CN111366597B - Method for detecting breathing exposure hazard of rock wool material micro-nano fibers - Google Patents
Method for detecting breathing exposure hazard of rock wool material micro-nano fibers Download PDFInfo
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- G01N23/00—Investigating or analysing materials by the use of wave or particle radiation, e.g. X-rays or neutrons, not covered by groups G01N3/00 – G01N17/00, G01N21/00 or G01N22/00
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- G—PHYSICS
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- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
- G01N33/5008—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
- G01N33/5014—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics for testing toxicity
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- G01N5/00—Analysing materials by weighing, e.g. weighing small particles separated from a gas or liquid
- G01N5/02—Analysing materials by weighing, e.g. weighing small particles separated from a gas or liquid by absorbing or adsorbing components of a material and determining change of weight of the adsorbent, e.g. determining moisture content
Abstract
The invention discloses a method for detecting the breathing exposure hazard of rock wool material micro-nano fibers, and belongs to the technical field of biological detection. The method adopts an ultrasonic dispersion-Scanning Electron Microscope (SEM) to perform in-vitro release characterization on the rock wool material, and adopts low-temperature ashing-SEM to detect lung retention after drip irrigation of a rat trachea, so as to realize detection of the breathing exposure harm of the micro-nano fibers of the rock wool material. The evaluation detection scheme has the advantages of being simple in operation, capable of truly simulating the breathing exposure harm of the rock wool material in the lung tissue, high in practicability and the like, and can evaluate the breathing exposure harm of the rock wool material.
Description
Technical Field
The invention relates to a method for detecting the breathing exposure hazard of rock wool material micro-nano fibers, belonging to the technical field of biological detection.
Background
Rock wool material is an inorganic and amorphous fibrous silicate substance formed by melting or fiberizing natural ore and minerals at high temperature, and is an important variety of artificial mineral fibers. The rock wool material has wide application, can be processed into products such as plates, strips, pipe shells, felts and the like, and is mainly used for heat preservation, heat insulation, sound absorption, sound insulation and fire prevention of industrial equipment pipelines and building walls; the method is used for planting and culturing fruits, vegetables and flowers in agriculture; the vibration isolator is used for rail vibration isolation in the traffic field and the like. However, with the rapid production and development of rock wool products, the number of occupational population exposed to rock wool dust increases year by year, and the harm of the rock wool micro-nano fibers, such as lung tissue fibrosis, chronic lung obstruction, carcinogenesis and the like, causes wide attention.
Studies have shown that the health hazards of rockwool are closely related to the size of the released micro-nanofibers, the biological retention of lung tissue [ Methods for the determination of the halogenated properties for human health of human parent mineral fibers, David M. Bernstein et al, 1999 ]. Currently, the environmental health safety evaluation on rock wool fiber materials mainly comprises three methods. The first is to adopt a mode of on-site touch and observation experience [ rock wool: the method is not objective and accurate enough and is difficult to quantitatively describe, and the method is a new method for researching the safety of external thermal insulation materials of external walls, namely 'construction science' in 2011, 13 th and 72-74 pages. The second method is a method for detecting cytotoxicity, for example, the method for detecting cytotoxicity of rock wool material by using hamster lung V79 cells by dungjian army et al [ comparison study on in vitro cytotoxicity of chrysotile and fibrous asbestos substitutes, dungjian army et al, chinese occupational medicine, 2010, 5 th year, page 365-. The third method is to evaluate the safety of rockwool material by examining the health of the lung ventilation function of professional workers [ the influence of occupational exposure to rockwool on the lung ventilation function and respiratory symptoms, junjicheng billow, [ industrial hygiene and occupational diseases ], 2012, 2 nd, pages 68-72 ], which, although accurate in result, has hysteresis and is not predictive for rockwool product safety evaluation. Therefore, there is an urgent need to develop a new safety evaluation method for performing a systematic evaluation of the safety of industrial rock wool materials.
Disclosure of Invention
In order to solve the problems, the invention provides a method for performing in-vitro release characterization on rock wool materials by adopting an ultrasonic dispersion-Scanning Electron Microscope (SEM), and a novel method for detecting lung retention by low-temperature ashing-SEM after rat tracheal drip irrigation. The evaluation detection scheme has the advantages of being simple in operation, capable of truly simulating the breathing exposure harm of the rock wool material in the lung tissue, high in practicability and the like, and can evaluate the breathing exposure harm of the rock wool material.
The invention aims to provide a method for detecting the breathing exposure hazard of rock wool material micro-nano fibers, which comprises the following steps:
s1, dispersing rock wool materials in 0.01-10% NaCl aqueous solution, performing ultrasonic treatment for 0.1-100 hours to obtain rock wool fiber dispersion solution, and performing SEM characterization on the rock wool fiber dispersion solution, wherein the concentration of the rock wool fiber dispersion solution is 0.5-100 mg/mL;
s2, drip-irrigating the rock wool fiber dispersion solution into rat lung tissues by using a trachea, wherein the dropping amount is 0.1-4 mg/Kg;
s3, weighing 0.1-5g of lung tissues of the cultured rat, and ashing at low temperature for 1-48 hours to obtain low-temperature ashes;
s4, dissolving the low-temperature ashes in a 0.01-10% NaCl aqueous solution to obtain a low-temperature ashes solution, and performing SEM representation on the low-temperature ashes solution;
s5, in vitro release and bioretention evaluation for SEM characterization in steps S1 and S4.
Further, in step S1, the power of ultrasonic wave is 1-100W, and the solution is placed in ice bath during ultrasonic wave. Preferably 10-20W is adopted, the treatment is carried out for 0.5-2 hours, and the treatment process adopts opening for 15-25 s and closing for 5-10 s.
Furthermore, the concentration of the rock wool fiber dispersion solution is preferably 0.5-10 mg/mL.
Further, in the step S2, the drip irrigation of the trachea is performed by dividing the rockwool fiber dispersion solution into four portions, continuously performing the drip irrigation for four days, and dissecting 35 to 45 days after the drip irrigation to take out the whole lung tissue of the rat.
Further, the volume of the drip irrigation is 100-300 mu L/day/machine.
Further, in step S3, the lung tissue is lyophilized before low-temperature ashing, wherein the power of the low-temperature ashing is 1-150W. Preferably, the lyophilized lung tissue is cryoashed using 80-120W for 1-5 hours.
Further, in step S4, the concentration of the low temperature ashing solution is 0.5 to 10 mg/mL.
Further, in step S5, in vitro release and bioretention was evaluated by measuring the diameter and length of the fibers by SEM images of the dispersion solution of rock wool fibers and the solution of low temperature ashes, and counting the fiber ratios of length >20 μm, length <5 μm and length between 5-20 μm.
Further, the aqueous NaCl solution is preferably a physiological saline solution with a concentration of 0.9%.
The purpose of the invention is mainly realized by two technical schemes:
the method is characterized in that the rock wool releases micro-nano fibers in vitro. This solution requires the design of conditions that optimize the in vitro release. An ice bath probe ultrasonic mode is adopted, and the obtained micro-nano fiber has physical and chemical properties meeting the requirements of further animal experiments.
And secondly, evaluating the retention of the rockwool micro-nano fiber in the lung. We used the rat model, through the way of trachea instillation, using plasma low temperature ashing to realize this technical scheme. The low-temperature ashing can effectively remove the interference of biological tissues on the detection of the nano fibers, and simultaneously effectively retain the residual micro-nano fibers in the lung tissues for the characterization of SEM.
The invention has the beneficial effects that:
the evaluation and detection scheme provided by the invention has the advantages of simplicity in operation, capability of truly simulating the breathing exposure hazard of the rock wool material in the lung tissue, strong practicability and the like, and can be used for evaluating the breathing exposure hazard of the rock wool material.
Drawings
FIG. 1 is a technical roadmap for the inventive arrangements;
FIG. 2 is a surface topography of a rock wool material after being dispersed for 1 hour by a shaker;
FIG. 3 is a surface topography of a rock wool material after 1 hour of probe ultrasound;
FIG. 4 is an SEM image of high temperature ashing lung tissue dispersion with tracheal drip irrigation for 40 days;
FIG. 5 is an SEM image of a dispersion of low-temperature ashed lung tissue after 40 days of tracheal drip irrigation, in which A is rat lung tissue tracheal drip exposure of 0.5mg/Kg of rockwool fiber, and B is rat lung tissue tracheal drip exposure of 2mg/Kg of rockwool fiber.
Detailed Description
The present invention is further described below in conjunction with the following figures and specific examples so that those skilled in the art may better understand the present invention and practice it, but the examples are not intended to limit the present invention.
Example 1:
a batch of rock wool material was randomly drawn, 0.5g of the material was weighed into a 250ml blue-capped bottle on a precision balance, 100ml of 0.9% sodium chloride solution was added, the glass rod was stirred well until no flocculent material was present, and further shaken on a shaker for 1 hour (200 rpm/min). And (3) dropping 10 mu l of the liquid transfer gun onto a clean silicon wafer, standing in a drying box overnight, and performing SEM characterization on the shape and the size of the dispersed rock wool material. As shown in fig. 2, the rockwool was not able to form monodisperse rockwool fibers after being subjected to probe ultrasound.
Example 2:
randomly extracting a certain batch of rock wool material, weighing 0.5g of the material into a 250ml blue-cap bottle by using a precision balance, adding 100ml of 0.9% sodium chloride solution, fully stirring the glass rod until no flocculent substance exists, and carrying out ultrasonic treatment on the glass rod for 1 hour by using a probe ice bath (opening for 20s, closing for 5s, and power of 15W). And (3) dropping 10 mu l of the liquid transfer gun onto a clean silicon wafer, standing in a drying box overnight, and performing SEM characterization on the shape and the size of the dispersed rock wool material. As shown in fig. 3, rock wool can form a uniform micro-nanofiber dispersion after being subjected to probe ultrasound. The average length of the fibers was less than 20 microns (table 1).
Example 3:
62.5mg and 250mg of rock wool fiber materials are respectively weighed by a precision balance, respectively dispersed in 100ml of 0.9% sodium chloride solution, and after the glass rod is fully and uniformly stirred, the probe is subjected to ultrasonic treatment for 1 hour (20 s is turned on and 5s is turned off; the power is 15W). The prepared material was drip-irrigated into anesthetized Wistar rats by a tracheal drip irrigation volume of 200 μ L/day/rat for four consecutive days. After the drip irrigation, the weight change of the rats was measured weekly, and the whole lung tissue of the rats was dissected and removed 40 days later. The tissue was freeze-dried in a-80 ℃ freezer and the tissue was weighed. After the lyophilized tissue was incinerated in a muffle furnace at 800 ℃ for 8 hours, the lyophilized tissue was re-concentrated to 5mg/ml with 0.9% NaCl solution, and 10. mu.l was taken for SEM characterization. The results are shown in fig. 4, and the samples obtained by this method are easily agglomerated, and it is difficult to obtain well-dispersed rock wool fibers.
Example 4:
62.5mg and 250mg of rock wool fiber materials are respectively weighed by a precision balance, respectively dispersed in 100ml of 0.9% sodium chloride solution, and after the glass rod is fully and uniformly stirred, the probe is subjected to ultrasonic treatment for 1 hour (20 s is turned on and 5s is turned off; the power is 15W). The prepared material was drip-irrigated into anesthetized Wistar rats by a tracheal drip irrigation volume of 200 μ L/day/rat for four consecutive days. After the drip irrigation, the weight change of the rats was measured weekly, and the whole lung tissue of the rats was dissected and removed 40 days later. The tissue was freeze-dried in a-80 ℃ freezer and the tissue was weighed. The freeze-dried tissue plasma is incinerated at low temperature (100W) for 2 hours to obtain the rock wool material retained in the tissue, the rock wool material is concentrated to 5mg/ml by 0.9% sodium chloride solution, and 10 mu l of the rock wool material is taken for SEM characterization. As shown in fig. 5, the remaining rockwool fibers were clearly observed by SEM. The length of the residual rockwool fibres was counted and it was found that after 40 days of exposure, rockwool fibres with a length of >20 microns in lung tissue accounted for approximately 8.3% (table 1) compared to that before the drop addition, which was less than the specified 50%. Indicating that the batch of rockwool material has a lower risk of respiratory exposure toxicity.
TABLE 1 characterization of physicochemical properties of rockwool micro-nanofibers in vitro and in mouse lung tissue
The above-mentioned embodiments are merely preferred embodiments for fully illustrating the present invention, and the scope of the present invention is not limited thereto. The equivalent substitution or change made by the technical personnel in the technical field on the basis of the invention is all within the protection scope of the invention. The protection scope of the invention is subject to the claims.
Claims (9)
1. A method for detecting the breath exposure hazard of rock wool material micro-nano fibers is characterized by comprising the following steps:
s1, dispersing rock wool materials in 0.01-10% NaCl aqueous solution, performing ultrasonic treatment for 0.1-100 hours to obtain rock wool fiber dispersion solution, and performing SEM characterization on the rock wool fiber dispersion solution, wherein the concentration of the rock wool fiber dispersion solution is 0.5-100 mg/mL;
s2, drip-irrigating the rock wool fiber dispersion solution into rat lung tissues by using a trachea, wherein the dropping amount is 0.1-4 mg/Kg;
s3, weighing 0.1-5g of lung tissues of the cultured rat, and ashing the rat for 1-48 hours at low temperature by adopting plasma to obtain low-temperature ashes;
s4, dissolving the low-temperature ashes in a 0.01-10% NaCl aqueous solution to obtain a low-temperature ashes solution, and performing SEM representation on the low-temperature ashes solution;
s5, in vitro release and bioretention evaluation for SEM characterization in steps S1 and S4.
2. The method of claim 1, wherein in step S1, the power of the ultrasound is 1-100W, and the solution is placed in an ice bath during the ultrasound.
3. The method of claim 1, wherein the concentration of the rockwool fiber dispersion solution is 0.5-10 mg/mL.
4. The method as claimed in claim 1, wherein the tracheal drip irrigation is performed by dividing a dropping amount of the rockwool fiber dispersion solution into four parts, continuously performing the drip irrigation for four days, and dissecting 35-45 days after the drip irrigation to remove the whole lung tissue of the rat at S2.
5. The method as claimed in claim 4, wherein the volume of the drip irrigation is 100 μ L/day/machine.
6. The method according to claim 1, wherein in step S3, the lung tissue is lyophilized before low-temperature ashing, and the power of the low-temperature ashing is 1 to 150W.
7. The method as claimed in claim 1, wherein the concentration of the low temperature ashing solution is 0.5 to 10mg/mL at S4.
8. The method of claim 1, wherein in step S5, the in vitro release and bioretention assessment is by measuring the diameter and length of the fibers from SEM images of the rockwool fiber dispersion solution and the cryo-ashing solution, with statistical fiber ratios of >20 μm length <5 μm and length between 5 and 20 μm.
9. The method according to claim 1, wherein the aqueous NaCl solution is 0.9% physiological saline.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106796217A (en) * | 2014-07-21 | 2017-05-31 | 泰克年研究发展基金会公司 | For the composition of directly breathing sampling |
| CN207036849U (en) * | 2017-08-15 | 2018-02-23 | 魏丽 | A kind of medical breath sample detection means |
| KR101956346B1 (en) * | 2018-05-09 | 2019-03-08 | 서울대학교산학협력단 | Compositions for Diagnosing of cancer using Volatile Organic Compounds |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106796217A (en) * | 2014-07-21 | 2017-05-31 | 泰克年研究发展基金会公司 | For the composition of directly breathing sampling |
| CN207036849U (en) * | 2017-08-15 | 2018-02-23 | 魏丽 | A kind of medical breath sample detection means |
| KR101956346B1 (en) * | 2018-05-09 | 2019-03-08 | 서울대학교산학협력단 | Compositions for Diagnosing of cancer using Volatile Organic Compounds |
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| Title |
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| 岩棉粉尘对动物细胞毒性作用的实验研究;韩明禄等;《中国公共卫生学报》;19981231;第17卷(第1期);第52页 * |
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