CN101871927B - Online monitoring equipment and method for acute toxicity of water quality - Google Patents
Online monitoring equipment and method for acute toxicity of water quality Download PDFInfo
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- CN101871927B CN101871927B CN2010102000794A CN201010200079A CN101871927B CN 101871927 B CN101871927 B CN 101871927B CN 2010102000794 A CN2010102000794 A CN 2010102000794A CN 201010200079 A CN201010200079 A CN 201010200079A CN 101871927 B CN101871927 B CN 101871927B
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Abstract
The invention discloses online monitoring equipment for acute toxicity of water quality. The equipment comprises a luminescent bacteria continuous culture system, a sample feeding system, a liquid draining system, a photoelectric detection system and a data display system, and can realize automatic operation. The invention also discloses a method for performing online monitoring on the acute toxicity of the water quality by utilizing the equipment. The online monitoring equipment for the acute toxicity of the water quality of the invention has the characteristics of simple structure, low cost and convenient operation, and can be operated and maintained by non-professionals only after a simple training. By using the method, continuous and automatic acute toxicity monitoring of the water quality can be realized completely; sensitivity is equivalent to that of an international luminescent bacteria method; and monitoring steps are simplified greatly. By using the equipment and the method, an early warning is provided for a sudden environment accident and corresponding measures are taken timely so as to reduce loss greatly.
Description
Technical field
The present invention relates to a kind of Apparatus and method for of acute toxicity of water quality on-line monitoring, relate in particular to a kind of equipment and method of the acute toxicity of water quality on-line monitoring based on the immobilization luminous bacteria biosensor, belong to the environmental monitoring field.
Background technology
Along with the aggravation of development of industry and agriculture and mankind's activity, Sudden Pollution Accident takes place frequently in recent years, because lack online early warning measure, not only causes great economic loss to the locality, also can cause casualties.For the early warning and the management of the protection of strengthening river and drinking water source, industrial wastewater discharge etc., launch respectively relevant regulations in the world, require to realize the continuous toxicity control of water quality.But traditional toxicity test is all long because of the time, expense is high, complex operation, the not early warning of suitable unexpected environmental accident.Therefore be badly in need of a kind of quick, easy, economic detection method so that reference is provided to the decision maker.
Along with the development of environmental microbiology, adopt microorganism to analyze water quality toxicity as indicator organism more, wherein photobacteria is suitable for on-line analysis and application owing to have higher sensitivity and easy to operate.Abroad begin one's study and use the photobacteria testing environment from middle 1960s; The eighties, the Beckman company of the U.S. developed the pertinent instruments and the reagent of Microtox inspection technology, just issued photobacteria detection method of toxicity (GT/T15441-1995) in China up to the mid-90.But they need a series of blank of preparation and titer in measurement, complicated operating process, consuming time, loaded down with trivial details can only under lab detect, and detect and can not be used for on-the-spot continuous in-situ robotization, have greatly weakened the application of this method.So utilizing the advantage of photogen toxicity detection on monitoring water environment to develop different photogen sensors, domestic and international research personnel are used for on-the-spot continuous in-situ robotization detection; Develop the Lumistox300 of Deltatox water quality toxicity detector, Hach company etc. such as U.S. SDI company; They adopt the photogen bacterium liquid of cryopreservation are recovered the back as detecting liquid; This false positive occurs with regard to running into the luminous shakiness of photogen unavoidably; Developed the photogen immobilization technology so have in recent years; Such as the oak Ridge National Laboratory of the U.S. and Tennessee university joint research and development based on noctilcent toxicity detection chip, the mycoderm that agar is fixing combines detection with circuit; The researcher of Israel and Korea S then adopts the structure of fibre-optical probe, is fixed in the bare fibre top to photobacteria, or photobacteria is fixed in the sleeve pipe that connects optical fiber.In more domestic researchs photobacteria absorption is fixed on the composite fibre film, sensitive membrane directly links to each other with silicon photo diode.But use several problems that face: photobacteria is from generation to generation very short, can not guarantee the luminous of its long-term stability; Photonasty is relatively poor when inserting the fibre-optical probe detection in the gel; Can not reuse etc.
Summary of the invention
Technical matters to be solved by this invention provides a kind of equipment of the acute toxicity of water quality on-line monitoring based on the immobilization luminous bacteria biosensor.
The technical matters that the present invention also will solve provides the method for the said equipment to the acute toxicity of water quality on-line monitoring.
For solving the problems of the technologies described above, the technical scheme that the present invention adopts is following:
A kind of equipment of acute toxicity of water quality on-line monitoring, it comprises photobacteria continuous culture system, sample adding system, drainage system, program control system, photodetector system, data processing and transmission system and data presentation system;
The photobacteria continuous culture system is made up of nutrient solution deposit bottle, reservoir, pump, reaction tube and flexible pipe; Nutrient solution deposit bottle upside down is connected with reservoir through flexible pipe; Reservoir is connected with reaction tube through flexible pipe; And by the nutrient solution flow acceleration of pump control reservoir to reaction tube 8; But reaction tube places in the darkroom of folding, is placed with 1～10 luminous bacteria biosensor in the reaction tube;
Sample adding system is made up of sample slot, pump and flexible pipe; Sample slot is connected with reaction tube in the photobacteria continuous culture system through flexible pipe; And by the flow acceleration of pump control sample to reaction tube, the flexible pipe that inserts in the reaction tube will be near the bottom of reaction tube;
Drainage system is made up of waste liquid tank, pump and flexible pipe; Waste liquid tank is connected with reaction tube in the photobacteria continuous culture system through flexible pipe; And by the mass rate of emission of the waste liquid after detection finishes in the pump control reaction tube to waste liquid tank, the flexible pipe that inserts in the reaction tube will be near the bottom of reaction tube;
Photodetector system is arranged on reaction tube bottom, and data processing is connected with photodetector system through circuit with transmission system, calculates relative luminous intensity automatically, and with data storage in data storage card, data result shows in data presentation system;
Photobacteria continuous culture system, sample adding system, drainage system, photodetector system, data processing and transmission system and data presentation system all are to realize full automatic working by program control system control.
Wherein, said luminous bacteria biosensor is that diameter is a gelled pill; Prepare by following method: photobacteria bacteria suspension and sodium alginate soln are mixed, mixed liquor is dropwise splashed in the calcium chloride solution, solidify.
The preparation method of above-mentioned luminous bacteria biosensor comprises the steps:
(1) inclined-plane photobacteria bacterial classification inoculation was cultivated 15～18 hours in the ASW nutrient culture media, cultivation temperature is 18～22 ℃, and rotating speed is 180～250r/min;
(2) get the bacteria suspension that step (1) makes, being diluted to optical density is 0.1～0.6, dilution is added in the sodium alginate soln of sterilized cooling, under 18～22 ℃, mixes;
(3) mixed liquor that step (3) is made dropwise splashes in the calcium chloride solution with syringe, solidifies then 4～6 hours, cleans with distilled water, promptly gets.
In the step (2), preferably condition is, bacteria suspension, and being diluted to optical density is 0.1～0.2.
In the step (2), the concentration of described sodium alginate soln is 40～80g/L, preferred 40～50g/L.
In the step (2), the volume ratio of said dilution and said sodium alginate soln is 1: 1～10, preferred 1: 1～2.
In the step (3), the concentration of described calcium chloride solution is 20～30g/L, preferred 20～23g/L.
In the step (3), the volume ratio of said mixed liquor and said calcium chloride solution is 2～3: 100.
The store method of above-mentioned luminous bacteria biosensor is stored in luminous bacteria biosensor in the liquid storage under 4 ℃ of conditions, and said liquid storage is for containing 30～40g/L NaCl and 0.5～1g/L CaCl
2The WS.
What wherein, described photodetector system used can be any photoelectric sensor.
Utilize the said equipment to carry out the method for acute toxicity of water quality on-line monitoring; In reaction tube, put into 3～5 luminous bacteria biosensors, utilize the photobacteria continuous culture system in reaction tube, to add nutrient solution, utilize sample adding system in reaction tube, to add sample again; Photodetector system detects luminous intensity; Data processing and the transmission system image data line data of going forward side by side is handled, detect finish after, by drainage system the waste liquid in the reaction tube is entered waste liquid tank; Luminous bacteria biosensor continues to be retained in the operation that repeats to add photobacteria nutrient culture media, adding sample, detection luminous intensity, image data and discharge opeing in the reaction tube; After the continuous monitoring 7～10 days, the luminous bacteria biosensor that more renews continues on-line monitoring.
Beneficial effect: acute toxicity of water quality on-Line Monitor Device of the present invention is simple in structure, with low cost, easy to operate, and the layman only need pass through simple training and get final product operation and maintenance; This method can realize the monitoring of water quality continuous and automatic acute toxicity fully, and the photobacteria method of sensitivity and GB is suitable, simplifies monitoring step greatly.Use these apparatus and method to provide early warning to take corresponding measure timely, can greatly reduce the loss as the outburst surroundings accident.
Description of drawings
Fig. 1 is the structural representation of acute toxicity of water quality on-line monitoring equipment of the present invention.Wherein, the 1st, nutrient solution deposit bottle, the 2nd, reservoir, the 3rd, sample slot, the 4th, waste liquid tank, the 5th, pump, the 6th, pump, the 7th, pump, the 8th, reaction tube, the 9th, photodetector system, the 10th, data presentation system, the 11st, luminous bacteria biosensor.
Fig. 2 is the luminous intensity figure after the luminous bacteria biosensor recovery of the present invention.
Fig. 3 is that luminous bacteria biosensor of the present invention is to the continuous operational effect figure of distilled water.
Fig. 4 is the response diagram of luminous bacteria biosensor of the present invention to pH.
Fig. 5 is the response diagram of luminous bacteria biosensor of the present invention to toxicant.
Fig. 6 is luminous bacteria biosensor reappearance experiment of the present invention.
Embodiment
According to following embodiment, can understand the present invention better.Yet, those skilled in the art will readily understand that embodiment is described only to be used to explain the present invention, and the present invention that should also can not limit in claims to be described in detail.
Embodiment 1: the preparation of luminous bacteria biosensor.
Inclined-plane photobacteria bacterial classification inoculation was cultivated about 16 hours in the ASW fluid nutrient medium, and cultivation temperature is 20 ℃, and rotating speed is 200r/min; Get above-mentioned bacteria suspension, carry out 50 times dilution, optical density adds dilution in the 40g/L sodium alginate soln of sterilized cooling about 0.1, and the volume ratio of dilution and sodium alginate soln is 1: 1, at 20 ℃ of abundant mixings; Above-mentioned mixed liquor dropwise splashed in the 20g/L calcium chloride solution with syringe solidified 4 hours, the volume ratio of mixed liquor and calcium chloride solution is 2%, contains 30g/L NaCl and 0.5g/L CaCl with being soaked in after the distilled water cleaning
2The WS in, be placed on 4 ℃ and store for future use.
The photobacteria sensitive element of preserving is placed on 20 ℃ environment recovery 10 minutes (said recovery is from 4 ℃ of processes of taking out under the environment that is placed on 20 ℃ with sensitive element) down, and its luminous intensity is as shown in Figure 2.
Embodiment 2: acute toxicity of water quality on-line monitoring equipment.
As shown in Figure 1, acute toxicity of water quality on-line monitoring equipment comprises photobacteria continuous culture system, sample adding system, drainage system, program control system, photodetector system, data processing and transmission system and data presentation system.The photobacteria continuous culture system is made up of nutrient solution deposit bottle 1, reservoir 2, pump 7, reaction tube 8 and flexible pipe; Nutrient solution deposit bottle 1, upside down is connected with reservoir 2 through flexible pipe, and reservoir 2 is connected with reaction tube 8 through flexible pipe, and by the nutrient solution flow acceleration of pump 7 control reservoirs 2 to reaction tube 8, but reaction tube 8 places in the darkroom of folding.Sample adding system is made up of sample slot 3, pump 6 and flexible pipe; Sample slot 3 is connected with reaction tube 8 in the photobacteria continuous culture system through flexible pipe; And by the flow acceleration of pump 6 control samples to reaction tube 8, the flexible pipe that inserts in the reaction tube 8 will be near the bottom of reaction tube 8.Drainage system is made up of waste liquid tank 4, pump 5 and flexible pipe; Waste liquid tank 4 is connected with reaction tube 8 in the photobacteria continuous culture system through flexible pipe; And by the mass rate of emission of the waste liquid after detection finishes in the pump 5 control reaction tubes 8 to waste liquid tank 4, the flexible pipe that inserts in the reaction tube 8 will be near the bottom of reaction tube 8.Photodetector system is arranged on reaction tube 8 bottoms.Data processing is connected with photodetector system through circuit with transmission system; Automatically calculate relative luminous intensity; And with data storage in the SD card; Data result shows in data presentation system, for example adds reagent, sample, 5 minutes, 10 minutes, 15 minutes luminous intensity and relative luminous intensity.Photobacteria continuous culture system, sample adding system, drainage system, photodetector system, data processing and transmission system and data presentation system all are to realize full automatic working by program control system control.
In the nutrient solution deposit bottle 1 nutrient solution is housed, nutrient solution comprises following component: the nutrient culture media 10% (v/v) of photobacteria, CaCl
25g/L, NaCl 300g/L, solvent are water.
Wherein, the nutrient media components of photobacteria is (g/mL) as follows: Tryptone 0.5%, Yeast extract 0.5%, and NaCl 3%, Na
2HPO
40.5%, KH
2PO
40.1%, glycerine 0.3%, solvent are water.
During use, in reaction tube, put into 3～5 luminous bacteria biosensors, utilize the photobacteria continuous culture system in reaction tube, to add 35 μ L nutrient solutions with the inventive method preparation; Utilize sample adding system in reaction tube, to add 315 μ L samples again; Photodetector system detects luminous intensity, and per 5 minutes, 10 minutes and 15 minutes, the image data line data processing of going forward side by side respectively of data processing and transmission system; After detecting end; By drainage system the waste liquid in the reaction tube is entered waste liquid tank, luminous bacteria biosensor continues to be retained in the operation that repeats to add photobacteria nutrient culture media, adding sample, detection luminous intensity, image data and discharge opeing in the reaction tube, and continuous monitoring is after 7～10 days; The luminous bacteria biosensor that more renews continues on-line monitoring.
Embodiment 3: luminous bacteria biosensor is to the continuous operation of distilled water.
Adopt the continuous monitoring of embodiment 2 described acute toxicity of water quality on-line monitoring equipment to distilled water, the effect of monitoring is as shown in Figure 3.Explanation is along with the carrying out of test; The upgrowth situation of bacterium can change thereupon; Cause it to press down the light rate and change, but except that individual point, can in the regular hour, keep relative stablizing; Can calculate and set dynamic alarming line and eliminate this influence according to the survey historical data, make calculating more accurate.
Embodiment 4: luminous bacteria biosensor is to the response of pH.
Adopt embodiment 2 described acute toxicity of water quality on-line monitoring equipment, the distilled water water sample that changes the sample of sample slot into different pH when proceeding to 100 round-robin is tested successively, and its effect is as shown in Figure 4.Explain that biological elements of the present invention has the pH tolerance range of a broad, the influence that can resist soda acid in the environment.
Embodiment 5: luminous bacteria biosensor is to the response of toxicant.
Adopt embodiment 2 described acute toxicity of water quality on-line monitoring equipment, the zinc sulfate water sample that changes the sample of sample slot into variable concentrations when proceeding to 100 round-robin is tested successively, and its effect is as shown in Figure 5.Explain that the present invention can make response by the sensitive toxicant to variable concentrations, and recovery that can be very fast prolongs along with the raising of exposure concentrations accordingly release time, for follow-up test provides guarantee.
Embodiment 6: the experiment of luminous bacteria biosensor reappearance.
Carried out 7 days test with distilled water as sample, the parameter of test is 35 μ L nutrient solutions, 315 μ L samples, and the time interval is 2 minutes, 15 minutes reaction time, application of sample discharge opeing automatically continuously, the effect of test is as shown in Figure 6.Explanation bead in the process of follow-on test can move the time in a week continuously and not disintegrate; And luminous value also is stabilized in the good range; Along with the prolongation of test duration, thereby the decay that self can take place makes and presses down the light rate and rise gradually, but does not influence the carrying out of test.
Claims (3)
1. the equipment of an acute toxicity of water quality on-line monitoring is characterized in that it comprises photobacteria continuous culture system, sample adding system, drainage system, program control system, photodetector system, data processing and transmission system and data presentation system;
The photobacteria continuous culture system is made up of nutrient solution deposit bottle (1), reservoir (2), the 3rd pump (7), reaction tube (8) and flexible pipe; Nutrient solution deposit bottle (1) upside down is connected with reservoir (2) through flexible pipe; Reservoir (2) is connected with reaction tube (8) through flexible pipe; And by the nutrient solution flow acceleration of the 3rd pump (7) control reservoir (2) to reaction tube (8); Reaction tube (8) but place in the darkroom of folding, be placed with 1～10 luminous bacteria biosensor (11) in the reaction tube (8);
Sample adding system is made up of sample slot (3), second pump (6) and flexible pipe; Sample slot (3) is connected with reaction tube (8) in the photobacteria continuous culture system through flexible pipe; And by the flow acceleration of second pump (6) control sample to reaction tube (8), the flexible pipe that inserts in the reaction tube (8) will be near the bottom of reaction tube (8);
Drainage system is made up of waste liquid tank (4), first pump (5) and flexible pipe; Waste liquid tank (4) is connected with reaction tube (8) in the photobacteria continuous culture system through flexible pipe; And by the mass rate of emission of the waste liquid after detection finishes in first pump (5) the control reaction tube (8) to waste liquid tank (4), the flexible pipe that inserts in the reaction tube (8) will be near the bottom of reaction tube (8);
Photodetector system (9) is arranged on reaction tube (8) bottom; Data processing is connected with photodetector system (9) through circuit with transmission system; Automatically calculate relative luminous intensity, and with data storage in data storage card, data result shows in data presentation system (10) lining;
Photobacteria continuous culture system, sample adding system, drainage system, photodetector system, data processing and transmission system and data presentation system all are to realize full automatic working by program control system control.
2. the equipment of acute toxicity of water quality on-line monitoring according to claim 1 is characterized in that said luminous bacteria biosensor (11) is a gelled pill; Prepare by following method: photobacteria bacteria suspension and sodium alginate soln are mixed, mixed liquor is dropwise splashed in the calcium chloride solution, solidify.
3. utilize the described equipment of claim 1 to carry out the method for acute toxicity of water quality on-line monitoring; It is characterized in that in reaction tube, putting into 1～10 luminous bacteria biosensor, utilize the photobacteria continuous culture system in reaction tube, to add nutrient solution, utilize sample adding system in reaction tube, to add sample again; Photodetector system detects luminous intensity; Data processing and the transmission system image data line data of going forward side by side is handled, detect finish after, by drainage system the waste liquid in the reaction tube is entered waste liquid tank; Luminous bacteria biosensor continues to be retained in the operation that repeats to add photobacteria nutrient culture media, adding sample, detection luminous intensity, image data and discharge opeing in the reaction tube; After the continuous monitoring 7～10 days, the luminous bacteria biosensor that more renews continues on-line monitoring.
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| CN102539640A (en) * | 2010-12-17 | 2012-07-04 | 中国科学院生态环境研究中心 | Method used for testing high throughput acute toxicity of early life stage of fish |
| CN102175606B (en) * | 2011-01-19 | 2012-07-11 | 西安建筑科技大学 | Method for detecting acute biological toxicity of sewage |
| CN103076318A (en) * | 2012-12-27 | 2013-05-01 | 力合科技（湖南）股份有限公司 | Toxicity detection device |
| CN103645163B (en) * | 2013-12-20 | 2016-01-27 | 北京城市排水集团有限责任公司 | The continuous on-line monitoring Apparatus and method for of resurgent water acute toxicity of water quality |
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| CN104480007B (en) * | 2014-12-05 | 2016-03-30 | 力合科技（湖南）股份有限公司 | A kind of storage facilities of water quality biological toxotest bacterial classification and cultural method |
| CN106442483A (en) * | 2016-11-22 | 2017-02-22 | 暨南大学 | Luminous bacterium flow injection method for quickly detecting and warning food-borne toxin pollution and application of luminous bacterium flow injection method |
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|---|---|---|---|---|
| CN101131384A (en) * | 2007-09-27 | 2008-02-27 | 上海交通大学 | Method for detecting subaqueous acute biological toxicity using photobacteria |
| CN101218499A (en) * | 2005-05-26 | 2008-07-09 | 希姆陶克斯有限公司 | Water monitoring system |
| CN101477056A (en) * | 2009-01-22 | 2009-07-08 | 浙江清华长三角研究院 | Multichannel photobacteria on-line water quality toxicity monitoring device and monitoring method |
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| AT505306B1 (en) * | 2007-07-09 | 2008-12-15 | Mbonline Gmbh | DEVICE FOR MONITORING WATER TO MICROBIAL CASES |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101218499A (en) * | 2005-05-26 | 2008-07-09 | 希姆陶克斯有限公司 | Water monitoring system |
| CN101131384A (en) * | 2007-09-27 | 2008-02-27 | 上海交通大学 | Method for detecting subaqueous acute biological toxicity using photobacteria |
| CN101477056A (en) * | 2009-01-22 | 2009-07-08 | 浙江清华长三角研究院 | Multichannel photobacteria on-line water quality toxicity monitoring device and monitoring method |
Non-Patent Citations (1)
| Title |
|---|
| 彭强辉等.水质生物毒性在线监测技术研究进展.《环境监测管理与技术》.2009,第21卷(第4期),第12-16页. * |
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