Method for separating impurity minerals from archaeological site unearthed bone biological apatite

Abstract

The invention discloses a method for separating impurity minerals in archaeological site unearthed bone biological apatite, and belongs to the technical field of pretreatment of archaeological site unearthed bone samples. The method specifically comprises the following steps: 1) putting a pretreated block-shaped skeleton sample into deionized water, and then freezing for 80-100 minutes in an environment of-45 DEG C to-55 DEG C; transferring the frozen skeleton sample to an environment of 75-85 DEG C to be unfrozen for 8-12 minutes, and completing a freezing and thawing cycle; (2) repeating the steps for a plurality of freezing and thawing cycles to obtain a freezing and thawing product; and (3) carrying out particle size sorting on the freeze-thaw product in a leaching manner to respectively obtain skeleton samples with the particle size ranges of 0.5-2mm, 2-5mm and greater than 5mm for subsequent analysis. On the basis of thermal expansion difference and ice expansion stress, a bonding interface of impurity minerals and biological apatite is destroyed, particle size distribution characteristics are combined, gravity screening is replaced by liquid flow screening, re-adsorption caused by triboelectrification is avoided, and precise impurity mineral separation is achieved.

Description

Technical Field

[0001] The present invention belongs to the technical field of pretreatment of bone samples unearthed from archaeological sites, and particularly relates to a method for separating impurity minerals in bone bioapatite unearthed from archaeological sites. Background Art

[0002] In the field of bioarchaeological research, the carbon and oxygen isotope ratios of bone bioapatite unearthed can reflect the diet, migration and paleoenvironmental information of ancient humans or animals, and are important analysis carriers. However, impurity minerals from the soil, such as calcite, quartz, muscovite, feldspar, etc., are often densely attached to the surface of bioapatite crystals or within the pores of cancellous bone. The presence of these impurity components will interfere with the pollution identification results and reduce the accuracy of isotope testing. With the continuous improvement of the testing accuracy requirements, it has become an urgent task to develop an efficient pretreatment technology to remove these impurities.

[0003] Currently, the methods for removing exogenous impurity minerals are mainly divided into two categories: physical methods and chemical methods, but both of these methods have obvious limitations. In physical methods, although ultrasonic cavitation can strip the attached impurity particles, due to some particles being tightly adhered or deeply embedded in the bone pores, they cannot be fully exposed to the ultrasonic action environment, resulting in poor treatment effects and difficulty in completely removing impurities. This indicates that the ultrasonic cleaning technology is limited by the strong binding effect between impurity minerals and the bioapatite matrix. This binding has multiple characteristics: firstly, a heterogeneous interface solid-phase connection is formed through interface actions such as electrostatic adsorption and mechanical interlocking; secondly, there are significant differences in the thermal expansion coefficients between different mineral components (e.g., quartz α = 5.5×10 -7 / K, bioapatite α = 13.3×10 -6 / K); thirdly, the relative particle size and distribution characteristics of various mineral particles. In chemical methods, acetic acid has been proven to be able to effectively dissolve calcite impurities. However, for other chemically stable oxides and silicate impurities such as quartz (quartz is the most common pollution mineral except calcite), although hydrofluoric acid or strong alkalis (such as NaOH) have good dissolution effects, the use of such strongly corrosive reagents requires dedicated protective facilities, and the waste liquid treatment cost is high, which is difficult to popularize and promote for ordinary archaeological laboratories.

[0004] In summary, the existing physical and chemical methods have insurmountable limitations in removing exogenous impurity minerals in bioapatite, which not only affects the accuracy of isotope testing but also restricts the in-depth development of bioarchaeological research. Therefore, it is urgent to design a new separation technology to remove impurity substances to meet the increasing high-precision analysis requirements. Summary of the Invention

[0005] The object of the present invention is to solve the deficiencies in the prior art and provide a method for separating impurity minerals from bone bioapatite unearthed from archaeological sites.

[0006] The specific technical solution adopted by the present invention is as follows:

[0007] The present invention provides a method for separating impurity minerals from bone bioapatite unearthed from archaeological sites, and the specific steps are as follows:

[0008] S1: Place the pretreated massive bone sample in water, and then place it in an environment of -45°C to -55°C for freezing for 80 to 100 minutes; transfer the frozen bone sample to an environment of 75°C to 85°C for thawing for 8 to 12 minutes to complete one freeze-thaw cycle;

[0009] S2: Repeat step S1, and after performing several freeze-thaw cycles, obtain a freeze-thaw product;

[0010] S3: Perform particle size sorting on the freeze-thaw product obtained in step S2 by leaching method to obtain bone samples with particle size ranges of 0.5 to 2 mm, 2 to 5 mm, and particle size greater than 5 mm respectively for subsequent analysis.

[0011] Preferably, the pretreatment method of the bone sample is specifically as follows:

[0012] S11: Place the massive bone sample to be treated in a test tube containing deionized water so that the bone sample is immersed in the deionized water; repeatedly clean the bone sample with ultrasonic waves to remove surface dust and loose impurities;

[0013] S12: Immerse the cleaned bone sample in a sodium hypochlorite solution for dissolving organic matter, let it stand for 24 hours, and then wash the bone sample with deionized water until it is neutral;

[0014] S13: Place the bone sample washed in step S12 in an acetic acid - calcium acetate buffer solution for dissolving carbonate, shake it to react fully, then let it stand for 24 hours, and wash the bone sample with deionized water until it is neutral to complete the pretreatment of the bone sample.

[0015] Furthermore, in step S11, the frequency of the ultrasonic wave is 40 to 50 kHz, and the cleaning time is 15 to 20 minutes.

[0016] Furthermore, in step S12, the volume concentration of the sodium hypochlorite solution is 2%.

[0017] Furthermore, in step S13, the concentration of the acetic acid - calcium acetate buffer solution is 1 mol / L.

[0018] Preferably, the freeze-thaw cycle in step S2 is performed 5 to 67 times.

[0019] Preferably, in step S3, deionized water is used for leaching, and the water flow rate is controlled at 0.5 - 1.5 L / min.

[0020] Preferably, an X-ray diffractometer is used to perform phase analysis on bone samples with particle size ranges of 0.5 - 2 mm, 2 - 5 mm, and particle size greater than 5 mm obtained after particle size sorting, and bone samples with impurity mineral content lower than the detection limit of 5 wt% are used for subsequent analysis.

[0021] Compared with the prior art, the present invention has the following beneficial effects:

[0022] The present invention innovatively proposes a freeze-thaw cycle separation method based on the thermal expansion mismatch of minerals and the cyclic freeze-thaw effect of water. Through the synergistic effect of using an ultra-low temperature refrigerator (-50 °C, 90 min) and a constant temperature metal bath (80 °C, 10 min), the periodic stress generated by the difference in thermal expansion performance of heterogeneous materials and the expansion of water during freezing is fully utilized to effectively break the bonding interface between impurity minerals and bioapatite. Since these two devices are relatively popular in ordinary archaeological laboratories, and separating the freezing and thawing functions can significantly improve the freeze-thaw cycle efficiency, it has good application prospects.

[0023] Furthermore, in combination with the relative particle size distribution characteristics (from micron to millimeter level) of various mineral particles in archaeological bones, a multi-stage particle size screening system is designed. By using the liquid flow medium screening method, the problem of re-adsorption of bioapatite particles and impurities caused by frictional electrification of dry samples during the traditional gravity screening process is effectively avoided, thus achieving more accurate separation of impurity minerals. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 XRD patterns of bone samples of sample DR12 at different particle sizes;

[0025] Figure 2 XRD patterns of bone samples of sample DR11 at different particle sizes. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0026] The present invention will be further described and explained below in conjunction with the drawings and specific embodiments. Without conflict, the technical features of each embodiment in the present invention can be combined accordingly. The bone samples in the following specific examples are from bones containing impurities unearthed from different archaeological sites.

[0027] Example 1

[0028] In this example, the separation of impurity minerals in bone bioapatite of 3 different bone samples is carried out, and the specific method is as follows:

[0029] I. Pretreatment of bone samples

[0030] (1) Use a cutter to cut three bone block samples from different sources into pieces of about 2 g. While cutting the samples, use the cutter to gently scrape off the surface contaminants. The samples are numbered DR10, DR11, and DR12.

[0031] (2) Three different block bone samples to be processed are placed in a test tube containing deionized water, so that the bone samples are immersed in the deionized water. The test tube is placed in an ultrasonic cleaner, and the bone samples are repeatedly cleaned with ultrasound to remove surface dust and loose impurities; the frequency of the ultrasound is 40 kHz, the total power is 1200 W, and the cleaning time is 15 minutes, until the solution is clear.

[0032] (3) Immerse the cleaned bone sample in a 2% sodium hypochlorite solution, loosely cover the mouth of the test tube with aluminum foil, let it stand for 24 hours, and then wash the bone sample with deionized water until it is neutral to remove organic matter in the bone sample.

[0033] (4) Place the bone sample washed in step (3) in 1 mol / L acetic acid-calcium acetate buffer (pH adjusted to 4.8) and shake for 1 minute. Cover the mouth of the test tube loosely with aluminum foil. After standing for 24 hours, wash the bone sample with deionized water until it is neutral to remove carbonate in the bone sample, thereby completing the pretreatment of the bone sample.

[0034] 2. Freeze-thaw cycle treatment

[0035] The pretreated block bone sample was placed in deionized water and then frozen in an ultra-low temperature refrigerator at -50°C for 90 minutes; the frozen bone sample was transferred to a constant temperature metal bath at 80°C for 8-10 minutes to complete a freeze-thaw cycle. Both the ultra-low temperature refrigerator and the constant temperature metal bath can automatically control the constant temperature with an accuracy within the range of 1°C and 0.5°C. The freeze-thaw product was obtained after repeating the above freeze-thaw cycle 46 times and performing a total of 47 freeze-thaw cycles. According to the crushing of the bone sample during the freeze-thaw process, it needs to be screened every 10-20 times, and samples larger than 0.5 mm are retained to ensure the amount of samples for subsequent testing.

[0036] 3. Particle size sorting

[0037] The freeze-thaw products obtained in step 2 were sorted by particle size by leaching. Using 5mm, 2mm and 0.5mm aperture sieves, deionized water was used for leaching and sieving, and the water flow rate was controlled at 1L / min. Bone samples with particle sizes ranging from 0.5 to 2mm, 2 to 5mm and particle sizes greater than 5mm were obtained, and then placed in a 70℃ oven to dry for 24h for subsequent X-ray diffraction testing.

[0038] 4. X-ray diffraction analysis

[0039] The X-ray diffraction phase analysis of the bone samples after particle size sorting was carried out using an X-ray diffractometer. The specific method is as follows: an X-ray diffractometer of D8 Discover, test conditions: power 42 kV×100 mA, Cu Kα radiation, scanning range: 5 - 80°(2θ), step width 0.02°, scanning speed 0.1 s / step, one-dimensional continuous scanning. The results are shown in Table 1.

[0040] Table 1 Phase identification results of different bone samples in Example 1

[0041]

[0042] Note: "◎" indicates only containing single-phase bioapatite, "×" indicates containing exogenous impurity minerals, "N" indicates no sample obtained

[0043] Comparative Example 1

[0044] Compared with Example 1, in this comparative example, the separation of impurity minerals in bone bioapatite was carried out on 3 different bone samples. The specific method is as follows:

[0045] I. Pretreatment of bone samples, the same as in Example 1.

[0046] II. Freeze-thaw cycle treatment, the same as in Example 1.

[0047] III. Particle size sorting

[0048] The freeze-thaw products obtained in step II were subjected to particle size sorting by leaching. Using sampling sieves with pore sizes of 0.125 mm and 0.25 mm, deionized water was used for leaching and sieving, and the water flow rate was controlled at 1 L / min. Bone samples with particle size ranges of 0.125 - 0.25 mm and particle sizes less than 0.125 mm were obtained respectively, and then placed in an oven at 70°C for drying for 24 h for subsequent X-ray diffraction testing.

[0049] IV. X-ray diffraction analysis

[0050] The X-ray diffraction phase analysis of the bone samples after particle size sorting was carried out using an X-ray diffractometer. The specific method is the same as in Example 1. The results are shown in Table 2.

[0051] Table 2 Phase identification results of different bone samples in Comparative Example 1

[0052]

[0053] Note: "◎" indicates only containing single-phase bioapatite, "×" indicates containing exogenous impurity minerals, "N" indicates no sample obtained

[0054] Comparing Table 2 and Table 1, it can be seen that the particle size classification of the product after freeze-thaw, namely 0.125 - 0.25 mm and particle size less than 0.125 mm, cannot separate the impurity minerals in the bone sample, which is not conducive to subsequent analysis. According to the table, after the sample DR11 undergoes freeze-thaw cycles and particle size separation, the impurity particles are separated to smaller particle sizes (0.125 - 0.25 mm and <0.125 mm), and there are no impurities in the larger particle sizes (0.5 - 2 mm, 2 - 5 mm, and >5 mm).

[0055] Figure 1 XRD patterns of bone samples of sample DR12 at different particle sizes Figure 2 XRD patterns of bone samples of sample DR11 at different particle sizes. According to Figure 1 it can be seen that after the sample DR12 undergoes freeze-thaw cycles, the bone samples with particle sizes less than 0.125 mm and 0.125 - 0.25 mm contain impurity minerals (quartz, muscovite), while the bone samples with particle sizes of 0.5 - 2 mm only contain hydroxyapatite and no impurity minerals. According to Figure 2 it can be seen that after the sample DR11 undergoes freeze-thaw cycles, the bone samples with particle sizes less than 0.125 mm and 0.125 - 0.25 mm contain impurity minerals (quartz), while the bone samples with particle sizes of 0.5 - 2 mm, 2 - 5 mm, and particle size greater than 5 mm only contain hydroxyapatite and no impurity minerals. It shows that the method provided by the present invention can obtain bone samples without impurity minerals, significantly improve the utilization rate of contaminated bone samples, and achieve green separation without chemical pollution.

[0056] The above-described embodiments are only a preferred solution of the present invention, but they are not intended to limit the present invention. Those of ordinary skill in the relevant technical field can still make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, all technical solutions obtained by adopting equivalent replacement or equivalent transformation fall within the protection scope of the present invention.

Claims

1. A method for separating impurity minerals from apatite of bones unearthed from archaeological sites, characterized in that: The specific steps are as follows: S1: Place the pretreated block bone sample in water, and then freeze it at -45℃ to -55℃ for 80 to 100 minutes; transfer the frozen bone sample to 75℃ to 85℃ for thawing for 8 to 12 minutes, completing a freeze-thaw cycle; S2: repeat step S1 and perform several freeze-thaw cycles to obtain a freeze-thaw product; S3: The freeze-thaw product obtained in step S2 is sorted by particle size by leaching to obtain bone samples with particle sizes ranging from 0.5 to 2 mm, 2 to 5 mm, and larger than 5 mm for subsequent analysis.

2. The method for separating impurity minerals from apatite of skeletons unearthed from archaeological sites according to claim 1, characterized in that: The pretreatment method of bone samples is as follows: S11: placing the block bone sample to be processed in a test tube containing deionized water, so that the bone sample is immersed in the deionized water; repeatedly cleaning the bone sample with ultrasound to remove surface dust and loose impurities; S12: Immerse the cleaned bone sample in a sodium hypochlorite solution for dissolving organic matter, let it stand for 24 hours, and then wash the bone sample with deionized water until it is neutral; S13: The bone sample washed in step S12 is placed in an acetic acid-calcium acetate buffer for dissolving carbonate and shaken for sufficient reaction. After standing for 24 hours, the bone sample is washed with deionized water until it is neutral, thereby completing the pretreatment of the bone sample.

3. The method for separating impurity minerals from skeletal biological apatite unearthed from archaeological sites according to claim 2, characterized in that: In step S11, the frequency of the ultrasonic wave is 40 to 50 kHz, and the cleaning time is 15 to 20 minutes.

4. The method for separating impurity minerals from apatite of skeletons unearthed from archaeological sites according to claim 2, characterized in that: The volume concentration of the sodium hypochlorite solution in step S12 is 2%.

5. The method for separating impurity minerals from apatite of skeletons unearthed from archaeological sites according to claim 2, characterized in that: The concentration of the acetic acid-calcium acetate buffer in step S13 is 1 mol / L.

6. The method for separating impurity minerals from apatite of skeletons unearthed from archaeological sites according to claim 1, characterized in that: The freeze-thaw cycle in step S2 is performed 5 to 67 times.

7. The method for separating impurity minerals from apatite of skeletons unearthed from archaeological sites according to claim 1, characterized in that: In step S3, deionized water is used for leaching, and the water flow rate is controlled at 0.5-1.5 L / min.

8. The method for separating impurity minerals from apatite of skeletons unearthed from archaeological sites according to claim 1, characterized in that: X-ray diffractometer was used to perform phase analysis on bone samples with particle sizes ranging from 0.5 to 2 mm, 2 to 5 mm, and larger than 5 mm obtained after particle size sorting, and bone samples with impurity mineral content below the detection limit of 5wt% were used for subsequent analysis.

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