CN1284135A - Rapid method for assessment of inhibition and kill of anaerobic bacteria by toxic compounds - Google Patents
Rapid method for assessment of inhibition and kill of anaerobic bacteria by toxic compounds Download PDFInfo
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- CN1284135A CN1284135A CN98813419A CN98813419A CN1284135A CN 1284135 A CN1284135 A CN 1284135A CN 98813419 A CN98813419 A CN 98813419A CN 98813419 A CN98813419 A CN 98813419A CN 1284135 A CN1284135 A CN 1284135A
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Abstract
A rapid method for determining the minimum inhibitory concentration of biocides or biocidal agents for use in anaerobically contaminated aqueous systems is disclosed. This rapid technique provides for an opportunity to determine the minimum amount of antimicrobial agents, or biocides either singularly or combination, to control anaerobic growth in aqueous streams contaminated by microorganisms. The test involves the use of pH indicator dye systems which respond to the environmental changes caused by the production of byproducts of carbohydrate metabolism by anaerobes.
Description
Invention field
The invention discloses a kind of fast method that is used for determining the agricultural chemicals that uses for the aqueous system that is polluted by anerobe or kills the minimum inhibition concentration of biological agent.This rapid technology provides and has been defined as controlling by anaerobic growth thing in the current of microbial contamination, and required biocide independent or combination is killed the chance of the minimum of biological agent in other words.This test comprises the use of pH indicator dye system, pH indicator dye system to by the production of by-products of the carbohydrate metabolism of anaerob and the environment change that causes react.
With need be opposite up to the conventionally test program of 1-2 time-of-week, this technology is providing test answers in about 30 minutes to about 8-10 hour period.This test procedure uses the multiple row sample aperture on microtitration culture dish, and every row have a plurality of sample aperture, and shifts contaminated water system, substratum, the predetermined aliquots containig of pH indicator, and increase, concentration is by the technology of the biocide of serial dilution.
Background of invention
In many fields that service water is used, must use and kill biological agent and control bacterial growth, avoiding, to prevent or to control the microorganism accumulation of bacterial deposition thing, and the accumulation that comes from the by product of bacterial deposition thing.The existence of these bacteriums and other microbial organisms can be disturbed the manufacturing of water treatment or the various products under these various process water occasions of use usually.Process water can comprise various water, the water of circulation primary in cooling water system only for example, the water of recirculation in the recirculating cooling water system of the sealing that has or do not have discharging unit for discharging, be used and be collected so that the water of handling before in Industrial processes, using, before or after handling with the quality standard that satisfies regulation, water as sewage discharge, at various products, the water of productive use that uses in the production of paper or textiles for example, and other water of productive use, for example in the Oil extraction, perhaps those water of productive use that use in the processing of hydro carbons chemical preparations.
In every kind of application of process water, use, and using at present and killing biological agent and reduce, prevent and controlling microbial, the existence of unwanted bacteria and similar grown organism.These growers not only cause stopping up, but also can deposit on the metallic surface of these microbe colonies, cause the corrosion of metallic surface.
In the various application of killing biological agent, relate to a kind of method, this method comprises with such concentration and adding in the processed process water, so that control the growth of these microbial organisms killing biological agent.The growth of so-called controlling microbial, we refer to and not only comprise eliminating fully and the elimination of microorganism of biological growth, but also comprise the static state control of microbial population, though so that not all organism all be killed, but control growing population to greatest extent.In the method for test and the definite efficient of killing biological agent that will use and type, in order to determine specific effective application of killing biological agent, especially determine to use in the water surrounding for the particular industry that produces microorganism therein, realize the static state control or the elimination fully of processed specified microorganisms, usually need to continue about 48 hours of 24-, sometimes continued for 1 week, the longest 2 week or the longer test procedures that reach.
These long test periods not only lose time, and the waste resource, and the microbiological manipulation of determining to keep specific contaminated industry source required appropriately kill biological agent, and best working strength and concentration aspect, the expense of these test procedures is higher.
In various industry and public place of entertainment application, microbicide is added in the aqueous system.During these are used some comprise that the adding microbicide is with the control industrial cooling circulating water system, in the water system of strolling about or have a rest such as swimming pool and hot spring, algae, bacterium, fungi and protozoic growth, add microbicide, with the bacterium in the control paper production process, use microbicide to control bacterial growth in the raw sugar course of processing or the like.The present invention also finds to kill biological agent and is used at industry and town water water inlet system, and control comprises (but being not limited to) striped mussel (zebramussel), the application of the invertebrates of Mytilus edulis and Asia clam (Asiaticclam) in the industrial cooling circulating water system.Though be specially adapted to aqueous system, but the present invention also can be applicable to the nonaqueous phase system.Term used herein " microbicide " and " killing biological agent " are used alternatingly, and comprise the pharmaceutical chemicals that is used for " nuisance " that define in as federal insecticide mycocide and rodenticide decree (FIFRA) in the control of water and nonaqueous phase fluid system.
The method of the microbicide quantity that is used at present determining that fluid system exists is the time-consuming measuring method (a kind of indirect method) of bacterial growth quantity in the system often, or the wet-chemical analysis method of active microbicide sample (a kind of direct method).Indirect method comprises cultivates the sample of taking from fluid system, to determine bacterial growth.If there is bacterial growth, usually more microbicide is added in this fluid system, till microbial culture demonstrates quantity microorganism growth stable or that reduce gradually.The wet-chemical analysis method is time-consuming, and workload is big, and when carrying out wet-chemical analysis in other scene except being furnished with breadboard well (well), error is very big.Before making the present invention, do not exist by indirect method, fast and accurately determine need how much be killed the on-the-spot easily method of biological agent by the system that anaerob pollutes.
Utilize tracer material to monitor that the method for the effect of the processing chemical preparations such as inhibitor and rust-preventive agent in the industrial water system is well-known.The US.4783314 of Hoots discloses by utilizing fluorometry, uses inertia transition metal tracer material to monitor the concentration of inhibitor and rust-preventive agent.The US.4966711 of Hoots etc. and US.5041386 disclose the concentration that the inert fluorescent additive that uses add-on to be proportional to the quantity of inhibitor and/or rust-preventive agent monitors inhibitor in the given industrial water system and/or rust-preventive agent.US.4992380,5006311 and 5132096 disclose the method and apparatus of the fluorescent tracer that uses in the application of monitoring Treatment of Industrial Water.
US5128419 and 5171450 discloses water-soluble polymers, and this water-soluble polymers has the fluorescence part that is used for monitoring the concentration that it is used in Treatment of Industrial Water.Japanese Patent No.55003668 (1980) discloses a kind of method of killing biological agent concentration that is used to monitor, this method is determined the concentration of microbicide indirectly by adding and measuring the lithium salts material.This method needs to add separately tracer material, and in order to obtain the result, needs to use atomic absorption spectrometry.Compare with fluorometry, atomic absorption spectrometry is quite expensive, and because related complex apparatus, and the cause of naked light and inflammable gas supply, atomic absorption spectrometry is difficult for being applicable to on-the-spot the use.
US.4,242,602 disclose a kind of UV spectrum of multiple water treatment component that monitors learns a skill.This method relates to expensive Analytical equipment and the use with special computer hardware that writes software.In addition, must on the basis of particular location, calibrate this equipment, and if the condition changing in the water, also must calibrate again.European patent application 466303 discloses and a kind of material has been added in the processed water, and observe this material how with the method for microbicide reaction.Measure the material that reacts with microbicide continuously, and determine the concentration of microbicide by the loss of this material.This method is cumbersome, needs special equipment, and twice independently chemical preparations supply.
Kill biological agent and microbicide and be used to control or eliminate bacterial growth in the multiple different service water medium.Usually, a kind of biological agent or microbicide killed is not enough to control all bacterial growths in the processed water medium.The processing of processed process water is disturbed in the existence of bacterium or other microbial organisms, and can cause corrosion and other associated problem of the equipment that contacts with these polluted water.
US patent No.5206151 (this patent is contained in this as a reference) discloses a kind of by using the rapid screening technology, determine to kill in the various process waters method of the effectiveness of biological agent and microbicide fast, this rapid screening technology obtains a plurality of samples of the contaminated water medium that contains microbial organisms; To wherein adding indicator dye, the oxidation-reduction indicator dyestuff of the dehydrogenase reaction that can produce with microbial organisms preferably; And in the processed solution that contains this oxidation-reduction indicator dyestuff, add subsequently and cultivate accelerator.Processed sample is installed on the titration culture dish that biocide quantity dilutes gradually, is being present under the identical temperature of the working temperature of industrial water system wherein, to this titration culture dish insulation with microbial organisms.This method is facilitated growth, and quickens the microbial organisms activity, causes increasing with the concentration of the reductase enzyme of oxidation-reduction indicator dyestuff reaction, thereby causes change in color.Write down the variation of color aspect subsequently.
By with respect to the tower that is untreated (column), relatively first of the color aspect change the minimum inhibitor concentration (M.I.C) of the biocide of the growth of the microbial organisms that can determine to suppress contained in the contaminated water system in the dyestuff.
In supporting the essential reducing environment of anaerobic bacterium, the oxidation-reduction indicator dyestuff such as the resazurin dyestuff can change color with reducing environment.So because environment may cause false positive findings, color change can not indicate whether to exist anaerob.
US patent No.5,206,151 described a kind of can be used for the test not highly polluted by anaerob, so do not have the water of original low (bearing) ORP.ORP is to be the Eo+ that unit is measured with mV.Disclosed method can be used for assessing facultative population in capable and the 67th row-Di 7 hurdles the 4th, the 6th hurdle row at the 6th hurdle 52-57.This facultative population mainly carries out aerobic respiration in test water, but under anaerobic can utilize anaerobic respiration/fermentation.Therefore, when under moderate or severe anaerobic condition, monitoring anaerobism kind group time, utilize redox dye to monitor that effectively the test of the facultative bacteria under the slight anaerobic condition can not provide result accurately.Because in will influence redox dye to the severe anaerobic condition and provide false data.On the contrary, when anaerob (obligate or facultative) is main population, and ORP is too low, so that can not utilize resazurin to detect the variation of metabolic activity the time, will use method disclosed herein.
US patent No.5,374,536 disclose a kind of product selects testing method, be used for determining fast sewage kill biological agent the synergy adulterant existence or kill the existence of biological agent adulterant.This method is used reduction-oxidation fuel system, the substratum of supply, incubation time and temperature that one or more are killed mixtures of biological agent or its adulterant and provide for the multiple colour-change of fuel system.
US patent No.5 discloses the colour-change that the microorganism that detects felt is stopped up in 413,680, and wherein iodonitrotetrazolium is used as indicator.
At Clinical Microbiology, in January, 1988, a kind of readout device that is used for quick MIC test is disclosed in the 1-7 page or leaf.But, because the valuable equipment that this method need be not easy to transport, so this method is not easy to rig-site utilization.In addition, described technology is growth end user liquid making body substratum, rather than natural fluid.This liquid nutrient medium has high protein content, can not be used for utilizing paper industry two kinds of the most normal use kill biological agent: glutaraldehyde and isothiazoline are tested, because they will be because of the protein loss of activity.In addition, this test is only worked to pure organism culture.Owing in paper mill or tested cooling tower fluid, have the blended population usually, so this technology purposes of being unsuitable for planning here.
At US patent No.5, in 627,042, cold light is used to count the number of live microorganism in the test soln.
PCT application WO 92/19764 has described the sensitive absorbefacient dyestuff based on pH, and this dyestuff is used to detect the microorganism growth in the body fluid of collection.The document described how to detect microorganism growth existence whether.Method disclosed herein is determined the effectiveness of microbicide to anaerob.
The present invention is by providing a kind of wieldy, accurate and successive concentrations of microbiocibles is determined method indirectly, solved the many problems that describe in detail above, determining of concentrations of microbiocibles is that microorganism growth in controlling flow system, the especially industrial water system is necessary.
So an object of the present invention is to provide a kind of process water that can be used for handling, be particularly useful for handling the different multiple microbicide of the process water that uses in the paper-making process and/or the rapid screening method of toxic agent, with controlling microbial.
Another object of the present invention provides a kind of screening and test can be used for the toxic agent and the quick visual screening procedure of killing biological agent of controlling microbial growth, and controls or eliminate in the process water, particularly the microorganism growth in the paper-making process.
Another object of the present invention is in visual program, use some pH indicator (being commonly referred to the pH indicator dye), the pH indicator dye changes color in visible light, thus the minimum inhibition concentration of specific antimicrobial (being MIC) in the water system that can determine to be polluted by microbial organisms.
The purpose of this invention is to provide at the scene a kind ofly, rather than rapid screening can be used for handling the different multiple microbicide of process water and/or the method for toxic agent in laboratory environment.
Another object of the present invention provides a kind of easy-to-handle microtitration culture dish, this microtitration culture dish contains a plurality of sample tube (perhaps sample aperture), each sample aperture can keep the contaminated water system that determines volume, the aliquots containig of perhaps contaminated water system.In the aliquots containig of water sample, add the dose known amounts pH indicator dye of predesignating subsequently, this pH indicator dye can react with the environmental change that is caused by the carbohydrate metabolism thing, and the carbohydrate metabolism thing produces because of the increase of the metabolic rate of microflora.Subsequently, the colour-change that the carbohydrate metabolism thing that produces because of the metabolic rate that increases and the chemical reaction between the pH indicator dye produce this indicator dye, this is a kind of measuring method of the metabolic increase of microorganism that causes by adding substratum in the water aliquots containig contained in the sample aperture of microtitration culture dish.Subsequently with regard to colour-change test (visual observation) these a plurality of aliquots containigs.Purpose of the present invention also comprises a series of controlled dilution that is used for by the biocide that produces various concentration, determine in specific contaminated water system, for example a certain papermaking batching, perhaps in the water that uses in the paper-making process, specific biocide or kill biological agent, perhaps these biocides or kill the program of minimum inhibition concentration of the arbitrary composition of biological agent.
The invention main idea
The invention discloses a kind of fast method that is used for determining the agricultural chemicals that uses for the aqueous system that is polluted by anerobe or kills the minimum inhibition concentration of biological agent.This rapid technology provides and has been defined as controlling by anaerobic growth thing in the current of microbial contamination, and required biocide independent or combination is killed the chance of the minimum of biological agent in other words.This test comprises the use of pH indicator dye system, pH indicator dye system to by the production of by-products of the carbohydrate metabolism of anaerob and the environment change that causes react.With need be opposite up to the conventionally test program of 1-2 time-of-week, this technology is providing test answers in about 30 minutes to about 8-10 hour period.This test procedure uses the multiple row sample aperture on microtitration culture dish, and every row have a plurality of sample aperture, and shifts contaminated water system, substratum, the predetermined aliquots containig of pH indicator, and increase, concentration is by the technology of the biocide of serial dilution.Detailed description of the invention
Present method is a kind ofly to determine which kind of kills biological agent/biocide and be suitable for use in most in the paper machine fluid, the means of controlling microbial growth.Uncontrolled microorganism growth can cause ugly deposition, and this deposition can cause paper break or aperture, and paper break or aperture cause shutdown of a high price.When testing,, and in being present in the real fluid of dynamic system wherein, this population tests importantly according to the natural population in the population environment about best biocide.These two kinds of factors are killed influence the effect of biological agent/biocide.Because biocide is very expensive, and only can be applicable to the zone selected, to select optimum activity to kill biological agent extremely important for system.Test should be carried out at the scene, paper mill.This potential range is several thousand miles in the full service laboratory of test paper machine fluid design.In shipment, along with the development of time, the mixed population of microorganism can change, and perhaps chemical transformation may take place fluid self.These factors can influence test result, and it is bad to cause killing the biological agent matching.
Kill biological agent
Term " is killed biological agent " and clearly is defined as any biocide that can produce killing effect in biosystem in the art.Term " is killed biological agent " and also is used to represent biocide (killing the subclass of biological agent) in the art, and this biocide is an inhibition, and is to prevent biological growth or the positive metabolic medicament of biosystem.Biocide is defined as containing a kind ofly to be killed biological agent or contains the mixture that one or more kill biological agent.Usually, biocide is microbicidel (microcidal) or quiet bacterium (microstatic), but microbicidel (microcidal) also can be represented biocidal, mean that to biosystem be " killing " (lethal), perhaps microbial inoculum is killed in expression, means microflora (subgroup of biosystem) " killing ".These terms can be used alternatingly in industrial circle, and those skilled in the art may appreciate that these terms are interchangeable.
Enumerated the illustrative biological agent of killing in the table 1, but the present invention has more than and is limited to those that enumerate and kills biological agent.Present method should be applicable to kills biological agent arbitrarily.
Table 1 is killed biological agent
| Kill biological agent | Effective constituent | Typically used |
| ????A | 3,5 dimethyl-tetrahydrochysene-2H-1,3,5-thiadiazine-2-thioketones | The sanitas of the starch and the human body |
| ????B | The methylene-bis thiocyanate- | Kill the slime mould agent |
| ????C | 1-alkyl (C 16-C 18) amino-3-aminopropane acetate ﹠ two (trichloromethyl) sulfone | Kill the slime mould agent |
| ????D | 5-chloro-2-methyl-4-isothiazoline-3-Tong ﹠2-methyl-4-isothiazoline-3-ketone | Kill slime mould agent sanitas |
| ????E | Alkyl dimethyl phenmethyl ammonium chloride ﹠ dialkyl methyl phenmethyl ammonium chloride | Kill the slime mould agent |
| ????F | 2,2-two bromo-3-nitrilo-propionic acid amides (DBNPA) | Kill the slime mould agent |
| ????G | 2-(thiocyano methyl sulfenyl)-benzothiazole ﹠ two (trichloromethyl) sulfone | Kill the slime mould agent |
| ????H | Sodium dimethyldithiocarbamate 40min ﹠ vinyl is two-the dithiocarbamic acid disodium | Kill the slime mould agent |
| ????I | Glutaraldehyde (1,5 pentanediol) | Kill the slime mould agent |
| ????J | 1-(3-chlorallyl)-3,5,7-three nitrogen chloro diamantane | Sanitas |
| ????K | N-4-dihydroxyl-alpha-oxo-Benzene Chloride | Kill the slime mould agent |
| ????L | Clorox | |
| ????M | 4,5-two chloro-1,2-disulfide group-3-ketone | |
| ????N | The decyl thio-ethylamine |
Substratum
The substratum of name is food only, and food is defined as having any material of nutrition in the art.Nutritious being further defined as provides nutrition.Nutrition is further defined as by this nutrition, the process that organism obtains and assimilates food.
The term Nutrient broth obtains from the DIFCO handbook, and is the technical term that AmericanPublic Health Association stipulates in " Standard Methods for theExamination of Water and Waste Water ".Nutrient broth, a kind of simple case of substratum can obtain from the suppliers such as DIFCO or BBL, and usually identical with above-mentioned " standard method " required prescription.Prescription and explanation are by public health and the strict control of medicine industry, thereby the term Nutrient broth is not a generic terms, should be understood that to represent the mixture of dehydrated powder, and when using usually, this mixture contains 3 parts of beef extracts and 5 parts of peptones.Nutrient broth is a kind of substratum, and this substratum is to promote the activity of microbial organisms and the food of growth.Nutrient broth is a kind of substratum of specific type.
Glucose and disinfectant mixing butter (dairy half and half pasteurizedcreamer) also are substratum.Glucose is also referred to as dextrose, is a kind of simple crystallised sugar.According to the Code of Federal Regulations the 21st chapter, the 131.180th joint, dairy half and half are milk and the food mixture that contains the cream of 10.5-18% butterfat.Super pasteurize is to be defined as at minimum 280 °F by Pasteurized Milk Ordinance Code to handle the milk term in two seconds down.The adding of the substratum of regulation allows also to be used to promote growth, and this growth produces the carbohydrate metabolism thing with the dyestuff reaction, unless by the adding of biocide, stop this growth, this growth is become stablize constant.
The effect of the substratum that adds is to increase microbial metabolism, so that microorganism, perhaps the by product by microorganisms can react with the pH dyestuff.This reaction is indicated by the colour-change in the fluid.The purposes of the nutrition promotor in the method described herein is to increase microorganism metabolism.
Nearly all substratum or food can be called as Biomedia or agar, if agar-agar is added into, to produce separation or to count the material of various microorganisms.The material that finds in the promotor is used in the agar growth medium (except cream) usually.Nearly all food source can be with being substratum in growth medium.Some of them comprise V-8 juice and Semen Maydis powder, are used in environmental microorganism agar usually.
The concentration of the substratum that uses in the nutrition promotor is different from the concentration of those substratum that use in the growth medium.Equally, the purposes of substratum is also different.In promotor, substratum attempts to increase microorganism metabolism, can not change commodity fluidic state significantly simultaneously.The activity that the protein that uses with the concentration that contained in the agar can disturb some to kill biological agent is considered in the design of accelerator solution, and makes these medicament loss of activity.Accelerator solution is designed to make this problem to be reduced to minimum degree, still promotes microorganism metabolism fast simultaneously.
One of benefit of the present invention is not need sterilization, so sterilization is not described.Needn't carry out sterilization, therefore before or after adding substratum, not need to carry out sterilization.In addition, follow the sterilisation program of experimental arrangement both unnecessary usually, can not realize these sterilisation program again at the scene.
Here in the method for Lve Shuing, do not need the sterile state/sterilization of substratum, solution, material etc.Because the population in the process water (1-8 hour) at short notice is determined, therefore in the method, the natural population in the process water will provide the microorganism active of actual measurement just.Compare with the biological agent selection technology of killing of standard, required time weak point is one of great advantage of this method.
Usually, the population in the test sample book of process water will contain 500,000-10,000,000 colony forming unit (cfu) bacteria/milliliters.If population is lower than 500,000cfu/mL, then in 8 hours time ranges of this method, the content of test hole will not reacted, and the more important thing is, and this test can not propose appropriate demand to killing biological agent.Utilize almost any commercial biocide, can kill the bacterium of comparatively small amt at an easy rate.When having millions of organism in every milliliter of fluid,, will have this demand when attempting control kind of group time.Satisfied this demand here in the disclosed method.
Though needn't under aseptic condition, test necessary use cleaning, unpolluted material.Can not use in the test and show the arbitrary substance that pollutes sign.
Can be added into the substratum in the branch sample treated that utilizes that above-mentioned pH indicator dye handles and comprise any substratum in the aqueous solution or the aqeous suspension, perhaps any minimum medium mixture comprises glucose, sucrose, fructose, beef extract, peptone, (composition of beef extract and peptone is commonly called " Nutrient broth ") tryptone, milk, mixing butter, yeast extract, the perhaps any mixture of above-mentioned substance.Determine that at the fast method that uses us preferably defined medium is that volume ratio is 1: 8: 10 a glucose, the mixture of Nutrient broth and mixing butter under the situation of minimum inhibition concentration (MIC) of specific antimicrobial.But, the mutual fusion of proper proportion arbitrarily of these substratum, and can successfully promote the microbial organisms activity, the active promotion of microbial organisms makes the formation of metabolic speed and carbohydrate metabolism thing accelerate again, and the carbohydrate metabolism thing can react with pH indicator dye of the present invention again.Add substratum by direct solution on the microtitration culture dish, microorganism active or microbial respiratory are increased, the metabolic processes of metabolic speed or microorganism increases, thereby top result is provided.So transmit system by the organism electronic cell, the interaction of these metabolites and pH indicator is given security for monitoring cell viability.
The microtitration culture dish
The microtitration culture dish that uses is preferably has the transparent plastics plate that multiple row is called the depression of sample aperture, has a plurality of sample aperture in every row.The microtitration culture dish preferably has at least two row sample aperture, has at least three sample aperture in every row.Preferably the microtitration culture dish has about 6-12 row, and every row have about 6-12 pattern hole.But these culture dish also can be, for example, ceramic whiteware ware or be convenient to is recognized or any other ware shape structure of the appropriate background color of visual observation colour-change.These culture dish can be from Mass, Cambridge, and Co-star Corporation obtains.
The equal-volume aliquots containig of taking from tested water system is added in a series of sample aperture in the top of the multiple row sample aperture that traverses across the microtitration culture dish.And in these sample aperture, add pH indicator dye, substratum and the biocide of predetermined concentration.The adding concentration of biocide is about 0.1ppm～5000ppm.Subsequently by using a plurality of transfer pipets, the biocide that concentrations of biocide is about the concentration serial dilution of 0.1ppm～5000ppm adds in each sample aperture of each row sample aperture, thereby sample aperture contains by the test water of biological pollution, and in each sample aperture of each row sample aperture, contain the biocide of concentration serial dilution.Before or after adding biocide, in each sample aperture, add the pH indicator dye of known quantity, and to the developing medium that wherein adds known quantity.In every row, obtain the sample aperture of a series of aliquots containigs of a series of serial dilutions that contain contaminated water system like this, wherein added the biocide that quantity reduces gradually.Under the situation that has substratum and pH indicator dye, the biocide that this number of series reduces is gradually measured microorganism or metabolic activity subsequently, the active increase that microorganism or metabolic activity reflection organism electronic cell transmit system.A kind of difference of killing microbial organisms contained in the five equilibrium sample of the medium after substratum is handled of every row test is killed biological agent.
Subsequently under the temperature that equals contaminated water system substantially, heat insulating culture contains independently several row sample aperture, every row sample aperture has the titration culture dish of a plurality of sample aperture, contain the biocide that concentration reduces in proper order in the aliquots containig of microorganism in the sample aperture, and the substratum of equivalent and pH indicator dye, the sample that is polluted by microbial organisms is taken from this contaminated water system at first.Essentially identical temperature means and comprises about 10-15 ℃ of actual environment temperature plus-minus.These culture temperature are at least 20 ℃, but are no more than about 90 ℃ usually.The heat insulating culture time should be enough to form the variation of indicator dye color aspect by the reaction of indicator dye with the environmental change that is caused by the promoted active carbohydrate metabolism production of by-products that produces of substratum in the tested media.The sample culture dish must carry out anaerobic heat-preservation and cultivate.Can use for on-the-spot from the Anaero Pack System that Mitsubishi Gas Chemical Company buys by Remel.
The heat insulating culture time can be short to about 30 minutes, and can grow 8-10 hour.Test is compared with former antimicrobial acivity, this test duration, preferably be about 4-8 hour, and quite short with regard to the time, and also more sensitive for the existence of the minimum inhibition concentration of biocide.After time, take out the titration culture dish in the appropriate heat insulating culture between between about 30 minutes～8-10 hour, and visual inspection, with the colour-change of determining to take place in each sample aperture in every row sample aperture of titration culture dish.When untreated sample aperture becomes suitable color (dyestuff that depends on use), read the reading of titration culture dish.In every row sample aperture, to high concentrations of biocide, first observed is confirmed as the minimum inhibition concentration of biocide contained in this row sample aperture to the concentration of colour-change from low concentrations of biocide.Certainly, every row sample aperture contains the independent mixture of independent biocide (killing biological agent in other words) or biocide.
Can use the biocide of different starting point concentrations to repeat this process, till repeated experiments is determined control and kept the minimum inhibition concentration of controlling the microbial organisms in the aqueous media that utilizes this biocide processing.
The contaminated water system that the method for the minimum inhibition concentration of our definite biocide is specially adapted to test takes from the raw material or the batching water of different places in slurrying and the paper mill.Substratum preferably is selected from Nutrient broth, glucose, and the mixing butter of super pasteurize and their mixture, this mixture also can comprise above-described other substratum.The microtitration culture dish contains at least three row, best four row sample aperture, and every row preferably have at least eight sample aperture.The microtitration culture dish is usually by glass, and pottery is made, but preferably by transparent plastics or arbitrarily other medium make, described other medium arbitrarily can stand culture temperature and incubation time, and suitable background color is provided, by this background color, can observe and account for color.Titration culture dish after the processing carries out anaerobic heat-preservation usually under about 30～50 ℃ temperature cultivates, and the time is about 30 minutes～4-6 hour.
In addition, can be such as by nitrogen, helium is finished heat insulating culture in the inert gas atmosphere that argon gas and carbonic acid gas or the like form.When under inert gas atmosphere, carrying out heat insulating culture, will obtain other result, this result will give the experimenter about under the anaerobic condition, and contaminated water system organism is to the information of the reaction of killing biological agent.
The present invention is a kind of method that is used for the water system biocide minimum inhibition concentration of definite anaerob pollution, comprises the steps:
A) obtain described contaminated water systematic sample;
B) in described sample, add the pH indicator dye of the environmental change reaction that causes with carbohydrate metabolism thing by described anaerob;
C) substratum is added in the described sample, form the nutritive water phase system that dyestuff is handled;
D) obtain the aliquots containig of the nutritive water phase system that dyestuff handles;
E) biocide that will test of serial dilution repeatedly, and form the mixture of the antimicrobial agent solution of the described aliquots containig of the nutritive water phase system that dyestuff handles and each described serial dilution;
F) under the temperature that equals contaminated water system substantially, anaerobic heat-preservation is cultivated described mixture, and the heat insulating culture time should be enough to by the reaction of dyestuff with the described environmental change that is caused by the carbohydrate metabolism thing, the variation that produces dye colour;
G), determine to suppress the minimum biocide inhibition concentration of anaerob contained in the described contaminated water system by observing change in color.
In addition, the water system that anerobe pollutes is optional from slurrying and paper water, paper mill batching water, paper mill plain boiled water, brown raw material water (stock water), papermaking sewage, open circuit recirculated cooling water, closed cycle water coolant, oiler feed, sugar refinery water of productive use, chemical process liquid stream, fermentation broth stream foodstuff production water and oil and refinery's water of productive use and sewage.
In addition, substratum can be selected from mixing butter, yeast extract, glucose, sucrose, fructose, glycerine, beef extract, peptone, tryptone, milk and their mixture.
Can carry out serial dilution, so that on the cumulative volume basis of dye well substratum post-treatment aqueous phase system, the ultimate density of the described biocide of serial dilution is about 0.1ppm～10,000ppm.
The pH indicator dye can be selected from purpurum bromocresolis, tetrabromo-mcresolsulfonphthalein, coeruleum bromocresolis, dichlorophenol sulfonphthalein, methylene blue chloride, methyl red and phenol red.
The nutritive water phase system that can handle the described dyestuff of serial dilution on the microtitration culture dish carries out heat insulating culture, this microtitration culture dish contains at least four row sample aperture, every row have at least eight sample aperture, the temperature of heat insulating culture is about 25～60 ℃, and the time is about 60 minutes～and 24 hours.
In addition, in the heat insulating culture process, above processed titration culture dish, keep mainly containing inert gas atmosphere.
Rare gas element can be selected from CO
2And N
2
For enforcement of the present invention, step b) is delayed 30 minutes～and 20 hours.
For the more information about the method for biocide minimum inhibition concentration in our the definite contaminated water system is provided, provide the following examples.
Embodiment
From feed proportioning (stock furnish), tissue paper batching or other batching obtain about 100 milliliters filtration papermaking batching.
Subsequently 5 milliliters of culture medium solutions, 1 milliliter of pH indicator dye solution adds by strainer, for example by tissue paper or have in filtering these filtering solutions of strainer of about US Std Sieve Series No.90 screen size.Culture medium solution contains in 100 ml waters from 0.4 dried powder of BBL or Difco Nutrient broth and 1 gram glucose.The mixing butter that in this culture medium solution, adds 0.5 milliliter of ultrapasteurised that can obtain from local food supply retail shop.PH indicator dye solution contains 0.15 gram dyestuff in 100 milliliters of 0.01%NaOH.Use the hyperchannel micropipet subsequently, for example the Titertek that provides by the Flow laboratory
Micropipet, and with killing in the as many sample aperture of the biological agent row of will testing, the filtered water ingredients solution after this processing of about 150 microlitres is sucked in each microtitration sample aperture.Titertek is the breadboard registered trademark of Flow.
1 milliliter 1% the biological agent solution of killing is added 4 milliliters containing in substratum-dyestuff-batching sample, kill biological agent solution with formation 2000ppm in sample.These commerce or experiment are killed biological agent and can be included, but not limited to polyamine, isothiazoline, organosulfur compound (organosulfur), quaternary amine, organbromine compound, carbaminate, the methylene-bis thiocyanate-, perhaps their composition, it is perhaps known that other kills biological agent arbitrarily.
After forming this 2000ppm solution, take out this treatment soln of 150 microlitres immediately, and put it in the first micro-sample aperture of first tests column of microtitration culture dish.Make a row sample aperture keep not being killed the state that biological agent is handled.In the solution suction transfer pipet that forms in the microtitration sample aperture, this transfer pipet is used for shifting kills biological agent 2-3 time subsequently, subsequently the solution of sucking-off is put back in the same sample aperture, thus mixing solutions.After the mixing, from first sample aperture, extract 150 microlitre mixtures, and it is added in next sample aperture in the same row.Carry out this combination process once more, and utilize micropipet to draw this second sample aperture solution of 150 microlitres.This second aliquots containig is put into the 3rd sample aperture of same column on the microtitration culture dish.Repeat this program, all sample aperture in single-row all contain adding wherein and with it till the biocide of blended concentration serial dilution (killing biological agent).The 150 mul aliquots samples that extract from last sample aperture of this row are discharged from.
Kill biological agent for all that will test, in every row, repeat this program.Make a row sample aperture keep the state that is untreated,, promptly contain test soln with as contrast, substratum, the pH dyestuff, but do not add any tests column of killing biological agent or biocide.
To containing the multiple row sample aperture, every row have the microtitration culture dish heat insulating culture of a plurality of sample aperture subsequently.Temperature is preferably 10-15 ℃ of the temperature plus-minus of tested processing stream.These temperature are 20 ℃～80 ℃, and are typically about 25-30 ℃～approximately 50-60 ℃.Because carrying out anaerobic heat-preservation in carbon dioxide environment, cultivates by the microtitration culture dish, promptly before heat insulating culture, culture dish is provided by the anaerobic environment that is for example provided by Anaero Pack System, and Anaero Pack System can obtain from Mitsubishi Gas Chemical Company.Commercially available this system is converted into carbonic acid gas and H to the airborne oxygen of sample top
2O, thus CO is provided
2Atmosphere.
After said procedure, concentration gradient, promptly the serial dilution concentration of biocide will change to the about 8ppm (perhaps lower) in the minimum concentration sample aperture from the 1000ppm first high density sample aperture.Exist under the situation of dyestuff, the sample aperture that does not contain the capacity biocide is transformed into another color.The sample aperture that contains the capacity biocide keeps initial color.When the color in all check sample holes (untreated) is terminal colour-yellow or when transparent, read test result.By the concentration in comparative sequence diluted sample and the untreated sample aperture, can in its concentrations of biocide the sample aperture place of the minimum concentration of microbial organisms growth in the control test sample book, the visual MIC that determines each biocide.
Found this test is reacted, and in the paper mill, bacterium common in the cooling tower etc. comprises clostridium, sulfate-reducing bacteria and Klebsiella pneumoniae.By selecting biocide rightly or kill the starting point concentration of biological agent, can be about 10,000ppm-up to and comprise the concentration range that is lower than 1ppm concentration, biological agent is killed in screening.
Embodiment given below is used to illustrate the preferred embodiments of the invention and application, unless explanation in addition in additional claim does not limit the invention.
Embodiment 1
In order to experimentize, utilize NaOH or HCl that the pH value of the plain boiled water after the filtration of slurrying and paper mill acquisition is adjusted to 6.5.The 100mL aliquots containig after the adjusting of pH value, and 5mL hydration MiniTox
TM(can be from Illinois, the Nalco Chemical company of Naperville buys, and contains 0.4 gram Nutrient broth, 1.0 gram glucose, 100mLH
2O) nutrient solution, 0.15% dichlorophenol sulfonphthalein of 1mL, and the 0.5mL mixing butter adds Whirlpak together
In the bag.This solution of thorough mixing is till color even.To call the AnnaTox mixing processing to this solution below.
150 microlitre AnnaTox mixing processing are added in each prospect hole of microtitration pallet (microtitretray).Subsequently, 4 milliliters of AnnaTox mixing processing are added in 10 ml beakers (beaker is used for a kind of screened biological agent of killing).Next, 1 milliliter 1% (weight percent) killed biological agent solution add in each beaker, and mix.Extract 150 microlitres immediately, and beginning is along row prospect hole serial dilution from the top down.At the terminal point of this row prospect hole, this 150 microlitre is excluded.To interested all kill biological agent and repeat this program.One row's prospect hole is handled with killing biological agent, to be used as contrast.
Seal this pallet with scotch tape subsequently, avoiding evaporating, and, under the no stirring situation, carry out anaerobism and cultivate, till the flavescence of contrast prospect hole at 37 ℃.Periodically check changing to yellow of pallet from red.Dichlorophenol sulfonphthalein is pure red when pH value 6.8, is true yellow when pH value 5.1.By the fermentation byproduct (bi-product) that is present in the anaerobic bacterium population in the sample, realize the reduction of pH value.If there is no kill biological agent, can in 1-24 hour, colour-change take place, depend on the type of anaerobism population in the sample.Can effectively suppress or kill these bacteriums if kill biological agent, then can not observe colour-change.Behind all contrast prospect holes flavescence, read minimum inhibition concentration.With under this concentration, the form of the concentration of prospect hole bottom flavescence is read MIC.This is illustrated in this specified point, does not have to add the capacity that can tell on and kills biological agent.Like this, can carry out quantitative assay with regard to the effectiveness of killing the biological agent processing.
In order to ensure colour-change is because actual pH changes causes, but not because other external influence factors causes, uses AnnaTox method experiment arrangement; And after reading MIC, measure pH.The result shows that on average the MIC reading is corresponding to the remarkable decline of pH aspect between the concentration.The result is referring to the table IX.
Cause by microorganism growth in order to determine that pH changes, rather than by the CO that sucks
2Perhaps other influence factor causes, before chemically examining, under 121 ℃ to this batching autoclaving 30 minutes.Replace using scotch tape sealing prospect hole to avoid evaporating, in each prospect hole, can use a few dropstone waxes to seal prospect hole.Simultaneously not autoclaved batching is tested.Paraffin also is used to seal these prospect holes.In not autoclaved sample, with regard to killing the biological agent effect, MIC demonstrates normal change.In 21 hours, the chemical examination of carrying out for the sterilization batching does not demonstrate colour-change.At this moment, stop this test.
This test procedure also can be used for quantitatively determining to add killing after the biological agent, and it is to live that how many microorganisms are arranged.Obtain aliquots containig from each sample aperture, and utilize the plate culture of standard to determine bacterial count.
Be important to note that under anaerobic and test.The microorganism growth of indication comprises the biological and amphimicrobian biology of obligate anaerobic.The obligate anaerobic biology is those microorganisms that can not utilize oxygen, and the amphimicrobian biology is those microorganisms that can grow under aerobic or anaerobic situation.
Can use following any pH indicator to carry out this program: the table II
Trade(brand)name chemical name supplier
Purpurum bromocresolis 5,5-two bromo-o-cresolsulfonphthalein Baker
Tetrabromo-mcresolsulfonphthalein 4,4-(2,2-diepoxy-3H-1,2 benzo xathiol-3-EM ylidene) two [2,6-two bromo-3-methyl-sodium salts] Science
Dibromothymolsulfonphthalein 3,3-two dibromothymolsulfonphthalein Baker
Chlorophenol 4,4-(1,1-diepoxy-3H-2,1-benzoxathiol-3-Acros ylidene) two [2-chloro-9Cl]
Methylene blue chloride thiodiphenylamine-5,3, two (dimethylamino)-muriate (9Cl) EM Science of 7-
Methyl red (neighbour-[[right-(dimethylamino) phenyl] azo] phenylformic acid Baker
Phenol red phenolsulfonphthalein, sodium salt Baker
The benefit of using different indicator is the pH value that needn't regulate tested sample significantly, but can consider the intrinsic pH value of the particular system that will test, selects sample pH value.
Utilize above-described experimental arrangement to be the MIC in the determination of raw material table III that is obtained from the southeast (Southeastern) slurrying and paper mill.
The table III
Kill biological agent MIC (ppm)
A
1????????????????1000
B
2??????????????????8
C
3??????????????????8
D
4??????????????????8
E
5??????????????????32
F
6??????????????????8
G
7??????????????????16
1=tetrahydrochysene-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thioketones can be buied from Buckman.
2=Sodium dimethyldithiocarbamate 40min and vinyl be two-the dithiocarbamic acid disodium, can buy from Alco Chemical.
3=1,3 pentandial (glutaraldehyde) can buy from Union Carbide.
4=1,3 pentandial and N-alkyl dimethyl phenmethyl ammonium chloride can be buied from UnionCarbide.
5=1-alkyl (C
8-C
18) 3-amino-3-aminopropane one acetate and two (trichloromethyl) sulfone, can buy from Akzo.
6=5-chloro-2-methyl-4-isothiazoline-3-ketone and methyl-4-isothiazoline-3-ketone can be buied from Rohm﹠Haas.
7=N-alkyl dimethyl phenmethyl ammonium chloride and N-dialkyl methyl benzene ammonium chloride can be buied from Mason.
Embodiment 2
Use the program of describing among the embodiment 1, test obtains the result of table IV from the raw material in (Southeastern) slurrying of the different southeast and paper mill.The table IV
Kill biological agent MIC (ppm)
A
1???????????????1000
B
2?????????????????8
C
3?????????????????32
D
4?????????????????64
E
5?????????????????125
F
6?????????????????32
G
7?????????????????125
1=tetrahydrochysene-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thioketones can be buied from Buckman.
2=Sodium dimethyldithiocarbamate 40min and vinyl be two-the dithiocarbamic acid disodium, can buy from Alco Chemical.
3=1,3 pentandial (glutaraldehyde) can buy from Union Carbide.
4=1,3 pentandial and N-alkyl dimethyl phenmethyl ammonium chloride can be buied from UnionCarbide.
5=1-alkyl (C
8-C
18) 3-amino-3-aminopropane one acetate and two (trichloromethyl) sulfone, can buy from Akzo.
6=5-chloro-2-methyl-4-isothiazoline-3-ketone and methyl-4-isothiazoline-3-ketone can be buied from Rohm﹠Haas.
7=N-alkyl dimethyl phenmethyl ammonium chloride and N-dialkyl methyl benzene ammonium chloride can be buied from Mason.
Embodiment 3
Use the experimental arrangement of describing among the embodiment 1, test obtains the result of table V from the different material in (Southeastern) slurrying of another southeast and paper mill.
The table V
Kill biological agent MIC (ppm)
A
1??????????????1000
B
2????????????????8
C
3????????????????8
D
4????????????????8
H
8????????????????16
E
5????????????????8
F
6????????????????8
G
7????????????????8
1=tetrahydrochysene-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thioketones can be buied from Buckman.
2=Sodium dimethyldithiocarbamate 40min and vinyl be two-the dithiocarbamic acid disodium, can buy from Alco Chemical.
3=1,3 pentandial (glutaraldehyde) can buy from Union Carbide.
4=1,3 pentandial and N-alkyl dimethyl phenmethyl ammonium chloride can be buied from UnionCarbide.
5=1-alkyl (C
8-C
18) 3-amino-3-aminopropane one acetate and two (trichloromethyl) sulfone, can buy from Akzo.
6=5-chloro-2-methyl-4-isothiazoline-3-ketone and methyl-4-isothiazoline-3-ketone can be buied from Rohm﹠Haas.
7=N-alkyl dimethyl phenmethyl ammonium chloride and N-dialkyl methyl benzene ammonium chloride can be buied from Mason.
8=2-bromo-2-nitropropane-1,3-glycol (bronopol) can be buied from Angus.
Embodiment 4
Use (Southeastern) slurrying of another southeast of program test and the paper mill raw material described among the embodiment 1, obtain the result of table VI.
The table VI
Kill biological agent MIC (ppm)
A
1??????????????1000
B
2???????????????16
C
3???????????????8
D
4???????????????8
H
8???????????????32
F
5???????????????32
G
6???????????????32
1=tetrahydrochysene-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thioketones can be buied from Buckman.
2=Sodium dimethyldithiocarbamate 40min and vinyl be two-the dithiocarbamic acid disodium, can buy from Alco Chemical.
3=1,3 pentandial (glutaraldehyde) can buy from Union Carbide.
4=1,3 pentandial and N-alkyl dimethyl phenmethyl ammonium chloride can be buied from UnionCarbide.
5=5-chloro-2-methyl-4-isothiazoline-3-ketone and methyl-4-isothiazoline-3-ketone can be buied from Akzo.
6=N-alkyl dimethyl phenmethyl ammonium chloride and N-dialkyl methyl benzene ammonium chloride can be buied from Rohm﹠Haas.
8=2-bromo-2-nitropropane-1,3-glycol (bronopol) can be buied from Angus.
Embodiment 5
Use (Southeastern) slurrying of another southeast of program test and the paper mill raw material described among the embodiment 1, obtain the result of table VII.
The table VII
Kill biological agent MIC (ppm)
B
2?????????????????64
C
3?????????????????32
H
8?????????????????32
E
5?????????????????64
G
7?????????????????64
2=Sodium dimethyldithiocarbamate 40min and vinyl be two-the dithiocarbamic acid disodium, can buy from Alco Chemical.
3=1,3 pentandial (glutaraldehyde) can buy from Union Carbide.
5=1-alkyl (C
8-C
18) 3-amino-3-aminopropane one acetate and two (trichloromethyl) sulfone, can buy from Akzo.
7=N-alkyl dimethyl phenmethyl ammonium chloride and N-dialkyl methyl benzene ammonium chloride can be buied from Mason.
8=2-bromo-2-nitropropane-1,3-glycol (bronopol) can be buied from Angus.
Embodiment 6
Use (Southeastern) slurrying of another southeast of program test and the paper mill raw material described among the embodiment 1, obtain the result of table VIII.
The table VIII
Kill biological agent MIC (ppm)
A
1???????????????1000
B
2????????????????16
I
9????????????????250
J
10???????????????250
G
7????????????????64
1=tetrahydrochysene-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thioketones can be buied from Buckman.
2=Sodium dimethyldithiocarbamate 40min and vinyl be two-the dithiocarbamic acid disodium, can buy from Alco Chemical.
7=N-alkyl dimethyl phenmethyl ammonium chloride and N-dialkyl methyl benzene ammonium chloride can be buied from Mason.
9=methylene-bis (thiocyanate-) can be buied from Rohm and Haas.
10=1-(the rare propyl group of 3-chlorine)-3,5,7-three azepines-1-nitrogen diamantane can be from Michigan, and the Dow Chemical company of Midland buys.
Embodiment 7
The table IX understands that for example pH changes along with adding in this system killing biological agent with different concentration.In all cases, represent the pH breakpoint with boldface letter.Do not add the situation (H) of killing biological agent for having, can not produce tangible pH in the system and change.In the method for the invention, this just pH of indicator dye tracking changes.
The table IX
| PH variation along with the adding of killing biological agent | ||||||||
| Concentration (ppm) | ??A 1 | ??B 2 | ??C 3 | ??D 4 | ??E 5 | ??F 6 | ??G 7 | ??H 8 |
| ??1000 | ?4.66 | ?6.95 | ?6.34 | ?6.59 | ?7.08 | ?6.87 | ?6.87 | ?5.19 |
| ???500 | ?4.71 | ?6.89 | ?6.70 | ?6.70 | ?6.82 | ?6.94 | ?6.92 | ?4.76 |
| ???250 | ?5.20 | ?7.01 | ?6.76 | ?6.80 | ?6.82 | ?6.99 | ?6.85 | ?4.72 |
| ???125 | ?4.67 | ?7.21 | ?6.80 | ?6.80 | ?6.93 | ?6.95 | ?6.85 | ?4.92 |
| ????64 | ?4.86 | ?6.97 | ?6.70 | ?6.70 | ?7.06 | ?5.61 | ?6.12 | ?-- |
| ????32 | ?4.79 | ?5.95 | ?6.61 | ?6.60 | ?6.50 | ?4.87 | ?5.79 | ?-- |
| ????16 | ?4.67 | ?5.91 | ?5.76 | ?5.33 | ?4.56 | ?4.88 | ?4.62 | ?-- |
| ????8 | ?4.87 | ?4.48 | ?5.64 | ?5.52 | ?4.47 | ?4.82 | ?4.83 | |
| Initial pH | 7.21 | ?6.95 | ?6.95 | ?6.89 | ?7.20 | ?7.16 | ?6.81 | ?6.97 |
1=tetrahydrochysene-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thioketones can be buied from Buckman.
2=Sodium dimethyldithiocarbamate 40min and vinyl be two-the dithiocarbamic acid disodium, can buy from Alco Chemical.
3=1,3 pentandial (glutaraldehyde) can buy from Union Carbide.
4=N-alkyl dimethyl phenmethyl ammonium chloride and N-dialkyl methyl phenmethyl ammonium chloride can be buied from Mason.
5=2-bromo-2-nitropropane-1,3-glycol (bronopol) can be buied from Angus.
6=5-chloro-2-methyl-4-isothiazoline-3-ketone and methyl-4-isothiazoline-3-ketone can be buied from Rohm﹠Haas.
The mixture of 7=N-alkyl dimethyl phenmethyl ammonium chloride and N-dialkyl methyl benzene ammonium chloride (can buy) and 1,3 pentandial (glutaraldehyde) (can buy) from Union Carbide from Mason.
The 8=contrast does not add and kills biological agent.
Under the situation that does not break away from the principle and scope of the present invention that limit in the claim, can change composition, operation and the arrangement of the inventive method described herein.
Claims (9)
1. the method for the minimum inhibition concentration of biocide in the water system of a definite anaerob pollution comprises the steps:
A) obtain described contaminated water systematic sample;
B) in described sample, add the pH indicator dye that the environmental change that causes with carbohydrate metabolism thing because of described anaerob reacts;
C) substratum is added in the described sample, form the nutritive water phase system that dyestuff is handled;
D) obtain the aliquots containig of the nutritive water phase system that dyestuff handles;
E) biocide that will test of serial dilution repeatedly, and form the mixture of the antimicrobial agent solution of the described aliquots containig of the nutritive water phase system that dyestuff handles and each described serial dilution;
F) under the temperature that equals contaminated water system substantially, anaerobic heat-preservation is cultivated described mixture, and the heat insulating culture time should be enough to by the reaction of dyestuff with the described environmental change that is caused by the carbohydrate metabolism thing, the variation that produces dye colour;
G), determine to suppress the minimum biocide inhibition concentration of anaerob contained in the described contaminated water system by observing change in color.
2. in accordance with the method for claim 1, the water system that wherein said anerobe pollutes is selected from slurrying and paper water, paper mill batching water, paper mill plain boiled water, brown raw material water (stock water), papermaking sewage, open circuit recirculated cooling water, closed cycle water coolant, oiler feed, sugar refinery water of productive use, chemical process liquid stream, fermentation broth stream foodstuff production water and oil and refinery's water of productive use and sewage.
3. in accordance with the method for claim 1, wherein said substratum is selected from mixing butter, yeast extract, glucose, sucrose, fructose, glycerine, beef extract, peptone, tryptone, milk and their mixture.
4. in accordance with the method for claim 1, wherein carry out described serial dilution, so that on the cumulative volume basis of dye well substratum post-treatment aqueous phase system, the ultimate density of the described biocide of serial dilution is about 0.1ppm～10,000ppm.
5. in accordance with the method for claim 1, wherein said pH indicator dye is selected from purpurum bromocresolis, tetrabromo-mcresolsulfonphthalein, coeruleum bromocresolis, dichlorophenol sulfonphthalein, methylene blue chloride, methyl red and phenol red.
6. in accordance with the method for claim 1, wherein the nutritive water phase system of on the microtitration culture dish the described dyestuff of serial dilution being handled carries out heat insulating culture, this microtitration culture dish contains at least four row sample aperture, every row have at least eight sample aperture, the temperature of heat insulating culture is about 25～60 ℃, and the time is about 60 minutes～and 24 hours.
7. in accordance with the method for claim 6, wherein in the heat insulating culture process, above processed titration culture dish, keep mainly containing inert gas atmosphere.
8. in accordance with the method for claim 7, wherein rare gas element is selected from CO
2And N
2
9. in accordance with the method for claim 1, wherein step b) be delayed 30 minutes～20 hours.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US99514497A | 1997-12-29 | 1997-12-29 | |
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| JP (1) | JP2002500020A (en) |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101802610A (en) * | 2007-09-20 | 2010-08-11 | 陶氏环球技术公司 | A high throughput test method for evaluation of biocides against anaerobic microorganisms |
| CN105087361A (en) * | 2015-09-08 | 2015-11-25 | 湖州一控医疗科技有限公司 | Suspension type biological indicator and preparation method thereof |
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|---|---|---|---|---|
| BRPI0206375B1 (en) * | 2001-01-09 | 2015-08-04 | Kurita Water Ind Ltd | Method for selection of antimicrobial agent and its use |
| DK1639122T3 (en) | 2003-07-02 | 2009-05-04 | Dsm Ip Assets Bv | Improved test system for determining the presence of an antibiotic in a fluid |
| BRPI0917707A2 (en) * | 2008-08-27 | 2017-05-30 | Stepan Co | potentiated biocidal compositions and methods of use |
| US11071301B2 (en) | 2016-10-21 | 2021-07-27 | Ecolab Usa Inc. | Anti-microbial agent to control biomass accumulation in SO2 scrubbers |
| EP4288518A1 (en) * | 2021-02-08 | 2023-12-13 | C-Square Bioscience GmbH | Device and method for regulating the content of microorganisms |
Family Cites Families (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5094955A (en) * | 1988-03-15 | 1992-03-10 | Akzo N.V. | Device and method for detecting microorganisms |
| US5252484A (en) * | 1988-11-29 | 1993-10-12 | Minnesota Mining And Manufacturing Company | Rapid read-out biological indicator |
| US5206151A (en) * | 1990-06-11 | 1993-04-27 | Nalco Chemical Company | Rapid selection of biocide using a reduction oxidation indicator system |
| CA2058252A1 (en) * | 1991-03-18 | 1992-09-19 | Linda R. Robertson | Synergistic product selection test for biocides |
| US5885791A (en) * | 1995-11-16 | 1999-03-23 | The Research And Development Institute, Inc. | Antifungal and antibacterial susceptibility assay |
-
1998
- 1998-12-04 WO PCT/US1998/025864 patent/WO1999034013A1/en not_active Application Discontinuation
- 1998-12-04 JP JP2000526667A patent/JP2002500020A/en active Pending
- 1998-12-04 KR KR1020007007249A patent/KR20010033726A/en not_active Application Discontinuation
- 1998-12-04 NZ NZ505399A patent/NZ505399A/en unknown
- 1998-12-04 ID IDW20001264A patent/ID26631A/en unknown
- 1998-12-04 CA CA002316806A patent/CA2316806A1/en not_active Abandoned
- 1998-12-04 CN CN98813419A patent/CN1284135A/en active Pending
- 1998-12-04 AU AU16292/99A patent/AU1629299A/en not_active Abandoned
- 1998-12-04 EP EP98960774A patent/EP1051510A4/en not_active Withdrawn
- 1998-12-07 IN IN2139CA1998 patent/IN185760B/en unknown
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101802610A (en) * | 2007-09-20 | 2010-08-11 | 陶氏环球技术公司 | A high throughput test method for evaluation of biocides against anaerobic microorganisms |
| CN105087361A (en) * | 2015-09-08 | 2015-11-25 | 湖州一控医疗科技有限公司 | Suspension type biological indicator and preparation method thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| ID26631A (en) | 2001-01-25 |
| WO1999034013A1 (en) | 1999-07-08 |
| JP2002500020A (en) | 2002-01-08 |
| EP1051510A1 (en) | 2000-11-15 |
| NZ505399A (en) | 2002-10-25 |
| CA2316806A1 (en) | 1999-07-08 |
| IN185760B (en) | 2001-04-21 |
| EP1051510A4 (en) | 2003-01-22 |
| KR20010033726A (en) | 2001-04-25 |
| AU1629299A (en) | 1999-07-19 |
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