-deficient conditions have strong stability, which often makes it impossible to successfully extract phytoliths later. Therefore, when performing ashing treatment on plant samples, the sample amount should be strictly controlled to avoid carbonization. To achieve the ashing of more plant samples in a single operation and avoid carbonization, this invention provides a gas inlet 1-2 and a temperature control probe inlet 1-3 on the side wall of the crucible body 1. The gas inlet 1-2 is preferably located below the porous isolation plate 1-1 to introduce low-oxygen air. The introduction of low-oxygen air increases the contact area between the air and the plant sample surface, preventing carbonization due to oxygen deficiency. Furthermore, the low oxygen content avoids the risk of violent combustion and deflagration. The temperature control probe inlet 1-3 is designed to monitor the ashing temperature of the plant samples inside the crucible body 1. Therefore, this invention also includes a temperature-controlled electric furnace 3 for heating the crucible body 1. The temperature-controlled electric furnace 3 is equipped with a temperature controller 4, and a temperature control probe 5 is also installed in conjunction with the temperature controller 4. The temperature control probe extends into the cavity of the crucible body 1 through the temperature control probe inlet 1-3 to receive temperature signals and transmit them to the temperature controller 4. When the temperature is higher than the set temperature, a command is issued to stop heating in the temperature-controlled electric furnace 3; when the temperature is lower than the set temperature, the temperature-controlled electric furnace 3 starts heating. To introduce air with the required oxygen content, this invention also includes an air inlet duct 6, an airflow regulating valve 7, a first duct 8, and a second duct 9. The air inlet duct 6 passes through the gas inlet 1-2 and enters the central space between the bottom surface of the crucible body 1 and the porous isolation plate 1-1. The outlet end of the airflow regulating valve 7 is connected to the air inlet duct 6. The airflow regulating valve 7 has two inlet ends, which are respectively connected to the first duct 8 and the second duct 9 (see...). Figure 1 The air flowing through the first conduit 8 and the carbon dioxide flowing through the second conduit 9 are mixed in the airflow regulating valve 7 and then flow into the crucible body 1 through the air inlet conduit 6 at a set flow rate. The mixture is then diffused between the plant sample powders through the porous partition plate 1-1, avoiding carbonization caused by insufficient oxygen content during plant sample ashing. In use, the first conduit 8 is connected to the compressed air cylinder 10, and the second conduit 9 is connected to the compressed carbon dioxide cylinder 11 (see...). Figure 4 The compressed air cylinder 10 and the compressed carbon dioxide cylinder 11 contain air and carbon dioxide, respectively. The compressed air cylinder 10, the compressed carbon dioxide cylinder 11, and the flow rate control are all existing technologies and will not be described in detail here.

[0035] To prevent plant samples from falling from the contact point between the porous separator 1-1 and the inner wall of the crucible body to the bottom of the crucible body 1, this invention provides a ring of stepped edges 1-4 (see...) on the lower inner sidewall of the crucible body 1. Figure 3 The size of the porous partition plate 1-1 is matched with that of the step 1-4, which enables the porous partition plate 1-1 to be better positioned and avoids the consequences of uneven placement of the porous partition plate or excessive gaps at the contact points, which would cause a large number of plant samples to fall to the bottom of the crucible body 1.

[0036] Preferably, the diameter of the bottom circle of the crucible body 1 is 5.0-10.0 cm, the height of the frustum is 5.0-10.0 cm, and the volume is 100-785 cm³. 3 .

[0037] Example 2: Extraction of phytoliths using a combined method of ventilation-controlled temperature ashing and microwave digestion

[0038] This invention provides a method for extracting phytoliths using a combination of ventilation-controlled temperature ashing and microwave digestion. The specific steps are as follows:

[0039] Step 1: Place the collected plant samples in a 65℃ forced-air drying oven to dry, then pulverize them and pass them through a 10-mesh sieve before storing them for later use.

[0040] Step 2: Weigh a sufficient amount of plant sample and place it in a clean container. Record the sample weight as M1.

[0041] Step 3: Prepare the phytolith extraction device from Example 1;

[0042] Step 4: Place the crucible (including the crucible body 1, porous partition plate 1-1, and crucible lid 2) on a temperature-controlled electric furnace 3 and heat it at a high temperature (above 400℃) for 20 minutes. Then place it in a desiccator to cool and weigh it. After weighing, place the crucible body 1 on the temperature-controlled electric furnace 3 and press... Figure 4 Connect the ashing device to the compressed air cylinder 10 and the compressed carbon dioxide cylinder 11;

[0043] Step 5: Introduce the air and carbon dioxide mixture into the crucible body and control the inlet flow rate to 1.5 L / min. Specifically, loosen the valves of the compressed air cylinder 10 and the compressed carbon dioxide cylinder 11 and adjust the flow rate to mix the air and carbon dioxide at a volume ratio of 5:1 to 10:1. Adjust the airflow regulating valve 7 to control the flow rate into the inlet duct 6 to 1.5 L / min.

[0044] Step 6: Add the plant sample from which the phytoliths to be extracted into the crucible body 1. The amount added at one time shall not exceed 100g. Cover the crucible with the lid 2 and turn on the temperature-controlled electric furnace 3 to continue heating until the sample begins to burn.

[0045] Step 7: Turn on the temperature controller 4 and set the critical heating temperature to 350℃. The temperature control probe 5 measures the temperature inside the crucible body 1 in real time. When the temperature controller 4 receives a temperature from the temperature control probe 5 that is lower than the critical temperature, it controls the temperature-controlled furnace 3 to start heating. When the temperature is higher than the critical temperature, it triggers the temperature-controlled furnace 3 to stop heating until the sample is completely smoldering. Then, plant samples are added one by one until all the plant samples weighed in step 2 are added and completely smoldering, and the combustion residue is obtained.

[0046] Step 8: Close the airflow regulating valve 7, turn off the temperature control furnace 3 and the temperature controller 4, place the crucible containing the combustion residue in the desiccator to cool and weigh it again, calculate the weight of the sample combustion residue, and record it as M2;

[0047] Step 9: Weigh approximately 0.5000g of the combustion residue (denoted as M3) into a microwave digestion tube, add 4.0mL of concentrated nitric acid and 0.5mL of concentrated hydrochloric acid, let stand for 20 minutes, then cap the microwave digestion tube and place it in a microwave digester for digestion. The digestion parameters are: 1200W, 130℃, 15min. After digestion, the digestion solution and phytolith sample are obtained.

[0048] Step 10: Transfer all the digestion solution and phytolith sample from the microwave digestion tube to a 50ml centrifuge tube, dilute with distilled water, centrifuge at 3000 rpm for 5 min, and discard the supernatant.

[0049] Step 11: Wash the phytolith sample with water, centrifuge, and test the pH of the supernatant with pH test paper until the pH of the supernatant is neutral or close to neutral (pH range of 6.5-7.5). Discard the supernatant and place the extracted phytolith sample along with the centrifuge tube in a 65℃ oven to dry. Weigh the dried phytolith sample and record the weight as M4. Calculate the phytolith content (g / kg) in the plant sample using the following formula:

[0050]

[0051] Example 3: Comparison of the effects of different ashing devices on phytolith extraction

[0052] Apparatus 1: The ashing apparatus described in Example 1 is used. The crucible body 1 has the following specifications: bottom diameter 8cm, frustum height 7.0cm, and volume 350cm³. 3 The device is as follows Figure 4 As shown in the figure. The method for extracting the phytolith using device one is described in Example 2, wherein the amount added in step 6 varies, and is either 20g (added in 5 portions) or 100g (added all at once), as shown in Table 1.

[0053] Apparatus 2: Uses only the crucible from the ashing apparatus in Example 1 (crucible body specifications are the same as Apparatus 1), but differs from Apparatus 1 in that: no porous isolation plate is installed, and gas inlets 1-2 are blocked. Figure 5 As shown. The method for extracting the implant using device two is as follows:

[0054] Step 1: Place the collected plant samples in a 65℃ forced-air drying oven to dry, then pulverize them and pass them through a 10-mesh sieve before storing them for later use.

[0055] Step 2: Weigh a sufficient amount of plant sample and place it in a clean container. Record the sample weight as M1.

[0056] Step 3: Prepare the silicon extraction device, wherein the ashing device adopts... Figure 5 The structure shown;

[0057] Step 4: Place the crucible (including the crucible body 1 and the crucible lid 2) on a temperature-controlled electric furnace 3 and heat it at a high temperature (above 400°C) for 20 minutes. Then, place it in a desiccator to cool and weigh it. After weighing, place the crucible body 1 on the temperature-controlled electric furnace 3 and press... Figure 5 Connect the temperature-controlled electric furnace 3, temperature control probe 5, and other components to the crucible body 1;

[0058] Step 5: Add the plant sample from which the phytoliths to be extracted into the crucible body 1. The amount added at one time is 20g (added in 5 portions) or 100g (added all at once), as shown in Table 1; cover the crucible with lid 2 and turn on the temperature-controlled electric furnace 3 to heat until the sample begins to burn.

[0059] Step 6: Turn on the temperature controller 4 and set the critical heating temperature to 350℃. The temperature control probe 5 measures the temperature inside the crucible body 1 in real time. When the temperature controller 4 receives a temperature from the temperature control probe 5 that is lower than the critical temperature, it controls the temperature-controlled furnace 3 to start heating. When the temperature is higher than the critical temperature, it triggers the temperature-controlled furnace 3 to stop heating until the sample is ashed. Then, plant samples are added one by one until all the plant samples weighed in step 2 are added and ashed, and the combustion residue is obtained.

[0060] Step 7: Turn off the temperature-controlled furnace 3 and the temperature controller 4, place the crucible containing the combustion residue in the desiccator to cool and weigh it again, calculate the weight of the sample combustion residue, and record it as M2;

[0061] Step 8: Weigh approximately 0.5000 g (0.5000 ± 0.0010 g) of the combustion residue (denoted as M3) into a microwave digestion tube, add 4.0 mL of concentrated nitric acid and 0.5 mL of concentrated hydrochloric acid, let stand for 20 min, then cap the microwave digestion tube and place it in a microwave digester for digestion. The digestion parameters are: 1200 W, 130 °C, 15 min. After digestion, the digestion solution and phytolith sample are obtained.

[0062] Step 9: Transfer all the digestion solution and phytolith sample from the microwave digestion tube to a 50ml centrifuge tube, dilute with distilled water, centrifuge at 3000 rpm for 5 min, and discard the supernatant.

[0063] Step 10: Wash the phytolith sample with water, centrifuge, and test the pH of the supernatant with pH test paper until the pH of the supernatant is neutral or close to neutral (pH range of 6.5-7.5). Discard the supernatant and place the extracted phytolith sample along with the centrifuge tube in a 65℃ oven to dry. Weigh the dried phytolith sample and record the weight as M4. Calculate the phytolith content (g / kg) in the plant sample using the following formula:

[0064]

[0065] Two types of plant samples were used: banyan leaves and banyan trunks, with a total sample weight of 100g for each sample. The weight added to the ashing device was either 20g (added in 5 separate additions) or 100g (added all at once). The extraction efficiency of phytoliths using different ashing devices is shown in Table 1. The results show that device one did not cause carbonization when ashing the plant samples, while device two caused carbonization even with a single sample addition of 20g; when the single sample addition was 100g, carbonization was more severe, reaching 15-20%. Because samples carbonized in an oxygen-deficient environment have strong stability, microwave digestion of the carbonized combustion residue failed, leading to phytolith extraction failure. Therefore, carbonization must be avoided. Device one ensured that carbonization did not occur when adding no more than 100g of sample at a time, guaranteeing the reliability of the experimental results.

[0066] Table 1. Effects of different ashing devices on phytolith extraction

[0067]

[0068] Example 4: Extraction of phytoliths from different plant samples

[0069] Using the device from Example 3 ( Figure 4 Sixteen different plant material samples were ashed, and the amount and content of combustion residue for different plant samples are shown in Table 2. The results showed that the content of combustion residue varied among different plant materials, ranging from 0.33 to 30.93 g / kg.

[0070] Table 2. Amount and content of combustion residues from different plant samples

[0071]

[0072] After ashing, approximately 0.5000 g of the combustion residue was weighed into a microwave digestion tube, 4.0 mL of concentrated nitric acid and 0.5 mL of concentrated hydrochloric acid were added, and after standing for 20 min, the microwave digestion tube was capped and placed in a microwave digester for digestion. The results are shown in Table 3. The results show that phytoliths can be extracted by using the combined method of aeration-temperature controlled ashing-microwave digestion in one microwave digestion.

[0073] To verify the reliability of the results, the phytolith content of the same sample was simultaneously determined using the traditional microwave digestion method for all 16 samples mentioned above. The method was as follows: 0.5 g of plant sample was weighed, and 4.0 mL of concentrated nitric acid and 0.5 mL of concentrated hydrochloric acid were added. After standing for 20 minutes, the microwave digestion tube was capped and placed in a microwave digester for digestion. For samples with low phytolith content, multiple digestions (1-5 times) were performed, and the digestion solutions were combined and the phytoliths collected. If no phytolith solids were observed after combining 5 digestion solutions, the sample was considered undetectable. The results of the two methods were compared and are shown in Table 3. Figure 6 The results of measurements on different organs of different plants showed that traditional microwave digestion methods cannot accurately determine the phytolith content in samples of banyan, eucalyptus, Masson pine, camphor, and Chinese fir trunks. The combined method of aeration-controlled temperature ashing-microwave digestion can accurately and rapidly determine the phytolith content in different plants, and can be applied to the analysis of phytolith content in various plant samples, with relatively high measurement accuracy. While the single-sample digestion volume of microwave digestion alone is around 0.5g, the combined method of aeration-controlled temperature ashing-microwave digestion can increase the single-sample digestion volume to 150.8g or even higher, an increase of more than 300 times. This significantly reduces the workload of low-phytolith extraction, lowers the extraction and detection limits of phytoliths in plant samples, and makes it possible to determine the carbon content of phytoliths in plants with low phytolith content.

[0074] The results, compared with those obtained by the commonly used microwave digestion method, show that the phytolith content determined by the two methods is numerically comparable, with no significant systematic difference. A t-test (Table 3) was performed on the results of the two methods, yielding t = -1.27. Furthermore, when the degrees of freedom v = 10, the table shows t = 2.23 at the 5% level. <t 0.05 At the 1% level, t = 3.17, |t| < t 0.01 The results from the two methods showed no significant difference at the 1% and 5% levels. Linear correlation analysis was performed on the results of 16 different phytolith samples measured by the two methods (n = 11, R0). 2 = 0.9999 ( Figure 6The results showed a highly significant prior correlation, indicating that this method can reliably and accurately detect the phytolith content in different plants. Therefore, the results of determining the phytolith content in plants using this method are accurate and reliable.

[0075] Table 3. Results of phytolith content determination in 16 samples using different methods

[0076]

[0077] Based on the phytolith content results of different plant samples in Table 3, the number of microwave digestions and the volume of solvent used in the method of this invention (aeration-temperature controlled ashing-microwave digestion combined method) were calculated compared with the traditional microwave digestion method. The results are shown in Table 4. Clearly, the method of this invention significantly reduces the number of microwave digestions and greatly shortens the sample extraction time. The significantly reduced number of microwave digestions saves a large amount of extraction reagents, while also reducing experimental energy consumption and increasing the safety of the experimental process.

[0078] Table 4. Number of microwave digestion cycles and solvent volumes used for extracting 0.005 g of phytoliths using different methods.

[0079]

Claims

1. A method for extracting phytoliths, characterized in that: The method is a combined ventilation-temperature controlled ashing-microwave digestion method, including the following steps: Step 1: Place the collected plant samples in a forced-air drying oven to dry, then pulverize them and pass them through a 10-mesh sieve before storing them for later use. Step 2: Weigh the plant sample and place it in a clean container; Step 3: Prepare the phytolith extraction device; The aforementioned phytolith extraction device includes a microwave digester, characterized in that: the device further includes an ashing device, the ashing device including a crucible body (1), a crucible cover (2) that cooperates with the crucible body, and a temperature-controlled electric furnace (3) for heating the crucible body; the crucible body (1) is provided with a porous isolation plate (1-1) parallel to the bottom surface of the crucible body, the porous isolation plate (1-1) having a pore diameter of 1.0~2.0 mm; a gas inlet (1-2) and a temperature control probe inlet (1-3) are respectively provided on the side wall of the crucible body (1), the gas inlet (1-2) being located below the porous isolation plate for introducing a mixed gas of air and carbon dioxide, the volume ratio of air to carbon dioxide in the mixed gas being 5:1~10:1, and the flow rate being 1.5 L. / min; The temperature-controlled electric furnace (3) is equipped with a temperature controller (4) for setting the critical heating temperature to 350℃. The temperature controller (4) is equipped with a temperature control probe (5). The temperature control probe extends into the cavity of the crucible body (1) through the temperature control probe inlet (1-3). The device also includes an airflow regulating valve (7), an air inlet pipe (6) connected to the air outlet of the airflow regulating valve (7), and a first pipe (8) and a second pipe (9) connected to the air inlet of the airflow regulating valve (7). The air flowing through the first pipe (8) and the carbon dioxide flowing through the second pipe (9) are mixed in the airflow regulating valve (7) and then flow into the cavity of the crucible body (1) through the air inlet pipe (6) at a set flow rate. Step 4: Place the crucible body on a temperature-controlled electric furnace and heat for 20 minutes, then place it in a desiccator to cool and weigh it. After weighing, place the crucible body on the temperature-controlled electric furnace. Step 5: Introduce a mixture of air and carbon dioxide into the crucible body, controlling the inlet flow rate to be 1.5 L / min; Step 6: Add the plant sample from which the phytoliths are to be extracted into the crucible body. The amount added at one time shall not exceed 100 g. Cover the crucible with the lid and turn on the temperature-controlled electric furnace to heat it continuously until it starts to burn. Step 7: Turn on the temperature controller and set the critical heating temperature to 350℃. The temperature control probe measures the temperature inside the crucible in real time. When the temperature received by the temperature control probe is lower than the critical temperature, the temperature control furnace will start heating. When the temperature is higher than the critical temperature, the temperature control furnace will stop heating until the plant sample is completely smoldering and the combustion residue is obtained. Step 8: Close the airflow regulating valve, turn off the temperature control furnace and temperature controller, and place the crucible containing the combustion residue in the desiccator to cool and weigh it again; Step 9: Weigh 0.5000±0.0010 g of the combustion residue into a microwave digestion tube, add 4.0 mL of concentrated nitric acid and 0.5 mL of concentrated hydrochloric acid, let stand for 20 min, then cover the microwave digestion tube and place it in a microwave digestion apparatus for digestion. After digestion, digestion solution and phytolith sample are obtained. Step 10: Transfer all the digestion solution and phytolith sample from the microwave digestion tube to a 50 mL centrifuge tube, dilute with distilled water, centrifuge at 3000 rpm for 5 min, and discard the supernatant. Step 11: Add water to repeatedly wash and centrifuge the phytolith sample until the pH of the supernatant is 6.5-7.

5. After discarding the supernatant, place the extracted phytolith sample together with the centrifuge tube in an oven at 65°C to dry, and obtain the phytolith.

2. The method for extracting phytoliths according to claim 1, characterized in that: The temperature control probe inlet (1-3) mentioned in step 3 is located above the porous isolation plate.

3. The method for extracting phytoliths according to claim 1, characterized in that: In step 3, a ring of steps (1-4) is provided on the lower inner side wall of the crucible body (1), and the size of the porous isolation plate (1-1) matches that of the steps (1-4).

4. The method for extracting phytoliths according to claim 1, characterized in that: The crucible lid (2) mentioned in step 3 is provided with a vent hole (2-1) and a lid handle (2-2).

5. The method for extracting phytoliths according to claim 1, characterized in that: The holes on the porous isolation plate (1-1) in step 3 have a diameter of 1.70 mm.

6. The method for extracting phytoliths according to claim 1, characterized in that: The crucible body (1) mentioned in step 3 is frustum-shaped, with a base diameter of 5.0-10.0 cm, a height of 5.0-10.0 cm, and a volume of 100-785 cm³. 3 .

7. The method for extracting phytoliths according to claim 1, characterized in that: In step 5, air and carbon dioxide are mixed in a volume ratio of 5:1 to 10:

1.

8. The method for extracting phytoliths according to claim 1, characterized in that: The digestion parameters in step 9 are: 1200 W, 130℃, 15 min.

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