CN1173161C - Sampling diluting device for bio-reaction process in-line testing system - Google Patents
Sampling diluting device for bio-reaction process in-line testing system Download PDFInfo
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- CN1173161C CN1173161C CNB021383766A CN02138376A CN1173161C CN 1173161 C CN1173161 C CN 1173161C CN B021383766 A CNB021383766 A CN B021383766A CN 02138376 A CN02138376 A CN 02138376A CN 1173161 C CN1173161 C CN 1173161C
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Abstract
The present invention relates to a sampling and diluting device of an on-line detection system for the biological reaction process, more specifically to a micro sampling device and a cascade diluting device. A sample is taken and diluted in the mode of continuous flow and intermittent detection. A ceramic membrane filter piece is arranged at the end of a sampling probe, reaction liquid is filtered in situ in a membrane piece in a tank, the cross flow penetration is carried out, penetrating liquid is diverted and led out by threads of a diversion clamp sleeve outside a membrane, and the automatic micro sampling in the tank from the tank outside is realized. The cascade diluting device is mainly composed of a vertical three-stage cascade diluting plate with an overflowing cavity. Fluid flows by the fluid gravity, and the sample and dilute liquid stably flow by a micro complete flow path structure of an organic material. The sample taking amount of the device is little, the optimizing control of the biological reaction process can not be affected, the dilution ratio is easily controlled, and the present invention has the advantages of convenient and flexible operation, easy manufacture and low cost. Main members can be manufactured into standard members, and are easily cleaned and changed. The sample representation is good, and the present invention is suitable for the application of the on-line detection and the micro complete analysis (mu-TAS) of the biological reaction process.
Description
Technical field
The present invention relates to a kind of sampling air mix facilities of bioprocesses on-line detecting system, match with biology sensor and microcomputer, be specifically related to comprise trace sampling apparatus and cascade dilution device to realize the online detection of material concentration in the bioprocesses.
Prior art
Current fermentation industry is except traditional fermentation, and the fermenting and producing direction to microbiotic, amino acid, nucleic acid etc. develops.In recent years, just produce bioactivator in technology such as reorganization of research and utilization microorganism and cellular incubation.Be accompanied by above-mentioned evolution, the control of sweat and optimization are also from management develops to the direction with computer automation by experience.
Various physics and chemical sensor are widely used in the control of process.Detected temperatures, pH, dissolved oxygen DO, carbon monoxide, pressure, flow, the isoparametric sensor of agitating function are connected with microcomputer, have constituted the control system that is used for biochemical and sweat.But it can only be controlled biochemical reaction indirectly, can not be directly acquainted with the situation of change of material in the complicated biochemical process.And biology sensor has solved many species analysis of carrying out of being difficult in the past, can measure the concentration that direct representative sweat carries out the raw material and the tunning of state.Through the exploitation of 20 years of researches, biology sensor has formed the class of a uniqueness, gets up side by side with physical sensors and chemical sensor, regards the 3rd classification of sensor as.
Biology sensor has characteristics such as selectivity is good, enzyme can be repeatedly used, analysis speed is fast, accuracy is high, operating system is fairly simple, directly controls bioprocesses for microcomputer and has prepared feasible sensor condition.But, yet there are no report so far and can in general bioprocesses, realize online detection of biological, chemical quantitative analysis system.Biological, chemical amount can't realize online detection, and tracing sth. to its source is biology sensor and the detection interfacing of complicated bioprocesses can't reach a standard, and this has become " bottleneck " technology that biology sensor is realized online detection.
Bioprocesses on-line detecting system synoptic diagram comprises microscale sampler, diluter, microbiosensor group, bioengineering Data Management Analysis software and microcomputer as shown in Figure 1.And microscale sampler and diluter the technical problem to be solved in the present invention just.
In research work, the experimenter adopts the automatic small fermentation tank to carry out the optimization of technological condition for fermentation, generally use the canister of 2-30L volume, and sample volume is up to 15ml, repeatedly sampling causes fermentating liquid volume to reduce, sweat is brought bigger influence, can cause the error of process optimization to increase.So press for a kind of trace sampling apparatus of design, reduce sample volume, reduce the interference of sampling to detecting as far as possible.
Present existing air mix facilities, as mechanical arm, the FIA ON-LINE dilution technology is for the on-line determination of enriched sample provides one of effective method; Yet need time controller, computer control valve and the higher peristaltic pump of precision, thereby the machining requirement is high, costs an arm and a leg, the control of dilution ratio is dumb, has had a strong impact on biology sensor and has used in the online detection of bioprocesses.
Summary of the invention
(1) technical matters that will solve
A kind of trace sampling apparatus of bioprocesses on-line detecting system is provided, require to solve outside jar to carrying out the technical matters of microsampling in the fermentation tank, the heat labile characteristics of biology sensor had both been overcome, and overcome the influence of oxygen concentration and temperature to bioelectrode, reduced sample volume again, reduce the interference of sampling as far as possible, the needs of keep online serial sampling, intermittently measuring to detecting.
A kind of cascade dilution device of bioprocesses on-line detecting system also is provided, requires to solve the technical matters that to carry out the on-line continuous dilution.Adopt damping fluid that sample is carried out tens dilutions to hundred times, control dilution rate flexibly, to reach the range of linearity that biology sensor detects.Another effect of dilution is to regulate the sample pH value value, makes it coincidence senser working pH value, reduces and disturbs.
(2) technical scheme
Trace sampling apparatus
Fig. 2 is the trace sampling apparatus structural representation, and it is made up of a sampling probe and sample flow control section.The probe end is the membrane filtration part, and the other end is the penetrating fluid fairlead and be connected hermetic unit with jar.Sampling probe is made up of probe right cylinder, membrane filtration spare, the outer water conservancy diversion chuck of film, water conservancy diversion connector, fairlead, sleeve pipe.The flow control part is carried out flow control jointly by needle valve, metering valve.Connect by fairlead between sampling probe, needle valve, the metering valve.
Membrane filtration spare adopts ceramic membrane spare, and membrane filtration spare is a tubulose, and rete is at inwall, supporting layer outside, internal diameter is 6-10mm, external diameter is 10-14mm.The long 8-12mm of membrane filtration spare, membrane aperture 0.1-0.2 μ m.
The outer water conservancy diversion chuck of film, horizontal drilling on the vertical cylindrical end of placing of probe is the outer water conservancy diversion chuck of film.Water conservancy diversion chuck inwall is made screw-like outside film, is used for water conservancy diversion, and the difference of Major Diam and path is less than 2mm.Membrane filtration spare places the outer water conservancy diversion chuck boring of the cylindrical film of probe, and the outer water conservancy diversion chuck internal thread of film contacts with membrane filtration spare outer wall, and outer water conservancy diversion chuck of film and membrane filtration spare adopt interference fit, two ends, cooperation place silica gel sealing.The probe right cylinder is selected polyvinylidene fluoride or teflon material for use.
Open the cylindrical aperture of a conical head in the middle of the end disc that pop one's head in right cylinder and sleeve pipe join, the aperture column part has internal thread, and aperture communicates with water conservancy diversion chuck middle part, at the built-in water conservancy diversion connector of aperture.The water conservancy diversion connector is the small column of a band conical head, and the columnar portion of small column sets up external thread separately, and water conservancy diversion connector and probe right cylinder be with threaded engagement, the conical head of water conservancy diversion connector near the outer water conservancy diversion chuck of film to reduce dead volume.Fairlead is inserted in perforate in the middle of water conservancy diversion connector post cone.The water conservancy diversion connector is selected the teflon material for use.
Fairlead is the elasticity of plastics pipe, selects silicone elastic tube or polyvinylidene fluoride elastic tube for use.
One of sleeve pipe is provided with the external thread of internal thread and probe right cylinder one end with threaded engagement.Sleeve pipe is selected stainless steel for use, with the protection fairlead.
The cross-flow filtration infiltration technology is adopted in sampling, flows through film spare with in position filtrator by reactant liquor, directly filters in jar, utilizes film spare to realize separating of fermentation liquor and solid, and film spare adopts the screw thread water conservancy diversion outward, directly handles continuous filtration after drawing filtrate.Control permeate flow with valve, the diffusing capacity of film equates with permeate flow during steady state (SS), and penetrating fluid is drawn continuously.Get penetrating fluid at regular intervals as measuring samples, promptly intermittently measure, directly do not drain as the penetrating fluid of measuring samples.
Design parameter is: permeate flow 0.4ml/min, retardation time is less than 1min, sampler pipeline volume≤0.7ml, sampling amount 10-1000 μ l/min.
The cascade dilution device
The interface schematic flow sheet of three-stage cascade device in Fig. 3 on-line detecting system.The cascade dilution device is made up of the vertical three-stage cascade dilution plate in damping fluid overflow portion and carry potential overflow chamber.The damping fluid overflow portion comprises damping fluid storage tank, constant flow pump, three damping fluid overflow grooves, and the damping fluid flow is by the damping fluid Flux Valve Control on the damping fluid stream.Fig. 4 is the structural representation of the vertical three-stage cascade dilution plate in carry potential overflow chamber.Structure is: sample flow path and damping fluid stream by the carry potential overflow chamber merge into the diluted mixture stream, as the one-level diluter, three one-level diluters vertically are composed in series three-stage cascade dilution plate, first order diluted mixture stream and second level spill cavity are connected, second level diluted mixture stream and third level spill cavity are connected.First order diluted mixture liquid is continued dilution as partial sample, the sample of partial diluted mixture liquid as the third level diluted again, the diluted mixture liquid of the third level is detected as test sample.The spill cavity bottom is provided with tapping valve, and the spill cavity top is provided with air hole.Sample flow is by the control of the sample flow variable valve on the sample flow path, and the cascade dilution plate is selected organic material for use, as polyvinylidene fluoride or teflon.Fluid flowing path adopts and micro-total analysis system (the little full flow structure of μ-TAS) adapt, fully shorten the residence time of sample, but unlikely formation drop, fluid flows and relies on the gravity of fluid self, the static pressure that had both fully kept fluid, the least possible generation dynamic pressure, all operations must be that continous-stable flows, serial sampling, serial dilution, employing are intermittently measured.
The fluid flowing path caliber is 0.1-1mm, and the diluted mixture flow path length is 5-10cm, and the spill cavity chamber is 3-6mm deeply.The single-stage dilution ratio is controlled at 4-10, and dilution error is not more than 1%, three grade of dilution ratio and is up to 1000, and dilution error is not more than 2.5%.
(3) beneficial effect
The trace sampling apparatus of the present invention's design, the characteristics of three-stage cascade air mix facilities maximum are microsampling, Microdilution, trace for microbiosensor detects, its amount of samples is 10-100 μ L/min, fermented 100 hours, the sample loss amount of serial sampling only is 60-600ml also, can satisfy the requirement that the sampling of small-sized fermentation jar detects.
Trace sampling apparatus replaces manual sampling mode with automatic sampling mode, adopt the cross-flow filtration technology, utilize film spare to realize separating of fermentation liquor and solid, the screw thread water conservancy diversion is adopted in the film spare outside, as long as reactant liquor flows through in film spare pipe, will be flowed in the chuck screw thread by the filter membranous layer infiltration, formation cross-flow infiltration, realize the first in first out of liquid, the liquid that promptly responds flows in the film spare pipe, will see penetrating fluid and draw.
Adopt filtrator in position, directly in jar, filter, take out filtrate as sample, its advantage is: do not increase the complicacy of jar, at jar probe place dress filtering membrane spare is installed, can not increase the microbiological contamination chance, film spare and pipeline are easy to clean, and are standard component, can in time change.
The cascade dilution device has designed the operation that relies on the fluid self gravitation, be the stability that dilute sample and buffering flow quantity are controlled in overflow, need not time controller, computer control valve and high-precision peristaltic pump, easy to operate, flexible, be fit to the diluted sample requirement of the online detection of bioprocesses.The little full flow structure of its three grades of dilutions and fluid flowing path makes sample form stable continuous and flows, and dilution rate does not change with the change of sample or damping fluid flow or changes very little.
The method that on-line detecting system adopts continuously sampling automatically, dilutes, intermittently measures.The flow process of serial sampling dilution is to allow sample flow path sampling line and dilution pipeline according to this, and sample is in continuous flow, and its flow is controlled easily, do not need the dead sample of emptying, and memory effect is little, and is little to the sample representativeness influence.Serial sampling, dilution system manufacturing are simple, cheap, sample representativeness is good, are very suitable for on-line analysis and detect and use.Can further reduce biological respinse and control cost, improve the precision that biology sensor detects.
Description of drawings
Fig. 1 bioprocesses on-line detecting system synoptic diagram.
Fig. 2 trace sampling apparatus structural representation.(a) little sample sampler front elevational schematic: (b) sampling probe part schematic side view.
The interface schematic flow sheet of three-stage cascade device in Fig. 3 on-line detecting system.
The structural representation of the vertical three-stage cascade dilution plate in Fig. 4 carry potential overflow chamber.
1 probe right cylinder in the accompanying drawing, 2 membrane filtration spares, the outer water conservancy diversion chuck of 3 films, 4 water conservancy diversion connectors, 5 fairleads, 6 covers close, 7 needle valves, 8 metering valves, 9 spill cavities, 10 sample flow path, 11 sample flow variable valve, 12 tapping valves, 13 air holes, 14 damping fluid flow control valves, 15 damping fluid streams, 16 diluted mixture streams.
Embodiment
Embodiment 1
Trace sampling apparatus: membrane filtration spare adopts the ceramic membrane spare of certain company.Film spare is a tubulose, and rete is at inwall, supporting layer outside, inner diameter d 1=8mm, outside diameter d 2=12mm.Pressure drop is about 10KPa during filtration, and membrane aperture is 0.2 μ m, and requiring flow of filtrate is 0.4ml/min, gets the long 10mm of being of film spare, calculates according to correlation formula, can satisfy the permeate flow requirement.
The membrane aperture of membrane filtration spare is 0.1 μ m, and all the other conditions also can satisfy the permeate flow requirement with embodiment 1.
Three-stage cascade dilution plate current body stream length overall is 90mm, the stream caliber is 1mm, then cumulative volume is 0.09ml, consider that with the 0.1ml/min flow its residence time only is 0.9min, sampling 10-1000L/min, the single-stage dilution ratio is controlled at 4-10, and the single-stage dilution error is not more than 1%, three grades of dilution ratios are up to 1000, and dilution error is not more than 2.5%.Sample glucose, its original concentration are 278.54g/l, carry out three grades of dilution tests, and its dilution accuracy test result is:
The test result of table 1 three-stage cascade dilution precision
Three grades of total dilution ratio error rate % of dilution of one-level dilution secondary dilution
Mean concentration 52.23 10.89 2.45
Extension rate 5.33 4.80 4.44 113.69 0.6
Mean concentration 47.42 6.65 0.94
Extension rate 5.87 7.13 7.07 296.32 1.2
Mean concentration 28.83 3.30 0.45
Extension rate 9.66 8.74 7.33 618.98 1.8
Mean concentration 28.83 2.88 0.28
Extension rate 9.66 10.00 10.35 1,000 2.5
Claims (2)
1, a kind of trace sampling apparatus that is used for the online detection of bioprocesses, it is characterized in that forming by a sampling probe and sample flow control section, the probe end is the membrane filtration part, the other end is the penetrating fluid fairlead and be connected hermetic unit with jar, sampling probe is by probe right cylinder (1), membrane filtration spare (2), the outer water conservancy diversion chuck (3) of film, water conservancy diversion connector (4), fairlead (5), sleeve pipe (6) is formed, the flow control part is by needle valve (7), metering valve (8) carries out flow control, sampling probe jointly, needle valve, connect by fairlead between the metering valve;
Membrane filtration spare (2) adopts ceramic membrane spare, and membrane filtration spare is a tubulose, and rete is at inwall, supporting layer outside, internal diameter is 6-10mm, external diameter is 10-14mm, the long 8-12mm of membrane filtration spare, membrane aperture 0.1-0.2 μ m,
The outer water conservancy diversion chuck (3) of film, horizontal drilling on the end of the vertical probe right cylinder of placing (1), be the outer water conservancy diversion chuck (3) of film, water conservancy diversion chuck (3) inwall is made screw-like outside film, is used for water conservancy diversion, and membrane filtration spare (2) places the outer water conservancy diversion chuck boring of the film of probe right cylinder (1), the outer water conservancy diversion chuck internal thread of film contacts with membrane filtration spare (2) outer wall, outer water conservancy diversion chuck (3) of film and membrane filtration spare (2) adopt interference fit, two ends, cooperation place silica gel sealing
In the middle of the end disc that pop one's head in right cylinder and sleeve pipe join, open the cylindrical aperture of a conical head, the aperture column part has internal thread, aperture communicates with the outer water conservancy diversion chuck (3) of film middle part, at the built-in water conservancy diversion connector of aperture (4), water conservancy diversion connector (4) is the small column of a band conical head, the columnar portion of small column sets up external thread separately, water conservancy diversion connector (4) and probe right cylinder (1) are with threaded engagement, the conical head of water conservancy diversion connector (4) is near the outer water conservancy diversion chuck (3) of film, perforate in the middle of water conservancy diversion connector (4) post cone, insert fairlead (5), (6) one of sleeve pipes are provided with the external thread of internal thread and probe right cylinder (1) one end with threaded engagement.
2, on-line detecting system trace sampling apparatus as claimed in claim 1, the right cylinder (1) that it is characterized in that popping one's head in is selected polyvinylidene fluoride or teflon material for use, water conservancy diversion connector (4) is selected the teflon material for use, fairlead (5) is selected silicone elastic tube or polyvinylidene fluoride elastic tube for use, and sleeve pipe (6) is selected stainless steel for use.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB021383766A CN1173161C (en) | 2002-09-27 | 2002-09-27 | Sampling diluting device for bio-reaction process in-line testing system |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB021383766A CN1173161C (en) | 2002-09-27 | 2002-09-27 | Sampling diluting device for bio-reaction process in-line testing system |
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| CN1402009A CN1402009A (en) | 2003-03-12 |
| CN1173161C true CN1173161C (en) | 2004-10-27 |
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Families Citing this family (8)
| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2009132476A1 (en) * | 2008-04-30 | 2009-11-05 | 西门子公司 | Sampling device, detection system and method for culture components |
| CN101571460B (en) * | 2009-06-18 | 2011-08-24 | 中国原子能科学研究院 | Sampling device of small samples |
| CN102279118A (en) * | 2011-05-05 | 2011-12-14 | 山东省科学院生物研究所 | Concentration gradient diffusion sampler for bioreactor on-line testing system |
| CN103212230B (en) * | 2013-03-29 | 2015-07-15 | 成都易态科技有限公司 | Liquid cross flow filtration method and component and special flow guide mandrel bar |
| DE102014106918A1 (en) * | 2013-05-28 | 2014-12-04 | Endress + Hauser Conducta Gesellschaft für Mess- und Regeltechnik mbH + Co. KG | Method and apparatus for the automated execution of affinity-based assays |
| CN103913563B (en) * | 2014-04-09 | 2015-12-30 | 上海岗崎控制仪表有限公司 | A kind of three-phase biologic matter sensor and filtering and turbulent method |
| CN104111248A (en) * | 2014-07-03 | 2014-10-22 | 中国科学院东北地理与农业生态研究所 | Sampling needle used for determining elements in soil filtrate |
| CN105890923B (en) * | 2014-12-11 | 2019-08-09 | 山东省科学院生物研究所 | A kind of sterile micro on-line period device and sampling method |
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