CN107881109B - Method for preparing fecal strain liquid by removing copper, lead and zinc ions in human feces through lithium illite - Google Patents
Method for preparing fecal strain liquid by removing copper, lead and zinc ions in human feces through lithium illite Download PDFInfo
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Abstract
The invention discloses a method for preparing a fecal bacterial solution by removing copper, lead and zinc ions in human feces by utilizing lithium illite. The invention can remove copper, lead and zinc ions in human excrement by specific adsorption, so that excrement donated by excrement donors with excessive copper, lead and zinc ions can reach the standard after treatment.
Description
Technical Field
The invention belongs to the technical field of human fecal bacteria transplantation, fermentation and detection, and relates to a method for removing copper, lead and zinc ions in human feces by utilizing lithium illite.
Background
"Fecal transplantation" (FMT) refers to the transplantation of functional flora in the feces of healthy people into the gastrointestinal tract of patients to reconstruct new intestinal flora and realize the treatment of intestinal and parenteral diseases. Fecal transplantation has been used for the treatment and exploratory studies of various flora-related diseases such as clostridium difficile infection, inflammatory bowel disease, intractable constipation, metabolic disease, intestinal immunodeficiency, intestinal allergy and the like as an effective means for reconstructing intestinal flora, and is considered as a breakthrough medical progress in recent years.
The fecal bacteria feeding way is divided into an upper digestive tract, a middle digestive tract and a lower digestive tract. The upper digestive tract route mainly refers to oral fecal bacteria capsule; the middle digestive tract route comprises passing through a nasointestinal tube, a gastroscope forceps channel hole and a percutaneous endoscopic gastrostomy jejunum tube; the lower gastrointestinal route includes colonoscopy, enema, colostomy, and endoscopic intestinal catheterization (TET). The physician should select the most appropriate input route for the patient according to his condition and the characteristics of the various routes.
The state of the coprophilous fungi transplantation can be divided into three forms of freshness, freezing storage and fermentation. Since the safety of fecal bacteria transplantation is the primary consideration in clinical decision making, in order to isolate a high quality fecal flora without pathogens, the fecal donors are highly demanding, at least within 3-6 months no diarrhea occurs, no antibiotics have been used, and there must be no infectious diseases and gastrointestinal diseases, in addition, there are limitations including age, physiology, pathology, psychology, integrity, time, environment, etc.
As the fecal bacteria provided by qualified fecal donors can not meet the clinical requirements at all, the fecal bacteria with certain ions exceeding the standard is processed by the technology, so that the fecal bacteria meets the requirements of fecal bacteria transplantation, and the method has very important practical significance.
Along with the change of human living environment, the exceeding of copper, lead and zinc ions in human excrement is more and more common, and when the copper, lead and zinc ions in the excrement of an excrement provider exceed the standard, the requirement of excrement bacterium transplantation is not met.
Disclosure of Invention
The invention aims to provide a method for removing copper, lead and zinc ions in human excrement by utilizing lithium illite.
The technical scheme adopted by the invention is as follows:
a method for preparing a fecal bacteria liquid by removing copper, lead and zinc ions in human feces by utilizing lithium illite,
1) adding a sterile NaCl solution into the excrement sample, adding Likelite, and uniformly stirring to obtain excrement slurry;
2) primarily filtering the excrement slurry, taking filtrate, and stirring the filtrate at 1800-2200 rpm for 3-6 min every 0.8-1.2 h under the protection of carbon dioxide, nitrogen, helium, neon or argon, and obtaining the excrement and bacterium slurry after 4-7 h;
3) filtering the fecal strain slurry to obtain filtrate to obtain crude fecal strain liquid; and centrifuging the crude fecal strain liquid, taking the precipitate, and re-suspending to obtain colorless and odorless fecal strain liquid containing no copper, lead and zinc ions.
Further, the preparation method of the lithium illite comprises the following steps: adding water and lithium metaphosphate into illite ore, heating to 70-80 ℃, and adding Na2CO3(ii) a Stirring for 2.5-3.5 h, standing for 20-30 h, then centrifugally washing with deionized water, recovering solids, and drying to obtain the Lidikite.
Further, the illite mineral, lithium metaphosphate and Na2CO3And the dosage ratio of water is 80-120 g: 8-12 g: 18-22 g: 1L of the compound.
Furthermore, the cation exchange capacity of the obtained lithium-based illite is 40-60 mmol/100 g.
Further, in the step 1), the dosage ratio of the excrement sample, the NaCl solution and the Lidikite is 1 g: 3-7 mL: 0.5 to 1.5 g.
Further, the concentration of the NaCl solution is 0.85-0.95% w/v.
Further, in the step 1), the stirring speed is 1800-2200 rpm, and the time is 2-4 min.
Further, in step 2), the specific operations of the primary filtering are as follows: and filtering the excrement slurry by using a filter screen with the aperture of 4.5-5.5 mm to remove large-particle substances.
Further, in step 3), the specific operation of filtering the fecal sewage sludge is as follows: filtering by a filter screen with the aperture of 1.5-2.5 mm, 0.8-1.2 mm, 0.4-0.6 mm and 0.2-0.3 mm in sequence.
Further, in the step 3), the rotation speed of the centrifugation is 5500-6500 rpm, and the time is 12-18 min.
The invention has the beneficial effects that:
at present, a method for removing copper, lead and zinc ions in human excrement is not reported temporarily, and the method can remove the copper, lead and zinc ions in the human excrement through specific adsorption, so that excrement donated by excrement donors with excessive copper, lead and zinc ions can reach the standard after treatment.
Detailed Description
A method for preparing a fecal bacteria liquid by removing copper, lead and zinc ions in human feces by utilizing lithium illite,
1) adding a sterile NaCl solution into the excrement sample, adding Likelite, and uniformly stirring to obtain excrement slurry;
2) primarily filtering the excrement slurry, taking filtrate, and stirring the filtrate at 1800-2200 rpm for 3-6 min every 0.8-1.2 h under the protection of carbon dioxide, nitrogen, helium, neon or argon, and obtaining the excrement and bacterium slurry after 4-7 h;
3) filtering the fecal strain slurry to obtain filtrate to obtain crude fecal strain liquid; and centrifuging the crude fecal strain liquid, taking the precipitate, and re-suspending to obtain colorless and odorless fecal strain liquid containing no copper, lead and zinc ions.
Preferably, the preparation method of the lithium illite comprises the following steps: adding water and lithium metaphosphate into illite ore, heating to 70-80 ℃, and adding Na2CO3(ii) a Stirring for 2.5-3.5 h, standing for 20-30 h, then centrifugally washing with deionized water, recovering solids, and drying to obtain the Lidikite.
Preferably, the illite mineral, lithium metaphosphate and Na2CO3And the dosage ratio of water is 80-120 g: 8-12 g: 18-22 g: 1L of the compound.
Preferably, the drying temperature is 100-110 ℃, and the drying time is 45-55 h.
Preferably, the cation exchange capacity of the obtained lithium illite is 40-60 mmol/100 g.
Preferably, in the step 1), the dosage ratio of the excrement sample to the NaCl solution to the lithium illite is 1 g: 3-7 mL: 0.5 to 1.5 g.
Preferably, the concentration of the NaCl solution is 0.85-0.95% w/v.
Preferably, in the step 1), the stirring speed is 1800-2200 rpm, and the time is 2-4 min.
Preferably, in step 2), the preliminary filtration specifically comprises: and filtering the excrement slurry by using a filter screen with the aperture of 4.5-5.5 mm to remove large-particle substances.
Preferably, in step 3), the specific operation of filtering the fecal sewage sludge is as follows: filtering by a filter screen with the aperture of 1.5-2.5 mm, 0.8-1.2 mm, 0.4-0.6 mm and 0.2-0.3 mm in sequence.
Preferably, in the step 3), the rotation speed of the centrifugation is 5500-6500 rpm, and the time is 12-18 min.
Preferably, in step 3), the liquid used for resuspension is a sterile 0.85% -0.95% w/v NaCl solution.
The present invention will be further described with reference to the following examples.
The embodiments of the present invention are described below with reference to specific embodiments, and other advantages and effects of the present invention will be easily understood by those skilled in the art from the disclosure of the present specification. The invention is capable of other and different embodiments and of being practiced or of being carried out in various ways, and its several details are capable of modification in various respects, all without departing from the spirit and scope of the present invention.
It should be understood that the processing equipment or devices not specifically mentioned in the following examples are conventional in the art; all pressure values and ranges refer to absolute pressures.
Furthermore, it is to be understood that one or more method steps mentioned in the present invention does not exclude that other method steps may also be present before or after the combined steps or that other method steps may also be inserted between these explicitly mentioned steps, unless otherwise indicated; it is also to be understood that a combined connection between one or more devices/apparatus as referred to in the present application does not exclude that further devices/apparatus may be present before or after the combined device/apparatus or that further devices/apparatus may be interposed between two devices/apparatus explicitly referred to, unless otherwise indicated. Moreover, unless otherwise indicated, the numbering of the various method steps is merely a convenient tool for identifying the various method steps, and is not intended to limit the order in which the method steps are arranged or the scope of the invention in which the invention may be practiced, and changes or modifications in the relative relationship may be made without substantially changing the technical content.
Wherein the concentration of Cu, Pb and Zn ions in fecal bacteria is measured by inductively coupled plasma spectrometer (ICP, Optima 2000DV, USA) (test methods reference Xiaoang Cai, Guang-Jian Dai, Shao-Zao Tan, Yu Ouyang, You-Sheng Ouyang, Qi-Shan Shi. synergistic interaction information and ceramic ions loaded. alpha. -zirconium phosphate. materials letters.2012,67(1): 199-201.).
Example 1 method for preparing fecal bacterial liquid by removing copper, lead and zinc ions in human feces with lithium illite
(1) Preparation of lithium illite: adding 100g illite mineral (commercial product, particle size of 0.5-2.0 mm) into 1L water, adding 10g lithium metaphosphate, heating to 75 deg.C, adding 20g Na2CO3(ii) a Stirring for 3h at an electromagnetic speed of 60rpm, standing for 24h, centrifugally washing for 4 times by using deionized water, removing insoluble substances at the bottom, and recovering solids; drying at 105 ℃ for 48h to obtain lithium illite; the Cation Exchange Capacity (CEC) of the resulting Liulilite was 50mmol/100 g.
(2) Taking 30 +/-0.1 g of lithium illite (the particle size is 0.5-2.0 mm), adding 100mL of sterile 0.9% NaCl solution, stirring at 500r/min for 5min, and filtering out water by a stainless steel filter screen of 0.25mm to obtain lithium illite slurry for later use.
(3) Placing 50 +/-1 g of donor fresh excrement in a stirrer, adding 250mL of sterile 0.9% NaCl solution, adding the prepared lithium illite slurry (the particle size is 0.5-2.0 mm), and homogenizing at 2000r/min for 3min to obtain excrement slurry;
(4) filtering the excrement slurry by a stainless steel filter screen with the thickness of 5.0mm to remove large-particle substances in the excrement slurry, and obtaining excrement slurry after primary filtration;
(5) transporting the primarily filtered excrement slurry ice to a laboratory and processing the excrement slurry ice within 2 hours after collection;
(6) transferring the primarily filtered excrement slurry into a nitrogen bioengineering kitchen, introducing carbon dioxide gas, and stirring for 5min at a speed of 2000r/min in a homogenizer every 1h, and obtaining excrement and bacteria slurry after 6 h;
(7) filtering the fecal strain slurry by stainless steel filter screens with diameters of 2.0mm, 1.0mm, 0.5mm and 0.25mm step by step to remove Li-based illite, lithium carbonate, unabsorbed food residues and small particle substances which adsorb copper, lead and zinc ions, and obtaining coarse fecal strain liquid;
(8) the crude fecal bacterial liquid is centrifuged at 6000r/min for 15min, and the fecal bacterial pellet is resuspended in 250mL of 0.9% NaCl solution to obtain 250mL of nearly colorless and odorless fecal bacterial liquid.
The total content of copper, lead and zinc ions in the obtained fecal strain liquid is determined to be zero (the total content of copper, lead and zinc ions is less than 4ppb, which cannot be detected by an instrument), and the content of sodium ions is 154 mmol/L.
Blank control group: the procedure was the same as in the above-mentioned steps (1) to (8) of this example, except that 250mL of a nearly colorless and odorless liquid was obtained without adding feces and Liulite, and the total content of copper, lead and zinc ions in the liquid was determined to be zero (no instrument detection, total content of copper, lead and zinc ions < 4ppb), and sodium ion content was determined to be zero (see Table 1).
Comparative group 1 (with feces and without illite): the procedure was as in the above-mentioned steps (1) to (8) of this example except that no Lidikite was added to obtain 250mL of a nearly colorless and odorless liquid, and the contents of copper, lead and zinc ions in the liquid were measured as 1.3mmol/L, 3.1mmol/L and 1.6mmol/L, respectively, and the total content of copper, lead and zinc ions was measured as 6mmol/L and the content of sodium ions was measured as 154 mmol/L.
Comparative group 2 (illite with stool): the procedure was as in the above-mentioned steps (1) to (8) of this example except that no Likite was added to obtain 250mL of a nearly colorless and odorless liquid, and the contents of copper, lead and zinc ions in the liquid were measured to be 1.1mmol/L, 2.7mmol/L and 1.4mmol/L, respectively, and the total content of copper, lead and zinc ions was measured to be 5.2mmol/L and the content of sodium ions was measured to be 156.1mmol/L (see Table 1).
TABLE 1 results of the determination of the ion content in the different treatment groups
Note:ammol/L, mmol/L;
bN/A, which cannot be detected by instruments, and the content is less than 4ppb (ppb: mu g/L).
As can be seen from Table 1, the donor fresh feces treated by Lidikite and having excessive copper, lead and zinc ions no longer contain copper, lead and zinc ions, so that the feces donated by feces donors having excessive copper, lead and zinc ions can reach the standard after treatment, and the content of sodium ions is basically kept unchanged.
Embodiment 2 method for preparing fecal bacterial liquid by removing copper, lead and zinc ions in human feces through lithium illite
(1) Preparation of lithium illite: adding 120g of illite mineral (commercial product, particle size of 0.5-2.0 mm) into 1L of water, adding 8g of lithium metaphosphate, heating to 80 ℃, and adding 22g of Na2CO3(ii) a Stirring at an electromagnetic speed of 60rpm for 3.5h, standing for 20h, centrifuging and washing with deionized water for 4 times, removing insoluble substances at the bottom, and recovering solids; drying for 45h at 100 ℃ to obtain the Liulite; the Cation Exchange Capacity (CEC) of the resulting Liulilite was 60mmol/100 g.
(2) Placing 50 +/-1 g of donor fresh excrement in a stirrer, adding 150mL of sterile 0.95% NaCl solution, adding 25g of lithium-based illite (the particle size is 0.5-2.0 mm), and homogenizing at 2200r/min for 4min to obtain excrement slurry;
(3) filtering the excrement slurry by a stainless steel filter screen with the thickness of 5.0mm to remove large-particle substances in the excrement slurry, and obtaining excrement slurry after primary filtration;
(4) transporting the primarily filtered excrement slurry ice to a laboratory and processing the excrement slurry ice within 2 hours after collection;
(5) transferring the primarily filtered excrement slurry into a nitrogen bioengineering kitchen, introducing nitrogen gas, and stirring for 5min at a speed of 2000r/min in a homogenizer every 1.2h, and obtaining excrement and bacteria slurry after 4 h;
(6) filtering the fecal strain slurry by stainless steel filter screens with diameters of 2.0mm, 1.0mm, 0.5mm and 0.25mm step by step to remove Li-based illite, lithium carbonate, unabsorbed food residues and small particle substances which adsorb copper, lead and zinc ions, and obtaining coarse fecal strain liquid;
(7) the crude fecal strain liquid is centrifuged at 6500r/min for 18min, and the fecal strain precipitate is resuspended in 250mL of 0.9% NaCl solution to obtain 250mL of nearly colorless and odorless fecal strain liquid free of copper, lead and zinc ions.
Example 3 method for preparing fecal bacterial liquid by removing copper, lead and zinc ions in human feces with lithium illite
(1) Preparation of lithium illite: 80g of illite mineral (commercially available product, particle size 0.5-2.0 mm) was added to the mixtureAdding 12g of lithium metaphosphate into 1L of water, heating to 70 ℃, and adding 18g of Na2CO3(ii) a Stirring for 2.5h at an electromagnetic speed of 60rpm, standing for 30h, centrifuging and washing for 4 times by using deionized water, removing insoluble substances at the bottom, and recovering solids; drying at 110 ℃ for 55h to obtain the Lidikite; the Cation Exchange Capacity (CEC) of the resulting Liulilite was 50mmol/100 g.
(2) Placing 50 +/-1 g of donor fresh excrement in a stirrer, adding 350mL of sterile 0.85% NaCl solution, adding 75g of lithium illite (the particle size is 0.5-2.0 mm), and homogenizing at 2200r/min for 4min to obtain excrement slurry;
(3) filtering the excrement slurry by a stainless steel filter screen with the thickness of 5.0mm to remove large-particle substances in the excrement slurry, and obtaining excrement slurry after primary filtration;
(4) transporting the primarily filtered excrement slurry ice to a laboratory and processing the excrement slurry ice within 2 hours after collection;
(5) transferring the primarily filtered excrement slurry into a nitrogen bioengineering kitchen, introducing nitrogen gas, and stirring for 5min at a speed of 2000r/min in a homogenizer every 1.2h, and obtaining excrement and bacteria slurry after 4 h;
(6) filtering the fecal strain slurry by stainless steel filter screens with diameters of 2.0mm, 1.0mm, 0.5mm and 0.25mm step by step to remove Li-based illite, lithium carbonate, unabsorbed food residues and small particle substances which adsorb copper, lead and zinc ions, and obtaining coarse fecal strain liquid;
(7) the crude fecal strain liquid is centrifuged at 6500r/min for 12min, and the fecal strain precipitate is resuspended in 250mL of 0.9% NaCl solution to obtain 250mL of nearly colorless and odorless fecal strain liquid free of copper, lead and zinc ions.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Claims (9)
1. A method for preparing a fecal strain solution by removing copper, lead and zinc ions in human feces by utilizing lithium illite is characterized by comprising the following steps:
1) adding a sterile NaCl solution into the excrement sample, adding Likelite, and uniformly stirring to obtain excrement slurry;
2) primarily filtering the excrement slurry, taking filtrate, and stirring the filtrate at 1800-2200 rpm for 3-6 min every 0.8-1.2 h under the protection of carbon dioxide, nitrogen, helium, neon or argon, and obtaining the excrement and bacterium slurry after 4-7 h;
3) filtering the fecal strain slurry to obtain filtrate to obtain crude fecal strain liquid; centrifuging the crude fecal strain liquid, taking the precipitate, and re-suspending to obtain colorless and odorless fecal strain liquid containing no copper, lead and zinc ions;
the preparation method of the lithium illite comprises the following steps: adding water and lithium metaphosphate into illite ore, heating to 70-80 ℃, and adding Na2CO3(ii) a Stirring for 2.5-3.5 h, standing for 20-30 h, then centrifugally washing with deionized water, recovering solids, and drying to obtain the Lidikite.
2. The method of claim 1, wherein the illite mineral, lithium metaphosphate, Na2CO3And the dosage ratio of water is 80-120 g: 8-12 g: 18-22 g: 1L of the compound.
3. The method according to claim 1, wherein the cation exchange capacity of the obtained lithium illite is 40 to 60mmol/100 g.
4. The method according to claim 1, wherein in the step 1), the dosage ratio of the excrement sample, the NaCl solution and the Liulite is 1 g: 3-7 mL: 0.5 to 1.5 g.
5. The method according to claim 1, wherein the concentration of the NaCl solution is between 0.85% and 0.95% w/v.
6. The method according to claim 1, wherein in the step 1), the stirring speed is 1800-2200 rpm and the time is 2-4 min.
7. The method according to claim 1, wherein in step 2), the preliminary filtering is performed by: and filtering the excrement slurry by using a filter screen with the aperture of 4.5-5.5 mm to remove large-particle substances.
8. The method according to claim 1, wherein in the step 3), the specific operation of the fecal sewage sludge filtration is as follows: filtering by a filter screen with the aperture of 1.5-2.5 mm, 0.8-1.2 mm, 0.4-0.6 mm and 0.2-0.3 mm in sequence.
9. The method according to claim 1, wherein in the step 3), the rotation speed of the centrifugation is 5500-6500 rpm, and the time is 12-18 min.
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| CN104922158A (en) * | 2015-06-05 | 2015-09-23 | 中国人民解放军第三军医大学第三附属医院 | Fecal microbiota capsule, as well as preparation method and application thereof |
| CN104984736A (en) * | 2015-06-23 | 2015-10-21 | 中南大学 | Blood heavy metal ion adsorbent, preparation method thereof and blood perfusion device |
| CN106946485A (en) * | 2017-03-24 | 2017-07-14 | 吉林大学 | A kind of lithium is modified illite cement intensifier and preparation method |
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