Method and device for on-line biological toxicity detection of water quality by lactic acid bacteria ph inhibition

Abstract

The application discloses a kind of lactic acid bacteria PH inhibition method water quality online biological toxicity detection method and detection device, wherein detection method includes the following steps: preparation first water sample and second water sample;Get the initial PH voltage value of first water sample and second water sample;Constant-temperature culture is carried out to first water sample and second water sample;Respectively obtain the PH voltage value of first water sample and second water sample after cultivation;Biological toxicity analysis calculation is carried out to water sample to be measured.The method of the application determines the size of harmful substances in water by monitoring the PH change caused by lactic acid, a metabolic product of lactic acid bacteria;Based on the mature glass electrode electrochemical principle method to detect PH, its detection accuracy is not affected by water quality turbidity;The detection method of the application has high consistency with the luminescent bacteria method of prior art for detecting biological toxicity, and meets the related technical requirements in the determination of water quality acute toxicity.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological toxicity of water environmental pollutants, and particularly relates to a lactic acid bacteria PH inhibition method for online biological toxicity detection of water quality and a detection device. BACKGROUND

[0002] At present, most of the online biological toxicity analyzers on the market use luminescent bacteria (Fisher arc bacteria or bright luminescent bacteria) for detection. The culture and long-term preservation of luminescent bacteria have relatively harsh conditions, and water turbidity can also affect accuracy.

[0003] In view of the above problems, the skilled in the art have begun to study the use of lactic acid bacteria for microbial toxicity monitoring. For example, a Chinese patent application with the application number 201910729523.2 discloses a sewage biological toxicity monitoring method, which includes the following steps: S1, selecting 10 kinds of microorganisms including lactic acid bacteria, Escherichia coli, sulfur bacteria, nitrous acid bacteria, nitrous acid bacteria, denitrifying bacteria, actinomycetes, bacillus, rotifer and vorticella as the characteristic community of the sewage treatment plant; S2, selecting the influent COD concentration, influent total nitrogen concentration, influent water temperature, influent dissolved oxygen content, color, SS, sludge age and sludge concentration as environmental factors, and classifying all sewage treatment plants according to the selected environmental factors; S3, establishing a reference group for each type of sewage treatment plant obtained in step S2; S4, when a certain sewage treatment plant in each type of sewage treatment plant is unstable, the community structure is determined and compared with the standard community structure of the reference group of each type of sewage treatment plant, and the sewage treatment plant is diagnosed to analyze the reason from the microbial level and provide a basis for the repair of the sewage treatment plant. Although this method uses lactic acid bacteria to evaluate the sewage treatment capacity of the sewage treatment plant and monitor the effluent quality, the existing sewage biological toxicity monitoring method has high detection cost and complex detection method. SUMMARY

[0004] The present application aims to provide a lactic acid bacteria PH inhibition method for online biological toxicity detection of water quality and a detection device to solve the problems raised in the background art.

[0005] To achieve the above-mentioned purpose, the present application provides a lactic acid bacteria PH inhibition method for online biological toxicity detection of water quality, which includes the following steps:

[0006] S1, preparing a first water sample and a second water sample;

[0007] S2, obtaining the initial PH voltage value of the first water sample and the second water sample, respectively;

[0008] S3, incubating the first water sample and the second water sample at a constant temperature;

[0009] S4, respectively, obtain the PH voltage value of the first water sample and the second water sample after culture;

[0010] S5, biological toxicity analysis and calculation of the water sample to be tested.

[0011] Further, the specific steps of preparing the water sample in step S1 are:

[0012] S1.1, cleaning the culture tube: the control system controls the multi-way valve and peristaltic pump to add a certain amount of distilled water to the first culture tube and the second culture tube, respectively, to clean the first culture tube, the second culture tube and the PH probe in the first culture tube and the second culture tube;

[0013] S1.2, adding culture solution: the control system controls to add equal amount of lactic acid bacteria culture solution to the first culture tube and the second culture tube, respectively;

[0014] S1.3, the control system controls to add equal amount of distilled water to the first culture tube as the added lactic acid bacteria culture solution;

[0015] S1.4, the control system controls to add equal amount of water sample to be tested to the second culture tube as the added lactic acid bacteria culture solution;

[0016] S1.5, adding bacteria: the control system controls to add equal amount of lactic acid bacteria to the first culture tube and the second culture tube as the added lactic acid bacteria culture solution, and mix uniformly to prepare the first water sample and the second water sample.

[0017] Further, in step S2, the control system reads the PH voltage value of the first PH sensor and the second PH sensor, thereby obtaining the initial PH voltage value A1 of the first water sample and the initial PH voltage value B1 of the second water sample.

[0018] Further, in step S3, the temperature of the first culture tube and the second culture tube is kept constant at 25-37℃, and the constant temperature culture time is 15-25min.

[0019] Further, in step S4, after constant temperature culture, the control system reads the PH voltage value of the first PH sensor and the second PH sensor again, thereby obtaining the PH voltage value A2 of the first water sample after culture and the PH voltage value B2 of the second water sample.

[0020] Further, step S5 is specifically: using the following formula to calculate the inhibition rate of the water sample to be tested:

[0021]

[0022] According to the calculation result, the water sample to be measured is evaluated for toxicity; if CF is between 15% and 20%, it is mild toxicity; if CF is between 20% and 35%, it is moderate toxicity; and if CF is greater than 35%, it is severe toxicity.

[0023] The application also provides a lactic acid bacteria PH inhibition method water quality online biological toxicity detection device, the detection device includes a multi-way valve, a peristaltic pump, a first culture tube, a second culture tube, a metering unit, a positive sample bottle, a distilled water bottle, a water sample bottle to be measured, a waste liquid barrel, a lactic acid bacteria bottle, a culture solution bottle, a first PH sensor, a second PH sensor, a constant temperature module and a control system, the multi-way valve is connected with the first culture tube, the second culture tube, the metering unit, the positive sample bottle, the distilled water bottle, the water sample bottle to be measured, the waste liquid barrel, the lactic acid bacteria bottle and the culture solution bottle respectively, the first PH sensor and the second PH sensor are arranged in the first culture tube and the second culture tube respectively, the constant temperature module is used for constant temperature culture of the first culture tube and the second culture tube, the peristaltic pump is connected with the multi-way valve through the metering unit, and the control system is connected with the multi-way valve, the peristaltic pump, the metering unit, the first PH sensor, the second PH sensor and the constant temperature module respectively.

[0024] Compared with the prior art, the application has the following beneficial effects:

[0025] (1) The lactic acid bacteria PH inhibition method water quality online biological toxicity detection method of the application, by culturing lactic acid bacteria in the culture tube, the metabolic products of the lactic acid bacteria will produce acid as the lactic acid bacteria rapidly multiply and grow, the amount of the metabolic products of the lactic acid bacteria will be affected when the lactic acid bacteria meet harmful substances, the influence of the toxic substances on the growth of the lactic acid bacteria is determined by detecting the change of the PH in the culture tube, and the size of the harmful substances in the water sample to be measured is determined by monitoring the PH change caused by the metabolic product lactic acid of the lactic acid bacteria. The method has the characteristics of high detection precision, low use cost and simple maintenance.

[0026] (2) The lactic acid bacteria PH inhibition method water quality online biological toxicity detection method of the application detects the PH based on the mature glass electrode electrochemical principle method, and the detection precision is not affected by the turbidity of the water quality; the method has high consistency with the existing technology of the luminescent bacteria method for detecting biological toxicity, and meets the technical requirements of GB / T15441-1995 Water Quality Determination of Acute Toxicity.

[0027] In addition to the purposes, features and advantages described above, the application has other purposes, features and advantages. The application will be further described in detail below with reference to the accompanying drawings. BRIEF DESCRIPTION OF DRAWINGS

[0028] The accompanying drawings are included to provide a further understanding of embodiments of the application, and are incorporated in and constitute a part of this specification, illustrate embodiments of the application, and together with the description serve to explain the principles of the application. In the drawings:

[0029] Figure 1 is a structural schematic diagram of a water quality online biological toxicity detection device provided by the embodiments of the application by using a lactic acid bacteria PH inhibition method;

[0030] Figure 2 is a flow chart of a water quality online biological toxicity detection method provided by the embodiments of the application by using a lactic acid bacteria PH inhibition method;

[0031] wherein 1 is a multi-way valve, 2 is a peristaltic pump, 3 is a first culture tube, 4 is a second culture tube, 5 is a metering unit, 6 is a positive sample bottle, 7 is a distilled water bottle, 8 is a water sample bottle to be detected, 9 is a waste liquid barrel, 10 is a lactic acid bacteria bottle, 11 is a culture solution bottle, 12 is a first PH sensor, 13 is a second PH sensor, and 14 is a constant temperature module. DETAILED DESCRIPTION

[0032] The application will be described in detail below with reference to the embodiments shown in the drawings, but it should be noted that these embodiments are not a limitation on the application, and equivalent transformations or substitutions of functions, methods, or structures made by those skilled in the art according to these embodiments are within the protection scope of the application.

[0033] Please refer to Figure 1The embodiment of the present application also provides a water quality online biological toxicity detection device based on lactic acid bacteria PH inhibition method, which comprises a multi-way valve 1, a peristaltic pump 2, a first culture tube 3, a second culture tube 4, a metering unit 5, a positive sample bottle 6, a distilled water bottle 7, a water sample bottle 8 to be detected, a waste liquid barrel 9, a lactic acid bacteria bottle 10, a culture solution bottle 11, a first PH sensor 12, a second PH sensor 13, a constant temperature module 14 and a control system, the inlet of the multi-way valve 1 is communicated with the first culture tube 3, the second culture tube 4, the metering unit 5, the positive sample bottle 6, the distilled water bottle 7, the water sample bottle 8 to be detected, the waste liquid barrel 9, the lactic acid bacteria bottle 10 and the culture solution bottle 11 respectively, the inlet of the multi-way valve 1 is communicated with the first culture tube 3 and the second culture tube 4 respectively, the first PH sensor 12 and the second PH sensor 13 are arranged in the first culture tube 3 and the second culture tube 4 respectively, and the constant temperature module 14 is used for constant temperature culture of the first culture tube 3 and the second culture tube 4, the peristaltic pump 2 is connected with the multi-way valve 1 through the metering unit 5, and the control system is connected with the multi-way valve 1, the peristaltic pump 2, the metering unit 5, the first PH sensor 12, the second PH sensor 13 and the constant temperature module 14 respectively. In the structure, the positive sample bottle 6, the distilled water bottle 7, the water sample bottle 8 to be detected, the waste liquid barrel 9, the lactic acid bacteria bottle 10 and the culture solution bottle 11 are used for containing positive test liquid, distilled water, water sample to be detected, waste liquid, lactic acid bacteria and lactic acid bacteria culture solution respectively, and the first PH sensor 12 and the second PH sensor 13 are used for testing the PH voltage value of the liquid in the first culture tube 3 and the second culture tube 4 respectively.

[0034] See Figure 2 The embodiment provides a water quality online biological toxicity detection method based on lactic acid bacteria PH inhibition method, which utilizes the detection device to detect, and comprises the following steps.

[0035] S1, preparing a first water sample and a second water sample; the specific steps are as follows:

[0036] S1.1, cleaning the culture tubes: the control system of the detection device controls the multi-way valve 1 and the peristaltic pump 2 to add a certain amount of distilled water into the first culture tube 3 and the second culture tube 4 respectively, so that the two culture tubes and the PH probes in the two culture tubes are cleaned, and the two culture tubes are kept clean.

[0037] S1.2, adding culture solution: the control system controls the multi-way valve 1 and the peristaltic pump 2 to add 2mL of lactic acid bacteria culture solution into the first culture tube 3 and the second culture tube 4.

[0038] S1.3, adding distilled water (zero point calibration) to the first culture tube: the control system controls the multi-way valve 1 and the peristaltic pump 2 to add 2mL of distilled water into the first culture tube 3.

[0039] S1.4 Add the water sample to be tested to the second culture tube (water sample test): The control system controls the multi-way valve 1 and the peristaltic pump 2 to add 2 mL of the water sample to be tested to the second culture tube 4.

[0040] S1.5 Adding bacterial culture: The control system controls the multi-way valve 1 and the peristaltic pump 2 to add 2 mL of lactic acid bacteria culture to the first culture tube 3 and the second culture tube 4 respectively. After mixing evenly, the first water sample and the second water sample are obtained.

[0041] S2. Obtain the initial pH voltage values ​​of the first water sample and the second water sample; the control system reads the pH voltage values ​​of the first pH sensor 12 and the second pH sensor 13, thereby obtaining the initial pH voltage value A1 of the water sample in the first culture tube 3 and the initial pH voltage value B1 of the water sample in the second culture tube 4.

[0042] S3. The first and second water samples are cultured at a constant temperature. The temperature of the first culture tube 3 and the second culture tube 4 is controlled by the constant temperature module 14 to be kept constant at 35℃. The constant temperature culture time is 20 minutes, so that the lactic acid bacteria produce lactic acid through metabolism.

[0043] S4. Obtain the pH voltage values ​​of the first and second water samples after cultivation. After the isothermal cultivation is completed, the control system reads and records the pH voltage value A2 of the first water sample and the pH voltage value B2 of the second water sample after cultivation.

[0044] S5. Calculate the biotoxicity of the water sample to be tested: Calculate the inhibition rate CF of the water sample to be tested using the following formula:

[0045]

[0046] Toxicity evaluation is conducted based on the calculation results; if the CF content is between 15% and 20%, it is considered mildly toxic; if the CF content is between 20% and 35%, it is considered moderately toxic; and if the CF content is greater than 35%, it is considered severely toxic.

[0047] S6. Cleaning the culture tubes: The control system adds distilled water to the first and second culture tubes by controlling the multi-way valve and the peristaltic pump to clean the culture tubes and the two pH probes in the culture tubes, so as to keep the two culture tubes clean.

[0048] Positive test:

[0049] The positive control test for online biotoxicity of water using the lactic acid bacteria pH inhibition method in this embodiment of the invention, i.e., the positive control test, involves a positive test solution prepared with 7.5 mg / L Zn. 2+ The solution is used to test the sensitivity of lactic acid bacteria to toxic substances. Under normal circumstances, the inhibition rate should be greater than 50%. The specific steps for a positive test are as follows:

[0050] Cleaning the culture tubes: The control system adds a certain amount of distilled water to the first and second culture tubes by controlling the multi-way valve and the peristaltic pump to clean the two culture tubes and the pH probes in the two culture tubes, so as to keep the two culture tubes clean.

[0051] Add culture medium: The control system adds 2 mL of lactic acid bacteria culture medium to the first culture tube and the second culture tube respectively.

[0052] Add distilled water to the first culture tube (zero point calibration): The control system adds 2 mL of distilled water to the first culture tube.

[0053] Add zinc sulfate standard solution (positive test) to the second culture tube: The control system adds 2 mL of zinc sulfate standard solution (7.5 mg / L) to the second culture tube.

[0054] Adding inoculum: The control system adds 2 mL of lactic acid bacteria inoculum to the first culture tube and the second culture tube respectively, and mixes them evenly.

[0055] Reading culture tube signals: The control system reads and records the pH voltage value A of the first culture tube. 11 pH voltage value B of the second culture tube 11 .

[0056] Constant temperature incubation: The temperature of the first and second culture tubes is kept constant at 35℃, and the incubation time is 20 minutes, so that the lactic acid bacteria can easily metabolize and produce lactic acid.

[0057] Reading culture tube signals: After 20 minutes, the control system reads and records the pH voltage value A of the first culture tube. 12 pH voltage value B of the second culture tube 12 .

[0058] Cleaning the culture tubes: The control system adds distilled water to the first and second culture tubes by controlling the multi-way valve and the peristaltic pump to clean the culture tubes and the two pH probes in the culture tubes, keeping the two culture tubes clean.

[0059] In the positive test, the inhibition rate (CF) of lactic acid bacteria in the water sample is calculated using the above-mentioned inhibition rate formula. At this time, the CF value should be greater than 50%.

[0060] Negative test:

[0061] The negative control test, or negative control test, of the online biotoxicity test for the lactic acid bacteria pH inhibition method in this invention tests the stability of the biotoxicity detection method using the lactic acid bacteria pH inhibition method. Under normal circumstances, the inhibition rate should be in the range of -10% to 10%. The specific steps of the negative test are as follows:

[0062] Cleaning the culture tubes: The control system adds a certain amount of distilled water to the first and second culture tubes by controlling the multi-way valve and the peristaltic pump, cleaning the two culture tubes and the pH probes in the two culture tubes, so as to keep the two culture tubes clean.

[0063] Add culture medium: The control system adds 2 mL of lactic acid bacteria culture medium to the first culture tube and the second culture tube respectively.

[0064] Add distilled water to the first culture tube (zero point calibration): The control system adds 2 mL of distilled water to the first culture tube.

[0065] Add distilled water to the second culture tube (zero point calibration): The control system adds 2 mL of distilled water to the first culture tube.

[0066] Adding inoculum: The control system adds 2 mL of lactic acid bacteria inoculum to the first culture tube and the second culture tube respectively, and mixes them evenly.

[0067] Reading culture tube signals: The control system reads and records the pH voltage value A of the first culture tube. 21 pH voltage value B of the second culture tube 21

[0068] Constant temperature incubation: The temperature of the first and second culture tubes is kept constant at 35℃, and the incubation time is 20 minutes, so that the lactic acid bacteria can easily metabolize and produce lactic acid.

[0069] Reading culture tube signals: After 20 minutes, the control system reads and records the pH voltage value A of the first culture tube. 22 pH voltage value B of the second culture tube 22 .

[0070] Cleaning the culture tubes: The control system adds a certain amount of distilled water to the first and second culture tubes by controlling the multi-way valve and the peristaltic pump to clean the two culture tubes and the pH probes in the two culture tubes, so as to keep the two culture tubes clean.

[0071] In the negative test, the inhibition rate of lactic acid bacteria in the water sample is calculated using the inhibition rate (CF) formula mentioned above. At this time, the CF value should be between -10% and 10%.

[0072] Test data

[0073] 1. Comparison of test data between the lactic acid bacteria pH inhibition method and the luminescent bacteria method in this invention embodiment.

[0074] Water samples were collected from a river at different times, and biotoxicity was detected using both the lactic acid bacteria method of this invention and the existing luminescent bacteria method. The test results are as follows:

[0075] Table 1 Comparison of test data from the lactic acid bacteria pH inhibition method and the luminescent bacteria method.

[0076]

[0077] As can be seen from the data in Table 1, the test data of the lactic acid bacteria pH inhibition method and the luminescent bacteria method in the embodiments of the present invention are highly consistent, further illustrating the feasibility of the lactic acid bacteria pH inhibition method for detecting biotoxicity.

[0078] 2. Positive test data:

[0079] Table 2 Positive test data

[0080]

[0081] As can be seen from the data in Table 2, the positive test CF value is greater than 50%, and the test data is stable, meeting the requirements for a positive test.

[0082] 3. Negative test data:

[0083] Table 3 Negative Test Data

[0084]

[0085] As can be seen from the data in Table 3, the negative test CF value is between -10% and 10%, which meets the requirements for a negative test.

[0086] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A method for online biotoxicity detection of water quality using lactic acid bacteria pH inhibition, characterized in that, Includes the following steps: S1. Prepare the first and second water samples; the specific steps are as follows: S1.1 Cleaning the culture tubes: The control system controls the multi-way valve (1) and the peristaltic pump (2) to add a certain amount of distilled water to the first culture tube (3) and the second culture tube (4) respectively, and cleans the pH probes in the first culture tube (3), the second culture tube (4) and the first culture tube (3) and the second culture tube (4); S1.2 Adding culture medium: The control system controls the addition of equal amounts of lactic acid bacteria culture medium to the first culture tube (3) and the second culture tube (4), respectively; S1.3 The control system controls the addition of distilled water equal to the amount of lactic acid bacteria culture medium added to the first culture tube (3); S1.4 The control system controls the addition of the same amount of test water sample as the added lactic acid bacteria culture medium to the second culture tube (4); S1.5 Adding bacterial strains: The control system controls the addition of an equal amount of lactic acid bacteria strains to the first culture tube (3) and the second culture tube (4), respectively, and after mixing evenly, the first water sample and the second water sample are obtained. S2. Obtain the initial pH voltage value of the first water sample respectively. and the initial pH voltage value of the second water sample ; S3. The first water sample and the second water sample are incubated at a constant temperature. S4. Obtain the pH voltage value of the first water sample after cultivation. pH voltage value of the second water sample ; S5. Perform biotoxicity analysis calculations on the water sample to be tested; specifically, calculate the inhibition rate of the water sample to be tested using the following formula. : Toxicity evaluation of the water sample to be tested is conducted based on the calculation results.

2. The detection method according to claim 1, characterized in that, In step S2, the control system reads the pH voltage values ​​of the first pH sensor (12) and the second pH sensor (13) to obtain the initial pH voltage value of the first water sample. and the initial pH voltage value of the second water sample .

3. The detection method according to claim 2, characterized in that, In step S3, the temperature of the first culture tube (3) and the second culture tube (4) is kept constant at 25~37℃, and the constant temperature culture time is 15~25min.

4. The detection method according to claim 3, characterized in that, In step S4, after the isothermal incubation is completed, the control system reads the pH voltage values ​​of the first pH sensor (12) and the second pH sensor (13) again to obtain the pH voltage value of the first water sample after incubation. pH voltage value of the second water sample .

5. A water quality online biotoxicity detection device using a lactic acid bacteria pH inhibition method, characterized in that, The detection device includes a multi-port valve (1), a peristaltic pump (2), a first culture tube (3), a second culture tube (4), a metering unit (5), a positive sample bottle (6), a distilled water bottle (7), a water sample bottle to be tested (8), a waste liquid tank (9), a lactic acid bacteria bottle (10), a culture medium bottle (11), a first pH sensor (12), a second pH sensor (13), a constant temperature module (14), and a control system. The multi-port valve (1) is connected to the first culture tube (3), the second culture tube (4), the metering unit (5), the positive sample bottle (6), the distilled water bottle (7), the water sample bottle to be tested (8), the waste liquid tank (9), the lactic acid bacteria bottle (10), and the... The culture medium bottle (11) is connected, the first pH sensor (12) and the second pH sensor (13) are respectively installed in the first culture tube (3) and the second culture tube (4), the constant temperature module (14) is used to perform constant temperature culture on the first culture tube (3) and the second culture tube (4), the peristaltic pump (2) is connected to the multi-way valve (1) through the metering unit (5), and the control system is connected to the multi-way valve (1), the peristaltic pump (2), the metering unit (5), the first pH sensor (12), the second pH sensor (13) and the constant temperature module (14).

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