CN101698862A - Detection method for acute biological toxicity of drinking water contained in plastic bottle - Google Patents
Detection method for acute biological toxicity of drinking water contained in plastic bottle Download PDFInfo
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- CN101698862A CN101698862A CN200910196924A CN200910196924A CN101698862A CN 101698862 A CN101698862 A CN 101698862A CN 200910196924 A CN200910196924 A CN 200910196924A CN 200910196924 A CN200910196924 A CN 200910196924A CN 101698862 A CN101698862 A CN 101698862A
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Abstract
The invention relates to a detection method for the acute biological toxicity of drinking water contained in a plastic bottle, belonging to the technical field of drinking water detection. The method comprises the following steps: preparing a bacterial liquid used in tests from a fresh bacterial liquid or a bacterial liquid of freeze-dried powder, preparing a drinking water sample liquid, a control sample liquid, and a detection sample liquid for a plastic bottle, preparing a set of sample parallel samples by using the drinking water sample liquid or the detection sample liquid for the plastic bottle, then preparing a set of control parallel samples, adding 0.1 ml of fresh bacterial liquid or 0.05 ml of bacterial liquid of freeze-dried powder to each set of parallel samples, respectively placing for 15 min, determining the illumination intensity of one sample in each small test cup by a luminescence detecting device, and then calculating the relative illumination intensity. Compared with the prior art, the invention uses a freshwater luminescent bacterium peculiar to China, Vibrio-qinghaiensissp, as a detection strain, and the luminescent bacteria can grow and give out light without sodium chlorides. The invention has the advantages that the physical and chemical characteristics of the samples can not be changed, the correctness of the tests is ensured, and the detection speed is high.
Description
[technical field]
The present invention relates to tap water detection technique field, relate in particular to a kind of detection method of acute biological toxicity of drinking water contained in plastic bottle.
[background technology]
People need drink water every day, and safety of drinking water can not be ignored.The sales volume of bottled drinking water is very big at present, kind is also various, existing pure water, distilled water, natural mineral water or artificial mineralized water are also arranged, and their majorities all use Plastic Bottle as container, for these commodity tap water, the standard that country has " bottle (bucket) dress sanitary standard for drinking water " GB 19298-2003 to determine, wherein stipulated the detection index that some are main, comprised sense organ, physics and chemistry and microbiological indicator, this type of detects the assessment that index does not relate to bio-toxicity basically.With regard to existing situation as can be known, the detection that lacks bio-toxicity is a great disappearance of safety control, and for example the people is for poisoning, the illegally appearance of foodstuff additive, or the existence of heavy metal ion, health that all can the serious threat drinking person.
With regard to existing detection means supervision is helpless, because the detection method of physics or chemistry often relatively lags behind, only detect at known chemical substance, and the appearance of at present a large amount of syntheticses, make the conventional sense method hard to guard against, if the detection method with present national standard detects, often want several talentes to have the result, can not before listing, just judge; In addition, except learning the concentration that detects thing, the judgement that lacks bio-toxicity, for example: can find the content of the heavy metal of mercury and so on, but can't judge when this kind content status, this tap water is to eater's safety whether, especially a certain method that adopts in the GB can only be at certain material, material for not regulation detection then can not detect, so often have omission, causes the generation of acute poisoning incident.
Be the incident that prevents that posing a health risk from appearring in tap water, be necessary to carry out a kind of brand-new safety detection method.
At present, photogenic bacterium toxicity detection method has been widely used in the pollution of water quality and bio-toxicity is judged, and is simple and easy to do with it, detect advantage such as with low cost and receive publicity, can be referring to the mensuration of acute toxicity of water quality---photogenic bacterium method, GB/T15441-1995; But if detect, then need in sample, to add sodium-chlor and reach 3% final concentration, might change the physicochemical property of sample after so the sodium-chlor of high density adds, the exactness of influence test with the ocean luminescent bacteria.But Shang Weijian is useful on the detection fast and safely of drinking bottled water.
[summary of the invention]
The objective of the invention is to overcome the deficiencies in the prior art, employing Qinghai Vibrion Q67 substratum is made fresh bacterium liquid or is adopted the Qinghai Vibrion lyophilized powder to make test and use bacterium liquid, and a kind of method that detects the bottled drinking water security fast of designing.
For achieving the above object, the present invention proposes a kind of detection method of acute biological toxicity of drinking water contained in plastic bottle, it is characterized in that step is as follows: bacterium liquid is used in (1) preparation test: adopt fresh bacterium liquid or lyophilisate bacterium liquid, described fresh bacterium liquid is the slant strains with Qinghai Vibrion Q67 bacterial strain, move and receive on the fresh inclined-plane, cultivate 12h-18h for 15-25 ℃, this moment, bacterial luminescence was bright, with 10ml 0.85% physiological saline lawn is softly washed away from the inclined-plane, shake up, adjust bacterial concentration with a small amount of physiological saline, get the adjusted bacterium liquid of 0.1ml, add and to be equipped with in the sample test cell of 2ml0.85% physiological saline, detect with photometer luminous, select for use luminous intensity at the bacterium liquid of 200-500 ten thousand photons/second as fresh bacterium liquid; Described lyophilisate bacterium liquid is to add the 5mL resuscitation fluid in Qinghai Vibrion Q67 lyophilisate, thorough mixing, and the following placement of room temperature 8-15 minute makes bacterium recover luminous, promptly can be used as test bacterium liquid; (2) preparation sample: get bottled drinking water to be detected, add NaCl to final concentration be 0.8% as the tap water sample liquid; The inferior boiling point distilled water that adopts secondary heavily to steam, add NaCl to final concentration be 0.8% sample liquid in contrast; Clip Plastic Bottle 2g puts into triangular flask, adds the NaCl solution of 15ml 0.8%, and with 108 ℃ of sterilization 30min, gained solution is the Plastic Bottle steeping fluid, makes extract, and is as the test sample liquid of Plastic Bottle, standby; (3) to the luminous detection of sample: get six test cuvettes, wherein three test cuvettes add 2ml tap water sample liquid or Plastic Bottle respectively test sample liquid as one group of parallel sample of sample, in addition three test cuvettes are put into 2ml control sample liquid respectively and are also contrasted parallel sample as one group; Add fresh bacterium liquid of 0.1ml or 0.05ml lyophilisate bacterium liquid successively every 15s in three test cuvettes in every group of parallel sample, after placing 15min respectively, just adopt the luminous detection instrument to test the luminous intensity of sample in the cuvette every 15s between every group of parallel sample like this, calculate relative luminous intensity then.
It is OD that a small amount of physiological saline of described usefulness is adjusted bacterial concentration
660Be 0.2～0.5.
Described resuscitation fluid is that volumetric molar concentration is 0.05-0.2M, and pH value is the phosphoric acid buffer of 6.0-9.0, or quality-concentration of volume percent is the Glucose Liquid of 0.5-8.0%.
The relative luminous intensity of tap water is 95%～150% safety, and＞150% or 80～95% is suspicious, and＜80% is dangerous.
The relative luminous intensity of tap water Plastic Bottle is 95%～120% safety, and＞120% or 80～95% is suspicious, and＜80% is dangerous.
The present invention compares with prior art, utilize photogenic bacterium to external world poisonous substance the characteristics of sensitive reaction are arranged, adopting the exclusive a kind of fresh water photogenic bacterium of China---Qinghai Vibrion is used bacterial classification as detecting, adopt the exclusive a kind of fresh water photogenic bacterium Qinghai Vibrion Q67 bacterial strain of China that all kinds of poisonous substances are all had the sensitive reaction, and used Qinghai Vibrion is a fresh water type luminescent bacteria, need not sodium-chlor just can grow with luminous, therefore need not to add high density chlorination sodium in the specimen, can not change the physicochemical property of sample, guarantee the exactness of test, the safety detection that is used for bottled water, at first be that detection speed is fast: in 1 hour, can obtain a result, generally can whether assess its edible safety selling the previous crops detection; Secondly, be judgement: whether have the acute biological toxicity material to be present in the sample, can make safe, dangerous or suspicious judgement to sample to edible safety.
[embodiment]
Adopt the distinctive fresh water photogenic bacterium of China-Qinghai Vibrion Q67 bacterial strain Vibrio qinghaiensis Q67 among the present invention; be to being present in the reflection of the comprehensive toxicity of institute's Toxic in the sample; even the new synthetics that never occurred; as long as it is poisonous to organism; can make a difference to photogenic bacterium; and reflect from the luminous power of photogenic bacterium delicately; modern photon detection technique is very sensitive; atomic weak light intensity has taken place to change and also can detect; so extremely sensitive with the luminous detection of photogenic bacterium to the reaction of poisonous substance, than the general biomass cells several magnitude that is quick on the draw.
The luminous detection instrument: the BHP95114 type water quality toxicity analyser that adopts Binsong Photon Technology Co., Ltd. Beijing to produce is the test instrument.
The Qinghai Vibrion Q67 bacterial strain Vibrio qinghaiensis Q67 that adopts among the present invention is by red legend outstanding person, Wang Jie; the novel species of a kind of photogenic bacterium that Chen Xiaoyun etc. find; be published in document: Oceanologia et Limnologia Sinica, 25 (3): 273-278, this Qinghai Vibrion Q67 medium component is as follows:
MgSO
42.47g, MgCO
30.79g, MgBr
20.09g, MgCl
20.109g, CaCO
30.103g, KCl 0.122g, NaCl 8.29g, Mg (HCO
3)
20.50g, yeast extract paste 5g, tryptone 5g, glycerine 3g, agar 20g, be dissolved in the 1000mL distilled water, with its bevel, in 121 ℃ of sterilization 20min, it is standby to store in 4 ℃ of refrigerators.
The luminescent bacteria lyophilisate, the lyophilisate that employing is made into by above-mentioned bacterial classification, the patent No. of this lyophilisate: 97106203.X, can buy by market, be stored among-20 ℃ of refrigerators, use front opening, make it dissolving with an amount of resuscitation fluid, about 10 minutes, recover luminous vigor, just can be used for detecting, this resuscitation fluid is with reference to East China Normal University's journal (natural science edition, 2008,4,58-65 delivers, " isolation identification of Qinghai Lake photogenic bacterium and the electron microscopic observation " write articles by Mi Qin etc.
The luminous detection data computing adopts following formula:
The tap water evaluation of result:
| The effect toxicity grading | Relative luminous intensity |
| Safety | ?95％～150％ |
| Suspicious | ＞150% or 80～95% |
| Dangerous | ?＜80％ |
Tap water Plastic Bottle evaluation of result:
| The effect classification | Relative luminous intensity |
| Safety | 95％～120％ |
| Suspicious | ＞120% or 80～95% |
| Dangerous | ＜80％ |
Embodiment 1
One, test is prepared with bacterium liquid
1, the preparation of fresh bacterium liquid:
With the slant strains of above-mentioned Qinghai Vibrion Q67 bacterial strain, move and receive on the fresh inclined-plane, cultivate 12h-18h for 15-25 ℃; this moment, bacterial luminescence was bright, with 10ml 0.85%NaCl solution, lawn was softly washed away from the inclined-plane; shake up, adjust bacterial concentration, make bacterial concentration OD with a small amount of physiological saline
660About ≈ 0.3, get 0.1ml bacterium liquid, adding is equipped with in the sample test cell of 2ml physiological saline, and is luminous with the photometer detection, and luminous intensity is when 200-500 ten thousand photons/second, and this fresh bacterium liquid just can be used as the bacterium liquid of test usefulness.
2, the preparation of lyophilisate bacterium liquid:
In Qinghai Vibrion Q67 bacterial strain lyophilisate, add the 5ml resuscitation fluid, thorough mixing, the following placement of room temperature about 10 minutes makes bacterium recover luminous, obtains lyophilisate bacterium liquid, also can be used as test bacterium liquid.
Embodiment 1
The bottled pure water water quality toxicity detects
One, sampling
(1) bottled water sample liquid: an amount of with bottled pure water, add NaCl to final concentration be 0.8%.
(2) contrast water sample liquid: with the inferior boiling point distilled water that secondary heavily steams, add NaCl to final concentration be 0.8%.
Two, luminous detection
Be ready to three test cuvettes, each cuvette respectively adds 2ml bottled water sample liquid, is made as three parallel samples; Control sample liquid is also got three test cuvettes, each adds 2ml, three parallel samples are set equally, add fresh bacterium liquid 0.1ml or lyophilized powder bacterium liquid 0.05ml successively as test bacterium liquid, the timed interval that each test cuvette adds bacterium liquid is 15s, after placing 15min, measure the luminous intensity of each measuring cup, calculating relative luminous intensity with the luminous detection instrument 15s of being separated by successively.
Three, luminous detection data computing:
Four, test result evaluation:
Embodiment 2
Bottled distilled water water quality toxicity detects:
One, sampling
(1) directly get the distilled water sample: it is an amount of to get bottled distilled water, add NaCl to final concentration be 0.8%.
(2) blank water sample contrast, with the inferior boiling point distilled water that secondary heavily steams, add NaCl to final concentration be 0.8%.
Two, luminous detection
The BHP95114 type water quality toxicity analyser that adopts Binsong Photon Technology Co., Ltd. Beijing to produce is the test instrument.
Can use lyophilized powder bacterium liquid, also can use fresh bacterium liquid.
Be ready to three test cuvettes, each cuvette respectively adds 2ml distilled water sample, be made as three parallel samples, three test cuvettes are also got in blank water sample contrast, and each adds 2ml, three parallel samples are set equally, each test cuvette 15s at interval adds fresh bacterium liquid 0.1ml or lyophilisate bacterium liquid 0.05ml successively, as test bacterium liquid, after each test cuvette is placed 15min, measure the luminous intensity respectively test cuvette with the luminous detection instrument 15s of being separated by successively, calculate relative luminous intensity.
Three, luminous detection data computing
Four, test result evaluation
Embodiment 3
The detection of bottled natural mineral water
One, sampling
(1) directly get the preparation that mineral water is cooked sample liquid: an amount of with bottled natural mineral water, add NaCl to final concentration be 0.8%.
(2) preparation of blank sample liquid: the inferior boiling point distilled water that heavily steams with secondary, add NaCl to final concentration be 0.8%.
Two, luminous detection
Be ready to three test cuvettes, each cuvette respectively adds 2ml mineral water sample liquid, is made as three parallel samples; Control sample liquid is also got three test cuvettes, each adds 2ml, three parallel samples are set equally, add fresh bacterium liquid 0.1ml or lyophilized powder bacterium liquid 0.05ml successively as test bacterium liquid, the timed interval that each test cuvette adds bacterium liquid is 15s, after placing 15min, measure the luminous intensity of each measuring cup, calculating relative luminous intensity with the luminous detection instrument 15s of being separated by successively.
Three, luminous detection data computing:
Test result is estimated:
Embodiment 4
Bottled artificial mineral water's detection
One, sampling
(1) directly get the preparation of artificial mineral water's sample liquid: an amount of with bottled natural mineral water, add NaCl to final concentration be 0.8%.
(2) preparation of blank sample liquid: the inferior boiling point distilled water that heavily steams with secondary, add NaCl to final concentration be 0.8%.
Two, luminous detection
Be ready to three test cuvettes, each cuvette respectively adds 2ml artificial mineral water sample liquid, is made as three parallel samples; Control sample liquid is also got three test cuvettes, each adds 2ml, three parallel samples are set equally, add fresh bacterium liquid 0.1ml or lyophilized powder bacterium liquid 0.05ml successively as test bacterium liquid, the timed interval that each test cuvette adds bacterium liquid is 15s, after placing 15min, measure the luminous intensity of each measuring cup, calculating relative luminous intensity with the luminous detection instrument 15s of being separated by successively.
Three, luminous detection data computing
Four, test result evaluation:
Embodiment 5
The detection of Plastic Bottle
One, sampling
(1) blank sample liquid: the inferior boiling point distilled water that heavily steams with secondary, add NaCl to final concentration be 0.8%.
(2) preparation of Plastic Bottle sample steeping fluid: the Plastic Bottle sample is cut into the square of 1cm * 1cm, takes by weighing 2g and put in the triangular flask, add 15ml 0.8%NaCl solution, with 108 ℃ of sterilization 30min, gained solution is Plastic Bottle sample extract, and is as Plastic Bottle detected sample liquid, stand-by.
Two, luminous detection
Be ready to three test cuvettes, each cuvette respectively adds the detected sample liquid of 2ml Plastic Bottle, is made as three parallel samples; Control sample liquid is also got three test cuvettes, each adds 2ml, three parallel samples are set equally, add fresh bacterium liquid 0.1ml or lyophilized powder bacterium liquid 0.05ml successively as test bacterium liquid, the timed interval that each test cuvette adds bacterium liquid is 15s, after placing 15min, measure the luminous intensity of each measuring cup, calculating relative luminous intensity with the luminous detection instrument 15s of being separated by successively.
Three, luminous detection data computing
Four, test result evaluation:
| Sample liquid (adopting the test of lyophilisate bacterium liquid) | Luminous intensity (%) | The effect toxicity grading |
| ?F | ??100.5 | Safety |
| ?Z | ??89 | Suspicious |
| ?S | ??74 | Dangerous |
Claims (5)
1. the detection method of an acute biological toxicity of drinking water contained in plastic bottle, it is characterized in that step is as follows: bacterium liquid is used in (1) preparation test: adopt fresh bacterium liquid or lyophilisate bacterium liquid, described fresh bacterium liquid is the slant strains with Qinghai Vibrion Q67 bacterial strain, move and receive on the fresh inclined-plane, cultivate 12h-18h for 15-25 ℃, this moment, bacterial luminescence was bright, with 10ml 0.85% physiological saline lawn is softly washed away from the inclined-plane, shake up, adjust bacterial concentration with a small amount of physiological saline, get the adjusted bacterium liquid of 0.1ml, add and be equipped with in the sample test cell of 2ml 0.85% physiological saline, detect with photometer luminous, select for use luminous intensity at the bacterium liquid of 200-500 ten thousand photons/second as fresh bacterium liquid; Described lyophilisate bacterium liquid is to add the 5mL resuscitation fluid in Qinghai Vibrion Q67 lyophilisate, thorough mixing, and the following placement of room temperature 8-15 minute makes bacterium recover luminous, promptly can be used as test bacterium liquid; (2) preparation sample: get bottled drinking water to be detected, add NaCl to final concentration be 0.8% as the tap water sample liquid; The inferior boiling point distilled water that adopts secondary heavily to steam, add NaCl to final concentration be 0.8% sample liquid in contrast; Clip Plastic Bottle 2g puts into triangular flask, adds the NaCl solution of 15ml 0.8%, and with 108 ℃ of sterilization 30min, gained solution is the Plastic Bottle steeping fluid, makes extract, and is as the test sample liquid of Plastic Bottle, standby; (3) to the luminous detection of sample: get six test cuvettes, wherein three test cuvettes add 2ml tap water sample liquid or Plastic Bottle respectively test sample liquid as one group of parallel sample of sample, in addition three test cuvettes are put into 2ml control sample liquid respectively and are also contrasted parallel sample as one group; Add fresh bacterium liquid of 0.1ml or 0.05ml lyophilisate bacterium liquid successively every 15s in three test cuvettes in every group of parallel sample, after placing 15min respectively, just adopt the luminous detection instrument to test the luminous intensity of sample in the cuvette every 15s between every group of parallel sample like this, calculate relative luminous intensity then.
2. the detection method of a kind of acute biological toxicity of drinking water contained in plastic bottle as claimed in claim 1 is characterized in that: it is OD that a small amount of physiological saline of described usefulness is adjusted bacterial concentration
660Be 0.2～0.5.
3. the detection method of a kind of acute biological toxicity of drinking water contained in plastic bottle as claimed in claim 1, it is characterized in that: described resuscitation fluid is that volumetric molar concentration is 0.05-0.2M, pH value is the phosphoric acid buffer of 6.0-9.0, or quality-concentration of volume percent is the Glucose Liquid of 0.5-8.0%.
4. the detection method of a kind of acute biological toxicity of drinking water contained in plastic bottle as claimed in claim 1, it is characterized in that: the relative luminous intensity of tap water is 95%～150% safety, and＞150% or 80～95% is suspicious, and＜80% is dangerous.
5. the detection method of a kind of acute biological toxicity of drinking water contained in plastic bottle as claimed in claim 1, it is characterized in that: the relative luminous intensity of tap water Plastic Bottle is 95%～120% safety, and＞120% or 80～95% is suspicious, and＜80% is dangerous.
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| CN102507447A (en) * | 2011-10-25 | 2012-06-20 | 中国科学院华南植物园 | Method for measuring mycotoxin fusaric acid by using Vibrio-qinghaiensis Sp.Nov. (strain Q67) |
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| CN102175828A (en) * | 2011-01-20 | 2011-09-07 | 济南市供排水监测中心 | Method for evaluating water quality health risk with chromium ion as standard toxic substance |
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Application publication date: 20100428 |