CN100408669C - Oral biological film dynamic model device and its oral biological film forming method - Google Patents
Oral biological film dynamic model device and its oral biological film forming method Download PDFInfo
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Abstract
The present invention relates to a dynamic model device of an oral cavity biological film, which comprises an alkali liquid jar (1), a culture medium jar (2), a fermentation jar (3), constant flow cultivation chambers (4) and a sample adding bottle (7). The dynamic model device of the oral cavity biological film is characterized in that the constant flow cultivation chambers (4) are formed by connecting a plurality of vertical standard cylindrical constant flow cultivation chambers in series; the alkali liquid jar (1) is connected with the fermentation jar (3) through a peristaltic pump P1; the culture medium jar (2) is connected with the fermentation jar (3) through a peristaltic pump P2; the fermentation jar (3) leads in noble gases N2 and CO2 through a filter F2 and a peristaltic pump P4; the fermentation jar (3) outwards discharges waste liquid and waste gases through a peristaltic pump P5 and a filter F1; the fermentation jar (3) is connected with the constant flow cultivation chambers (4) through a peristaltic pump P6; each of the constant flow cultivation chambers (4) outwards discharge waste liquid and waste gases through a peristaltic pump P7 and a filter F3; the culture medium jar (2) and the sample adding bottle (7) are connected with the constant flow cultivation chambers (4) in parallel respectively through the peristaltic pumps P2, P3.
Description
One, technical field
The present invention relates to the method for a kind of oral biological film dynamic model device and formation oral biological film thereof, belong to microbial film oral medical model apparatus or stomatology field.
Two, background technology
As single celled life form, bacterium in research for a long time all the time to be free on unicellular state in the liquid nutrient medium as research object, simplified the influence of external environment so to the full extent, also obtained a large amount of achievements in research, for modern microbiology is laid a good foundation.Yet under state of nature, bacterium seldom is free on the form in the pure culture and exists with this.Microbial film is the modal existence form of bacterium, is meant at solid-state organic or inorganic dielectric surface, and a large amount of microorganisms are collected in the matrix that exocellular polysaccharide constitutes and crosslinked mutually and microorganism ecological environment that form.
In medical field, the microbial film disease has occupied the infected by microbes disease more than 80%.Microbial film is by the effect of its physics and chemistry barrier, the good ecotope of resisting outside stimulus is provided to pathogenic bacterium, and the gene conduction of development therein and engaging, thereby help also that the level transmission to drug resistant gene causes very big difficulty to traditional antibiotic therapy between different microorganisms.The principal disease dental caries disease of harm humans oral Health, its main paathogenic factor also are to be the main cariogenicity microbial film (plaque) that constitutes with the streptococcus mutans.
In the bacteriological study of dental caries disease, because the complicacy of oral cavity ecotope, the relation that will study certain or some bacterium and dental caries disease in vivo is unusual difficulty, therefore, set up the oral biological film model that to simulate mouthful interior environment, help learning research, obtain than the more accurate and comprehensive result of traditional bacteriology experiment the external oral microorganism that carries out high confidence level.Biological film model can external creation a kind of with body in proximate ecotope, the growth metabolism condition of regulation and control bacterium, make it both to have consistence with natural surroundings, the remarkable advantage that has stdn and Modulatory character again, carrying out the oral biology experiment with biological film model compares with simple experiment in vitro, confidence level is higher, and is repeatable better, is a kind of very potential laboratory facilities.
Biological film model system main purpose is to simulate as much as possible ecotope in the oral cavity, and this artificial environment is maintained at stable status.Associated biomolecule membrane modle (the Chinese stomatology magazine .1996 of report such as present domestic Liu Zheng; 31 (2): 110-112.) only be easy chemostat microbial culture system, though can satisfy basic microorganism continuous culture requirement, but its structure is numerous and jumbled, core component is cultivated for the fermentor tank continuous static, still can not effectively simulate the continual flowing environment of nutritive medium such as the inner saliva in oral cavity, and not good enough for the control of polluting; And external like product, equal Journal of Dental Research as Sissons, 1991,70 (11): the perseverance of foundation such as the 1409-16 report artificial oral system of multiple-workstation (MAM) and the Kinniment thing membrane modle (Microbiology that improves people's living condition, 1996,142,631-638), although control accuracy height to envrionment conditions, but its complex structure is corresponding also high to the accuracy requirement of peripherals, so it costs an arm and a leg, application cost is high, although to the good but still application that can't put it over of oral cavity environmental simulation.
The present inventor has set up many fungus strains biological film model of being made up of streptococcus mutans, blood suis, streptococcus-salivarius, actinomyces naeslundii etc. in early-stage Study, Chinese patent 03135320.7 is entitled as " method that oral biological film mould unit and oral biological film thereof form ", experimental results show that this model has reliability and repeatability preferably, can be used in the biomembranous every in vitro study in oral cavity.Yet, we find that also also there are some defectives in this model in the middle of experiment, and at first, this biological film model only can be observed the microbial film formation situation of specific in the model and unique time point, the gained data are limited, are difficult to realize the omnidistance of experimental microbial film forming process detected in real time.Next is that the core component constant flow culture of this model is an irregular profile, is difficult to accomplish stdn, and the clearance rate that is difficult to control each constant flow culture in experiment reaches consistent, makes each constant flow culture be difficult to accomplish the parallel comparable of real meaning.Therefore, the present inventor has carried out certain transformation to the constant flow culture of existing biological film model, use the device of standard profile instead, and be provided with a plurality of openings at its top as required, can place the sample injector of a plurality of fixed preparations, the probe of multiple proofing unit (as the pH registering instrument, the redox-potential registering instrument) also can be installed simultaneously.On this basis, developed the method for a kind of oral biological film dynamic model device and formation oral biological film, realized detecting in real time continuously to artificial bacterial plaque is biomembranous, a large amount of significant experimental results have been obtained, for the cause of disease of dental caries disease and pathogenesis, screening anticariogenic agent, assessment medicine preventing decayed tooth usefulness and research mechanism of drug action provide strong technology platform, not seeing by retrieval has similar bibliographical information.
Three, summary of the invention
The objective of the invention is to provide at the deficiencies in the prior art a kind of can carry out the oral biological film dynamic model device of continuous detecting and form the method for oral biological film artificial bacterial plaque microbial film, be characterized in a plurality of hydroxylapatite plates (HA Discs) are placed the continuous culture system that contains the quantitative experiment bacterial strain, regularly give sucrose solution to provide experiment bacterium anaerobic glycolysis required substrate, making bacterium form experimental microbial film on the hydroxylapatite surface is artificial bacterial plaque, different times in the microbial film forming process takes out hydroxylapatite plate then, detect its surface and form biomembranous every biological indicator, so that experimental biomembranous forming process is comprehensively detected, realization detects in real time to various anticariogenic agent intervention experiment microbial films and sick form omnidistance of dental caries, and then the action effect of definite anticariogenic agent and mechanism of action.
Purpose of the present invention is realized by following technical measures:
The oral biological film dynamic model device contains alkali liquid tank, culture tank, fermentor tank, constant flow culture and application of sample bottle.Wherein constant flow culture is that a plurality of vertical standard cylindrical culturing room is connected in series, and alkali liquid tank is connected with fermentor tank by peristaltic pump, and culture tank is connected with fermentor tank by peristaltic pump, and fermentor tank is introduced rare gas element N by strainer and peristaltic pump
2And CO
2Fermentor tank discharges waste liquid, waste gas by peristaltic pump and strainer, fermentor tank is connected with constant flow culture by peristaltic pump, each constant flow culture is outwards held in both hands out waste liquid, waste gas by peristaltic pump and strainer, and culture tank is connected with a plurality of constant flow culture by peristaltic pump respectively with the application of sample bottle.
Fermentor tank is an interlayer, establishes following water-bath import and outlet in the interlayer, jar built-in pH control electrode, and fermentor tank places on the magnetic stirrer.
Constant flow culture is vertical standard cylindrical, the constant flow culture top is provided with a plurality of sample injectors, on a plurality of sample injectors, place a plurality of fixed preparations according to requirement of experiment, fixed preparation is a hydroxylapatite, sample injector vertically is positioned over constant flow culture inside, establish hyperchannel pH registering instrument electrode and waste liquid, outlet port in the constant flow culture, constant flow culture is placed in the constant incubator constant temperature and hatches, and provides stirring velocity about 30rpm with the dynamic environment of salivary flow in the simulation oral environment by magnetic stirrer.
Hydroxylapatite plate is provided by oral cavity biomedical engineering key lab of West China College of Stomatology Sichuan University investigation of materials chamber.
The formation method of oral biological film
1, after the aseptic connection of dynamic model device is finished, fermentor tank is placed on the magnetic stirrer, obtain the stirring velocity of 30-100ppm, temperature is maintained about 37 ℃ by the recirculated water bath cabinet, alkali liquid tank provides basic solution that the liquid phase PH valve of bacteria suspension in the fermentor tank is maintained about 7.0, the adding of basic solution is regulated by pH controller and peristaltic pump, treats that the stable back of each bacteria growing is standby.
2, fill the main source of nutrition of artificial saliva substratum BM-5 in the culture tank, add by peristaltic pump constant speed in fermentor tank and constant flow culture as experimental bacteria.
3, fill anticariogenic agent and sucrose solution in the application of sample bottle, add in constant flow culture by divider and peristaltic pump control, to provide the experiment microbial film to cause the substrate sucrose of dental caries and the anticariogenic agent of being studied, can adopt different medicines and drug level according to research purpose.
4, constant flow culture places in the constant incubator of sealing, provides stirring velocity about 30ppm with the dynamic environment of salivary flow in the simulation oral environment by magnetic stirrer.
5, according to requirement of experiment, in constant flow culture, place a plurality of sample injectors, in experimentation, the timed interval as requested, take out sample injector continuously, detect biomembranous every biological indicator that the hydroxylapatite surface forms on the sample injector, till whole sample injectors are exhausted, thereby realized vertical continuous real-time monitoring experimental microbial film forming process.
The present invention has following advantage:
1, can effectively simulate mouthful interior ecotope and also stably form experimental microbial film, verify its application usefulness by big quantity research at cariology and science of dental materials.
2, adopt hyperchannel culture technique in parallel, difference that can more different simultaneously microbial film treatment measures has also greatly been simplified workflow, is more convenient for using.
3, can take out sample continuously by a plurality of time points in experimentation, detect the formation situation of sample surface biological film.According to requirement of experiment the quantity of sample is set, can realizes experimental biomembranous whole process is monitored in real time.
4, the major equipment of selecting for use is product commonly used on the market, and cost control is good, compares with international like product, has goodish price advantage.
Thereby 5, adopt airtight training method to control the generation of polluting in model effectively.
Four, description of drawings
Fig. 1 is an oral biological film dynamic model device synoptic diagram
1, alkali liquid tank 2, culture tank 3, fermentor tank 4, vertical standard cylindrical constant flow culture 5, hydroxylapatite plate 6, sample injector 7, application of sample bottle 8, magnetic stirrer 9, constant water bath box 10, hyperchannel pH survey meter 11, magnetic stirrer 12, waste liquid outlet 13, waste gas outlet
P1-P7 is a peristaltic pump
F1-F3 is a filter
Fig. 2 is vertical standard cylindrical constant flow culture structural representation
Fig. 3 is the dynamic change of each treatment group pH of constant flow culture liquid phase
Fig. 4 is each each bacteria growing curve of treatment group hydroxylapatite surface biological film of different time
A, different time treatment group HAdisc surface biological film S.mutans growth curve
B, different time treatment group HAdisc surface biological film S.sanguis growth curve
C, different time treatment group HAdisc surface biological film A.naeslundii growth curve
D, different time treatment group HAdisc surface biological film L.rhamnosusi growth curve
Fig. 5 is each each bacterium pie graph of treatment group hydroxylapatite surface biological film of different time
A, the bacterial count of different time 25mM sucrose group HAdisc surface biological film
B, the bacterial count of different time 4000ppm Turkey-galls group HAdisc surface biological film
C, the bacterial count of different time distilled water group HAdisc surface biological film
Fig. 6 is that the fluorescent microscope of experimental group microbial film form is seen FM * 200
A:24 hour, B:48 hour, C:72 hour, D:96 hour, E:120 hour
The fluorescent microscope of the negative control group microbial film of Fig. 7 form is seen FM * 200
A:24 hour, B:48 hour, C:72 hour, D:96 hour, E:120 hour
Fig. 8 is that the fluorescent microscope of blank group microbial film form is seen FM * 200
A:24 hour, B:48 hour, C:72 hour, D:96 hour, E:120 hour
Five, embodiment
Below by embodiment the present invention is carried out concrete description; be necessary to be pointed out that at this present embodiment only is used for the present invention is further specified; but can not be interpreted as limiting the scope of the invention, the person skilled in the art in this field can make some nonessential improvement and adjustment according to the content of the invention described above.
Embodiment:
The oral biological film dynamic model device contains alkali liquid tank 1, culture tank 2, fermentor tank 3, constant flow culture 4 and application of sample bottle 7 shown in Fig. 1-2.Alkali liquid tank 1 is connected with fermentor tank 3 by peristaltic pump P1, and wherein, constant flow culture is that a plurality of vertical standard cylindrical constant flow culture are connected in series.Culture tank 2 is connected with fermentor tank 3 by peristaltic pump P2, and fermentor tank 3 is introduced rare gas element N by filter F2 and peristaltic pump P4
2And CO
2Fermentor tank 3 is by peristaltic pump P5 and filter F1 effluent discharge, waste gas, fermentor tank 3 is connected with a plurality of constant flow culture 4 in parallel by peristaltic pump P6, each constant flow culture 4 discharges waste liquid, waste gas by peristaltic pump P7 and filter F 3, and culture tank 2 and application of sample bottle 7 are connected in parallel by peristaltic pump P2, P3 and a plurality of constant flow culture 4 respectively.It is 37 ℃ that fermentor tank 3 keeps a jar interior temperature by circulator bath, places the pH controller in jar, keeps liquid phase PH valve about 7.0, and fermentor tank 3 places on the magnetic stirrer 8.Vertically be placed with a plurality of sample injectors 6 in each constant flow culture 4, be fixed with hydroxylapatite plate 5 on each sample injector 6, hyperchannel pH survey record instrument electrode 10 and waste liquid relief outlet 12, outlet port 13 are installed in the constant flow culture 4 simultaneously, constant flow culture 4 is positioned over constant incubator 9 interior 37 ℃ of constant temperature and hatches, and provides the stirring velocity about 30rpm to simulate the flow circuit of saliva in the oral environment by magnetic stirrer 11.Constant flow culture and sample injector useable glass, plastics or metal are made.Hydroxylapatite plate is made by oral cavity biomedical engineering key lab of West China College of Stomatology Sichuan University investigation of materials chamber and is provided.
The formation method of oral biological film:
1, after the aseptic connection of dynamic model device is finished, fermentor tank 3 is placed on the magnetic stirrer 8, obtain the stirring velocity of 30-100ppm, temperature is maintained about 37 ℃ by the recirculated water bath cabinet, alkali liquid tank 1 provides basic solution that the liquid phase PH valve of bacteria suspension in the fermentor tank is maintained about 7.0, the adding of basic solution is regulated by pH controller and peristaltic pump P1, treats that the stable back of each bacteria growing is standby.
2, fill the main source of nutrition of artificial saliva substratum BM-5 in the culture tank 2, add by peristaltic pump P2 constant speed in fermentor tank 3 and each constant flow culture 4 as experimental bacteria.
3, fill anticariogenic agent and sucrose solution in the application of sample bottle 7, add in each constant flow culture 4 by divider and peristaltic pump P3 control, to provide the experiment microbial film to cause the substrate sucrose of dental caries and the anticariogenic agent of being studied, can adopt different medicines and drug level according to research purpose.
4, constant flow culture 4 places in the constant incubator of sealing, provides stirring velocity about 30ppm with the dynamic environment of salivary flow in the simulation oral environment by magnetic stirrer 11.
5, according to requirement of experiment, in constant flow culture, place a plurality of sample injectors 6, in experimentation, the timed interval as requested, take out sample injector 6 continuously, detect biomembranous every biological indicators that hydroxylapatite 5 surfaces form on the sample injector, till whole sample injectors are exhausted, thereby realized vertical continuous real-time monitoring experimental microbial film forming process.
Below be the application example of this mould unit in cariology research:
1. experimental strain
Streptococcus mutans Streptococcus mutans ATCC 25175
Streptococcus sanguis Streptococcus sanguis ATCC 10556
Actinomyces naeslundii Actinomyces naeslundii WVU 627
Lactobacillus Lactobacillus rhamnosus AC 413
Above bacterial strain provides by School of Stomatology oral cavity, West China biomedical engineering key lab.
2. culture condition
2.1 fermentor cultivation condition
Temperature: keep by the recirculated water bath cabinet for 37 ℃;
Stir: low speed by magnetic stirring apparatus 8 controls, makes bacteria suspension mixing in the fermentor tank;
Anaerobic environment: 95%N
2, 5%CO
2, flow velocity 10ml/min is controlled by peristaltic pump P4;
PH: maintain about 0.7 with 0.1N NaOH solution, regulate by Ph controller and peristaltic pump Pl;
Remove filter: D=0.1/hr is controlled by peristaltic pump P5.
2.2 constant flow culture culture condition
Temperature: keep by the recirculated water bath cabinet for 37 ℃
Stir: low speed by magnetic stirring apparatus 11 controls, provides the shearing force that needs in the microbial film forming process
Clearance rate: D=0.75/hr is controlled by peristaltic pump P7
Give sample: after each experimental bacteria growth reaches and stablizes in the constant flow culture, gave sample 1 time in per 12 hours, be 1/10 of the continuous culture systems cubic capacity of biological symphysis to the sample amount at every turn, and application of sample is by peristaltic pump P3 and divider control.
3. experiment flow
This experiment is the biomembranous formation of simulation dental surface bacterial plaque, adopts hydroxylapatite plate (HA Discs) to place and contains the continuous culture system of testing hybrid bacterial strain, to form many fungus strains microbial film on its surface.Fermentor tank in the model is the bank of experiment mixed bacterium, and several parallel constant flow culture that link to each other with fermentor tank are the biological film formed place of experiment.
During the experiment beginning, streptococcus mutans, lactobacillus, Streptococcus sanguis and the actinomyces naeslundii that to recover in the TPY solid medium go down to posterity, through carrying out cultured continuously in the artificial saliva substratum that is inoculated into fermentor tank after the biochemical identification, its pH value is by pH controller control balance, and fresh artificial saliva substratum is added continuously by peristaltic pump.After the basis cultured continuously reaches and stablizes, mixed bacterium suspension in the fermentor tank is joined in each parallel constant flow culture by peristaltic pump, in each constant flow culture, add fresh artificial saliva substratum simultaneously, the ratio of bacterium and fresh culture is 1: 9 (v/v), stop inoculated bacteria after 24 hours, continue to add fresh artificial saliva and to each constant flow culture, carry out cultured continuously.Every day is to the detection of growing of the bacterium in the constant flow culture liquid phase environment, treat mixed bacterium cultured continuously in each constant flow culture reach stable after (continuous three days no significant differences), in each constant flow culture, insert a plurality of sample injectors that are fixed with HA Discs, HA Discs is immersed in the mixed bacterium suspension fully, simultaneously treatment soln was added in the constant flow culture (1 time/12 hours, each add volume be the constant flow culture liquor capacity 1/10), after this, from constant flow culture, take out a sample injector continuously according to 24 hours the timed intervals, the microbial film that HA Discs surface is formed carries out bacterial count, structure observation is till the sample injector in constant flow culture is exhausted.
4. artificial saliva
The artificial saliva prescription that this experiment is adopted is microbial film improved culture medium BM-5:
III type hog gastric mucin (Hog gastric mucin, Type III, Sigma) 2.50g/l
Peptide peptone (Proteose peptone, Fluka) 2.00g/l
Pancreatin decomposes casein peptone (Peptone from casein, tryptic digest, Fluka) 1.00g/l
Yeast extract (Yeast extract, Oxoid) 1.00g/l
Repone K (KCl) 2.50g/l
Glucose (Glucose) 0.50g/l
Cystine hydrochloride (Cystiene-Hydrochloride, Sigma) 0.10g/l
Protohemine (Heamin, Shanghai east wind Biochem Technology, INC.) 1.00mg/l
Sichuan University's doctorate paper 14
Dipotassium hydrogen phosphate (K2HPO43H2O) 0.114g/l
Potassium primary phosphate (KH2PO4) 0.20g/l
Adjusting its pH value with 1N NaOH solution is 7.5,121 ℃, 15psi, sterilization 70min.
5. experimental result example
This example is for studying the natural drug Turkey-galls to testing biomembranous influence
A tests grouping
According to difference, be divided into following each group to application of sample in the constant flow culture:
Negative control group: the 25mM sucrose solution, the final concentration that adds in the constant flow culture is 2.5mM.
The blank group: the distilled water that does not contain any carbohydrate is as blank.
Experimental group: contain the 25mM sucrose solution of the Galla Chinensis extract of 4mg/ml, add constant current and cultivate the back medicine
Final concentration is 0.4mg/ml.
The dynamic change of each treatment group pH of B constant flow culture liquid phase
As shown in Figure 3, in the process of 8 dosings, cyclic fluctuation had appearred in the pH value after positive controls added the sucrose solution that concentration is 25mmol/L, about 3 hours, touch the bottom, minimum pH drops to below 4.5, As time goes on, pH rises gradually, returns to the level before the sugaring in about 3 hours later on.In experimental group, reflect similar pH Changing Pattern equally.Along with gallic adding, slowly descending appears in pH, touches the bottom in about 3 hours, slowly rises subsequently, returns to the preceding level of dosing after 6 hours, about about pH7.0.In the whole culture of continuous cultivation, distilled water group pH does not see obvious fluctuation, maintains about pH7.5.
HAdic surface experiment microbial film live bacterial count in the C constant flow culture
In the microbial film culture cycle, different pharmaceutical is handled different time, different types of microbial film bacteria planting influence difference.The Turkey-galls group shows with the distilled water group bacteria planting stationary phase similar with the sucrose group (about 72 hours), microbial film bacterial count in 24 hours and microbial film bacterial count in 120 hours have significant difference (P＜0.05), for each experimental strain, the Turkey-galls group shows the effect of lactobacillus obvious suppression, its bacterial count of whole culture cycle is all lower, streptococcus mutans also there is certain inhibition, actinomyces naeslundii, blood suis are suppressed not obvious.To 120 hours microbial films, the Turkey-galls group was compared with the sucrose group, and lactobacillus is obviously suppressed (P＜0.05), and streptococcus mutans also is suppressed to a certain extent, and actinomyces naeslundii, blood suis suppress not obvious.The microbial film bacterium is constituted, though each experimental bacteria is sticked at microbial film certain difference is arranged quantitatively, each group is not seen have significant change (P＞0.05) on bacterium constitutes.
The fluorescence microscope of each treatment group experiment microbial film form of D
Biomembranous FM observes and finds that along with the increase of cultured continuously time, each organizes the increase that microbial film all shows bacterium and exocellular polysaccharide matrix, but each treatment group shows as diverse microbial film form outward appearance.Compare with the sucrose group, even at 120 hours microbial films, the Turkey-galls group does not see that fine and close bacterial polysaccharides matrix agglomerate agglomerating, in blocks in a large number, microbial film present comparatively loose outward appearance yet.
Claims (3)
1. oral biological film dynamic model device, contain alkali liquid tank (1), culture tank (2), fermentor tank (3), constant flow culture (4) and application of sample bottle (7), it is characterized in that constant flow culture (4) is connected in series for a plurality of vertical standard cylindrical culturing room, alkali liquid tank (1) is connected with fermentor tank (3) by peristaltic pump P1, culture tank (2) is connected with fermentor tank (3) by peristaltic pump P2, and fermentor tank (3) is introduced rare gas element N by filter F 2 and peristaltic pump
2And CO
2Fermentor tank (3) discharges waste liquid, waste gas by peristaltic pump P5 and filter F 1, fermentor tank (3) is connected with constant flow culture (4) by peristaltic pump P6, each constant flow culture (4) discharges waste liquid, waste gas by peristaltic pump P7 and filter F 3, and culture tank (2) and application of sample bottle (7) are connected in parallel by peristaltic pump P2, P3 and a plurality of constant flow culture (4) respectively; Wherein constant flow culture (4) is vertical standard cylindrical, the top of each constant flow culture (4) is provided with a plurality of sample injectors (6), can on a plurality of sample injectors, place a plurality of fixed preparations according to requirement of experiment, fixed preparation is a hydroxyapatite, sample injector (6) vertically is positioned over constant flow culture (4) inside, establish hyperchannel pH registering instrument (10) electrode in the constant flow culture (4), waste liquid relief outlet (12), outlet port (13), constant flow culture (4) is placed on the interior constant temperature of constant incubator and hatches, and the dynamic flow environment of the stirring velocity of 30rpm with saliva in the simulation oral environment is provided by magnetic stirrer (11).
2. oral biological film dynamic model device according to claim 1 is characterized in that fermentor tank (3) is an interlayer, establishes circulator bath import and outlet in the interlayer, jar built-in pH control electrode, and fermentor tank (3) places on the magnetic stirrer (8).
3. the oral biological film dynamic model device forms the method for oral biological film as claimed in claim 1 or 2, it is characterized in that this method contains following steps:
1) after the dynamic model device connection is finished, fermentor tank (3) is placed on the magnetic stirrer (8), obtain the rotating speed of 30～100rpm, temperature remains on 37 ℃ by circulator bath, alkali liquid tank (1) provides alkali lye to make the interior pH=7.0 of fermentor tank, regulate by pH controller and peristaltic pump P1, treat that the stable back of each bacteria growing is standby;
2) fill artificial saliva substratum BM-5 in the culture tank (2),, add by peristaltic pump P2 constant speed in fermentor tank (3) and each constant flow culture (4) as the main source of nutrition of experimental bacteria;
3) fill medicine and sucrose solution in the application of sample bottle (7), the experiment microbial film provides the substrate sucrose that causes dental caries and the anticariogenic agent of required research in each constant flow culture (4), can adopt different medicines and concentration according to the difference of research purpose;
4) constant flow culture (4) places in the crossing current incubator of sealing, and the dynamic environment of the stirring velocity of 30rpm with salivary flow in the simulation oral environment is provided by magnetic stirrer (11);
5) according to requirement of experiment, in each constant flow culture (4), place a plurality of sample injectors (6), in experimentation, the timed interval is as requested taken out sample injector (6) continuously, check upward biomembranous every biological indicator of hydroxylapatite surface formation of sample injector (6), till whole sample injectors (6) are exhausted, thereby realized vertical continuous real-time monitoring to experimental microbial film forming process.
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| CN101935611B (en) * | 2010-08-24 | 2015-04-22 | 好来化工(中山)有限公司 | Biomembrane culture and flow chamber in artificial mouth simulation device |
| CN101935612B (en) * | 2010-08-24 | 2013-10-16 | 好来化工(中山)有限公司 | Flow cell of manual oral cavity model used for culturing biological film |
| CN101948746B (en) * | 2010-09-09 | 2013-07-10 | 好来化工(中山)有限公司 | Artificial mouth simulating system device |
| CN101948745B (en) * | 2010-09-09 | 2013-06-26 | 好来化工(中山)有限公司 | Artificial mouth simulating device |
| CN107012159B (en) * | 2017-05-12 | 2019-06-21 | 四川大学 | A kind of streptococcus mutans biomembrane pH indicating gage and its application based on gene recombination plasmid |
| CN107418891A (en) * | 2017-09-20 | 2017-12-01 | 四川省人民医院 | A kind of perfusion type bladder bacterial biof iotalm dynamic is incubated simulation system |
| CN111349548A (en) * | 2020-05-06 | 2020-06-30 | 山西医纽恩医疗科技有限公司 | In-vitro fermentation system and evaluation system applying same |
| CN117264818B (en) * | 2023-09-15 | 2024-05-17 | 中山大学附属口腔医院 | Oral bacteria biological film culture medium and application thereof |
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| 口腔生物膜体外模型的建立和应用评估. 黄正蔚等.牙体牙髓牙周病学杂志，第14卷第9期. 2004 |
| 口腔生物膜体外模型的建立和应用评估. 黄正蔚等.牙体牙髓牙周病学杂志，第14卷第9期. 2004 * |
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