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WO1999011816A1 - Reactifs pour controler la proprete et procede de controle de la proprete - Google Patents

Reactifs pour controler la proprete et procede de controle de la proprete Download PDF

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Publication number
WO1999011816A1
WO1999011816A1 PCT/JP1998/003761 JP9803761W WO9911816A1 WO 1999011816 A1 WO1999011816 A1 WO 1999011816A1 JP 9803761 W JP9803761 W JP 9803761W WO 9911816 A1 WO9911816 A1 WO 9911816A1
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Prior art keywords
reagent
cleanliness
atp
adp
test
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Ceased
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PCT/JP1998/003761
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English (en)
Japanese (ja)
Inventor
Tatsuya Sakakibara
Seiji Murakami
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Kikkoman Corp
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Kikkoman Corp
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Priority to AU87503/98A priority Critical patent/AU8750398A/en
Publication of WO1999011816A1 publication Critical patent/WO1999011816A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions

Definitions

  • the present invention relates to an improvement of a cleanliness inspection method (hereinafter, referred to as a cleanliness inspection method by AT II method) using ATP as a contamination mark using a luciferin-luciferase luminescent reagent. More specifically, in addition to ATP, an indicator component of dirt, ADP, AMP and RNA, which were difficult to measure with conventional luciferin 'luciferase luminescent reagents, were measured at the same time.
  • the present invention relates to a cleanliness test reagent capable of detecting and accurately evaluating cleanliness, and a cleanliness test method using the reagent.
  • This method is generally performed according to the following procedure.
  • ATP is degraded by ATPase immediately after the death of the organism to become ADP or AMP.
  • Foods and beverages, their semi-finished products, and their raw materials contain significant amounts of ADP and AMP, but they do not react with the luciferin-norrecyclase luminescent reagent.
  • the conventional cleanliness inspection method using the ATP method cannot detect AMP, ADP, and RNA other than ATP, and therefore cannot perform a highly accurate cleanliness inspection.
  • the present invention is based on the conventional Luciferin 'Lucif
  • An object of the present invention is to provide a reagent for testing cleanliness, and a method for testing cleanliness using this reagent, with the accuracy of simultaneously measuring ADP, AMP and RNA, which was difficult to measure with a perase luminescent reagent.
  • the present inventors include pyruvate orthophosphate dikinase (hereinafter, sometimes referred to as PPDK), phosphoenolpyruvate, pyrophosphate, luciferin, luciferase and a metal salt.
  • PPDK pyruvate orthophosphate dikinase
  • phosphoenolpyruvate phosphoenolpyruvate
  • pyrophosphate pyrophosphate
  • luciferin luciferin
  • luciferase luciferase
  • a metal salt luciferase
  • PPDK-containing luminescent reagent (Hereinafter sometimes referred to as “PPDK-containing luminescent reagent”), it is possible to measure AMP in addition to ATP, which is an indicator of contamination, and to detect small contamination with high sensitivity. As a result, it was found that high-precision cleanliness inspection can be performed.
  • the above-mentioned luminescent reagent containing PPDK is used in combination with “enzyme that catalyzes the reaction that generates 80 to 8” and Z or “RNA degrading enzyme”, in addition to ATP and AMP, AD P and RNA was also measured at the same time, and it was found that even less dirt could be detected and a more accurate cleanliness inspection could be performed. That is, it has been found that a more accurate cleanliness test can be performed by measuring at least one selected from the group consisting of ADP, AMP and RNA and ATP as an index.
  • the present invention has been completed based on these findings.
  • the present invention is a cleanliness test reagent characterized by comprising a pyruvate orthophosphodiquinidase, a phosphoenolpenolepinoleic acid, a pyrophosphate, a luciferin, a luciferase and a metal salt.
  • the present invention provides at least one selected from the group consisting of an enzyme that catalyzes a reaction for producing ATP from ADP, a reagent for extracting a microbial intracellular component, and an RNA degrading enzyme.
  • a cleanliness inspection reagent characterized by comprising one or more of pyruvate orthophosphate dikinase, phosphoenolpyruvate, pivalate, luciferin, lumpulase and a metal salt,
  • the present invention also provides a cleanliness inspection method characterized by measuring at least one kind selected from the group consisting of ADP, AMP and RNA and ATP as an index, (4) Further, the present invention provides a method for producing a dirt-collecting carrier by contacting the carrier with the dirt with the inspection site, and then contacting the carrier with the dirt with sterile water.
  • a method for testing cleanliness comprising reacting the reagent with a cleanliness test reagent containing lysine, phosphoenorubyric acid, pyrophosphate, luciferin, luciferase, and a metal salt, and measuring the amount of luminescence generated.
  • the present invention provides a method for producing a carrier to which dirt is collected by bringing a carrier for collecting dirt into contact with an inspection site, and then contacting a reagent for extracting a component in a microbial cell with the carrier.
  • a reagent for extracting a component in a microbial cell At least one selected from the group consisting of an enzyme that catalyzes the reaction of producing ATP from ribonuclease and RNA degrading enzyme; pyruvate orthophosphoditokinase, phosphoenolpyruvate, pyrophosphate, noresiferin, and lumferase.
  • a cleanliness test reagent containing a metal salt and measuring the amount of luminescence generated.
  • the present invention provides a method for testing cleanliness containing a pyruvate orthophosphodikinase, phosphoenorubyric acid, pyrophosphate, luciferin, luciferase, and a metal salt to a test object or a processed product thereof.
  • a cleanness inspection method characterized by measuring the amount of light generated.
  • the present invention relates to a method for producing a test object or a processed product thereof, comprising: A cleanliness test reagent containing pyruvate orthophosphate dikinase, phosphoenolpyruvate, pyrophosphate, noresiferin, noresiferase and metal salts, and measuring the amount of luminescence generated. Cleanliness inspection method.
  • pyruvate orthophosphate dikinase used in the present invention acts on AMP, phosphoenorubyric acid and pyrophosphate in the presence of magnesium ion to catalyze a reaction to produce ATP, pyruvate and phosphate.
  • AMP phosphoenorubyric acid
  • pyrophosphate in the presence of magnesium ion to catalyze a reaction to produce ATP, pyruvate and phosphate.
  • These enzymes are already known for their physicochemical properties and manufacturing methods and are readily available. Such enzymes are known from microorganisms and plants.
  • microorganism-derived microorganisms include microorganisms belonging to the following genera or species.
  • Microbispora for example, Microbispora
  • plant-derived plants include the following plants.
  • This medium was inoculated with Microbispora sammorosa (Microbispsp0rathrermorosea) IFO14047, and this was cultured at 45 ° C with shaking to obtain a culture.
  • buffer A 20 mM HEPES buffer (pH 7.5) containing 5 mM EDTA, 1 ml MgS04 and 1 mM DTT (hereinafter referred to as buffer A) was added, and the cells were added. The suspension was sufficiently suspended to obtain 700 ml.
  • the above cell suspension (700 ml) was purified by the following procedure.
  • lysozyme manufactured by Nagase Seikagaku Co., Ltd.
  • diammonium phosphate 2 3.1 g was added, and the mixture was further stirred at room temperature for 2 hours to disrupt the cells.
  • This crushed liquid was centrifuged at 700 rpm for 15 minutes, and the supernatant was collected to obtain 620 ml of the liquid.
  • Ammonium sulfate was dissolved at a rate of 2 g / 100 ml in the above-mentioned solution (620 ml), and this was previously dissolved in the buffer A (pH 7.5) containing 0.15 M ammonium sulfate.
  • the enzyme was adsorbed on about 700 ml of QAE-Sephadex resin equilibrated in the above). This was washed with the buffer A (pH 7.5) containing 0.15 M ammonium sulfate to remove unnecessary proteins, and then the buffer A (pH 7.5) containing 0.6 M ammonium sulfate was removed. Eluted.
  • the eluate was concentrated using a hollow fiber ultrafiltration device (PAN 13-DX, manufactured by Asahi Medical Co., Ltd.), and then charged with QAE-Sephadex previously equilibrated with the buffer solution A (pH 7.5).
  • Column (diameter 6 cm x 15 cm)
  • the enzyme was adsorbed. Then, after washing with the buffer A containing 0.15 M ammonium sulfate (pH 7.5) to remove unnecessary proteins, the buffer A containing 0.15 M ammonium sulfate (pH 7.5 ) And a buffer containing 0.8 M ammonium sulfate (pH 7.5), and eluted with 3 liters of a concentration gradient buffer.
  • the active part was concentrated to 2 ml using an ultrafiltration membrane device (fraction: 100,000) manufactured by Namicon Co., Ltd. Gel filtration was performed on a TSK gel G300 SWXL column (0.76 cm x 30 cm x 2) equilibrated with 0 mM HE PES buffer (pH 7.5).
  • This fraction is a sample of this enzyme determined to be homogeneous by SDS-polyacrylamide gel electrophoresis and has a total protein content of 6.65 mg, a total activity of 66.0 U, and a specific activity of 9 .92 UZmg.
  • the amount of enzyme that produces 1 ⁇ mo 1 of ATP per minute at 37 ° C is defined as one unit.
  • examples of the luciferin used in the present invention include luciferin, a norrecifulin derivative, a luciferin precursor, and the like.
  • Examples of the luciferin derivative include the following.
  • Examples of the luciferin precursor include 2-cyano-6-methoxybenzothiazole (2-cyano-6-methoxyxybenzothiazol, D-cysteine, and the like).
  • luciferase for example, the following beetles (Coleoptera)
  • Luciferases derived from Coloptera mutated luciferases thereof, and luciferases derived from non-beetles.
  • metal salts include divalent metal salts, for example, magnesium salts such as magnesium sulfate and magnesium chloride, and manganese salts such as manganese sulfate and manganese chloride.
  • the following are examples of enzymes that catalyze the reaction of producing ATP from ADP.
  • Phospholipin kinase Phospho ribu 1okinase
  • D ribulose ⁇ 1,5-diphosphate
  • Substrate Carbamoinolenic acid (Carb amoyle ep pho s p a te) (7) Phosphoglycerate kinase (Pho spho gly c e r a te)
  • Substrate 3-Phospho-D-glyceric acid (3-Phospho-D-glycererte)
  • Substrate L-aspartic acid 4-monophosphate (L-Aspartate4—phosphate) and the like.
  • examples of the reagents for extracting intracellular components of microorganisms include surfactants (benzetonium chloride, benzalkonium chloride, triton XI 00, etc.), methanol, ethanol, a mixed solution of ethanol and ammonium, an aqueous solution of trichloroacetic acid, and perchloric acid.
  • surfactants benzetonium chloride, benzalkonium chloride, triton XI 00, etc.
  • methanol ethanol
  • ethanol a mixed solution of ethanol and ammonium
  • an aqueous solution of trichloroacetic acid trichloroacetic acid
  • perchloric acid perchloric acid
  • the RNase refers to an enzyme that catalyzes a reaction for producing 5′-mononucleotide (AMP, GMP, CMP, UMP) from RNA, and examples thereof include those described below.
  • nuclease 'S-Pin' has a nuclease 'Py' (Nuclease P,), Mung bean nuclease (Mu ngbeansnuc 1 ease), Neuroshola * classa 'nuclease (eurosporacrassa nu clease), etc. are strongly included.
  • ATP which is a component inherently contained in an enzyme activator, an enzyme stabilizer, a buffer, and a cleanliness test reagent and which causes background luminescence is also included. It is preferable to add one or more of erasing agents.
  • the activator for the enzyme include ammonium sulfate (an activator for pyruvate orthophospho-todikinase).
  • Enzyme stabilizers include dithiothreitol and EDTA (a luciferase stabilizer).
  • ATP elimination agent of the cleanliness test reagent examples include ATP-degrading enzymes, such as adenosine phosphate deaminase (a method for measuring enzyme activity: see Japanese Patent Application Laid-Open No. Hei 9-118600).
  • P PDK Pyruvate orthophosphate dikinase
  • Luciferase 'A concentration of 0.1 mg / m 1 (final concentration) or more, especially 0.5 to 20 mg / ml (final concentration).
  • OmM final concentration
  • Ammonium sulfate A concentration that is higher than, especially, 5.0 to 10 OmM (final concentration). (7) Ammonium sulfate:
  • Adenosine phosphate kinase a concentration of 0.001 UZm 1 or more, especially 0.01 U / m 1 to 10 U / m 1.
  • the carrier to which dirt is collected is brought into contact with the inspection site to obtain a carrier to which the dirt is adhered, and then sterile water is brought into contact with the carrier.
  • the measurement is performed by adding a cleanliness test reagent containing phosphate dikinase, phosphoenorubyric acid, pyrophosphate, luciferin, lumferase and a metal salt, and measuring the amount of luminescence generated.
  • a carrier for collecting dirt is brought into contact with the inspection site to obtain a carrier to which the dirt has adhered, and then a reagent for extracting an intracellular component of the microorganism is brought into contact therewith.
  • a cleanliness inspection reagent comprising at least one member selected from the group consisting of enzymes that catalyze the reaction of ribonucleic acid and RNase; The reaction is performed by measuring the amount of luminescence generated.
  • test object or its processed product is combined with pyruvate orthophosphate dikinase, phosphoenolpyruvate, pyrophosphate, noresifurin, lucifer
  • the reaction is performed by reacting with a cleanliness test reagent containing a razor and a metal salt, and measuring the amount of luminescence generated.
  • Inspection points include cooking utensils such as cutting boards, kitchen knives and shamogi, machinery for pharmaceutical production, and food production machinery.
  • a soil-collecting carrier such as a cloth such as a cotton swab or a gauze, a non-woven fabric, or a sponge is brought into contact with these inspection points to obtain a soiled carrier.
  • a contact liquid obtained by bringing the extract into contact with an extraction reagent for a component in a microbial cell is prepared.
  • the concentration of these microbial intracellular component extraction reagents is preferably 0.001% to 1% .c.
  • a contact solution from which microbial intracellular components, such as ATP, ADP, AMP, and RNA, are extracted is prepared. .
  • the reagent acts on ATP, ADP, AMP, and RNA to cause a luminescence reaction, and the amount of luminescence generated is measured.
  • This reaction has the general formula As shown in the dashed box (a), the AMP, pyrophosphate, phosphoeno-rubyruvic acid and magnesium ions are reacted with pyruvate orthophosphite dikinase to produce ATP, pyruvate and phosphate. As shown by the dashed line (mouth), ATP, luciferin, dissolved oxygen and magnesium ions are reacted with luciferase to produce AMP, pyrophosphate, oxylluciferin, carbon dioxide and light. According to this reaction formula, which is characterized by a combination of reactions, first, ATP is consumed by the (mouth) reaction, light is emitted, and AMP and pyrophosphate are generated. ) Regenerated into ATP by the reaction of. This ATP is again subjected to the reaction of (mouth), and the ATP is consumed to emit light.
  • the luminescence reaction has the general formula
  • the presence of phosphoenorubyruvic acid and magnesium ion in the reaction (a) (as described above) and the reaction (mouth) (as described above) are newly indicated by the two-dot chain frame (c). It is characterized by a combination of a reaction that converts ATP and pyruvate by pyruvate kinase, an enzyme that catalyzes the reaction that generates ATP from ADP.
  • acetate kinase or creatine kinase may be used instead of pyruvate kinase as the enzyme that catalyzes the reaction of generating ATP from ADP used here.
  • reaction formulas for acetate kinase and creatine kinase are shown below. Reaction formula of acetate kinase
  • reaction 2 first, as shown by the two-dot chain line, a reaction in which ADP is reacted with pyruvate kinase in the presence of magnesium ion and phosphoenolpyruvate to convert it to ATP and pyruvate ( C), whereby ADP is converted to ATP, and then the ATP is consumed by the reaction (mouth) to emit light, AMP is generated, and then the AMP is converted to ATP by the reaction (ii). Is converted.
  • This ATP is again supplied to the reaction (mouth), and the ATP is consumed to emit light and is converted to AMP.
  • these two reactions are simultaneously and continuously repeated.
  • luminescence generated by the ATP conversion reaction system is stable for at least 10 minutes at a high level without decay.
  • RNA shown in the three-dot chain frame was reacted with RNase, and AMP, GM It is characterized by a combination of reactions (2) that convert it to P, CMP and UMP.
  • the RNA is reacted with RNase, and the reaction (2) is performed to convert it to AMP, GMP, CMP and UMP.
  • the AMP is converted to ATP by the reaction (a), and then the ATP is consumed by the reaction (mouth) to emit light and to generate AMP.
  • This AMP is again subjected to the reaction (a) and is converted into ATP. This ATP is then consumed by the reaction (mouth), emits light, and is converted into AMP.
  • the luminescence generated in this series of ATP conversion reaction systems is stable for at least 10 minutes at a high level without decay.
  • the inspection location (for example, 10 cm 2 ) is wiped off with a cotton swab moistened with sterile water to obtain a swab 5 with dirt attached to the dirt collection section 5a.
  • the luminous reagent storage tank 4 is pushed up from below the cylinder 1, and the thin film member 4 a of the storage tank is pressed against the sharp portion of the projection 8 to be cleaved.
  • the piston 2 is further depressed from the stop position, the wiping part 5a penetrates (cleaves) the other wall surface 7b of the extraction reagent storage tank 7, and the extract of dirt is dripped to store the cleanliness inspection reagent.
  • the mixture is mixed with the luminescent reagent in tank 4 to perform a luminescent reaction, and the amount of generated luminescence is measured with a luminometer, and the cleanliness is measured from the result.
  • PPDK Pyruvate orthophosphate dikinase
  • Phosphoenolpyruvate 4.2 mM Sodium pyrophosphate 200 / iM Luciferin 1.5 mM Luciferase (manufactured by Kikkoman) 4.5 mg / m 1 Magnesium sulfate (metal salt) 15 mM
  • EDTA is used to prevent enzyme inhibition by metal
  • dithiothreitol is used for enzyme stabilization
  • ammonium sulfate is used to enhance the activation of pyruvate orthophosphoditokinase
  • adenosine phosphate is also used.
  • Deaminase is used as an ATP scavenger for cleanliness test reagents, and is not an essential component.
  • a cleanliness test reagent consisting of the following combination of (1) a luminescent reagent and (2) a reagent for extracting components from microbial cells was prepared.
  • a cleanliness test reagent consisting of the following combination of (1) a luminescent reagent and (3) an RNA-degrading enzyme reagent was prepared.
  • the following cleanliness test reagents were prepared by combining (1) a luminescent reagent, (2) a reagent for extracting components from microorganism cells, and (3) an RNase reagent.
  • a cleanliness test reagent comprising the following combination of (1) a luminescent reagent and (2) a reagent for extracting components from microorganism cells was prepared.
  • Example 1 Same as Example 1 except that pyruvate kinase (manufactured by Sigma) 500 U / m 1 was added to the luminescent reagent of Example 1.
  • pyruvate kinase manufactured by Sigma 500 U / m 1 was added to the luminescent reagent of Example 1.
  • the following cleanliness test reagents were prepared by combining (1) a luminescent reagent, (2) a reagent for extracting components from microorganism cells, and (3) an RNase reagent.
  • Example 1 Same as Example 1 except that pyruvate kinase (manufactured by Sigma) 500 U / ml was added to the luminescent reagent of Example 1.
  • pyruvate kinase manufactured by Sigma 500 U / ml was added to the luminescent reagent of Example 1.
  • the cleanliness test reagent prepared in Example 1 can measure ATP and AMP.
  • a cleanliness test reagent for comparison was prepared in exactly the same manner as in the cleanliness test reagent of Example 1 except that pyruvate orthophosphoditokinase was not contained.
  • the cleanliness test reagent prepared in Comparative Example 1 can measure ATP, but cannot measure AMP.
  • Example 7 In the cleanliness test method of Example 7 described above, the procedure was the same except that the “cleanness test reagent” prepared in Comparative Example 1 was used instead of the “cleanliness test reagent” prepared in Example 1. The luminescence was measured.
  • Table 1 summarizes the respective light emission amounts obtained in Example 7 and Comparative Example 2 and the ratio of the former light emission amount to the latter light emission amount.
  • the cleanliness test method using the conventional luciferin-luciferase luminescence reagent could not detect AMP, and the cleanliness test reagent of the present invention ( It shows a lower value of luminescence compared to (luminescence reagent), making it impossible to perform highly sensitive and accurate cleanliness tests.
  • ATP concentration of the sample is too low (1 X 10-13 M or less) and cannot be measured (samples marked with * in Table 2), measure the sample at a concentration of 1% by weight. Then, it was converted to 0.001% by weight concentration by proportional calculation.
  • the total concentration was determined using a calibration curve showing the linearity obtained from the relationship between the ATP standard solution having a known concentration and the luminescence amount for the following reason.
  • the calibration curve obtained from the ATP concentration and its luminescence amount is almost the same as the calibration curve obtained from the AMP concentration and its luminescence amount, so the total concentration of ATP and AMP is the ATP concentration and its luminescence. It can be measured using a calibration curve obtained from the amount.
  • the cleanliness inspection method using the conventional ATP method cannot detect AMP as a pollution index, and shows a lower luminescence amount than the PPDK-containing luminescent reagent of the present invention, and provides highly accurate cleanliness. No inspection can be performed.
  • the sample is too thin if (in Table 2, ⁇ marked with samples corresponds) can not be measured (1 X 1 0 13 M or less) measures a sample of darker density, 0.1 in proportional calculation 0 0 Converted to 1% by weight concentration.
  • the total concentration was measured using a calibration curve showing ATP concentration and luminescence for the following reasons.
  • the calibration curve showing ATP concentration and luminescence is almost the same as the calibration curve showing AMP concentration and luminescence, so the total concentration of ATP and AMP is the same as the calibration curve showing ATP concentration and luminescence. For L, it can be measured.
  • test tube A and test tube B add the cleanliness test reagent (luminescent reagent) 1001 prepared in Example 1 and Example 3, respectively.
  • the AMP concentration was determined by converting it to ATP.
  • the rice scoop (inspection area) (10 cm 2 ) was wiped off with a cotton swab moistened with ultrapure water. This rice scoop is made by rinsing the rice scoop used to separate cooked rice gently with water and spraying a suspension of various bacteria separated from old cooked rice. is there.
  • ultrapure water 1001 was added to test tubes A and B, respectively, and the RNase reagents prepared in Examples 4 and 6 were added to test tubes C and D, respectively. Add 1001 and leave at room temperature for 15 minutes (decompose the RNA contained in the sample and convert it to AMP).
  • the luminescent reagents 1001 prepared in Examples 2, 5, 5, and 6 were added to Examiners A, B, C, and D, respectively, and the amount of luminescence measured by Berthold was measured. The amount of luminescence after 10 seconds was measured with a vessel LB9501 (Noreminometer).
  • the AMP concentration was determined by converting it to ATP.
  • Example 1 the luminescent reagent 1001 prepared in Example 1 and Example 3 were added to test tubes A and B, respectively, and a luminescence amount meter L B9501 manufactured by Berthold was used.
  • Luminometer 1 the luminescence amount after 10 seconds was measured.
  • the AMP concentration was determined by converting it to ATP.
  • test tube A and test tube B contain 100/1 ultrapure water, respectively, and test tube C and test tube D contain RNase prepared in Example 4 and Example 6, respectively.
  • the AMP concentration was determined by converting it to ATP.
  • the present invention uses a PDKK-containing luminescent reagent, ADP, AMP and RNA, which were difficult to measure with a conventional luciferin-lumpulase luminescent reagent, in addition to ATP, which is an indicator of contamination, were used. By measuring at the same time, even a small amount of dirt can be detected with high sensitivity and the cleanliness can be evaluated accurately.
  • the enzyme that catalyzes the reaction to generate ATP from ADP and Z or ribonuclease are added to the PPDK-containing luminescent reagent.
  • ADP and RNA are also measured at the same time, Unclean dirt can be detected with high sensitivity and more accurate cleanliness inspection can be performed.
  • FIG. 1 is a schematic view showing one specific example of an instrument for performing the cleanliness inspection method of the present invention.

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Abstract

L'invention a pour objet des réactifs servant à contrôler la propreté et permettant de mesurer non seulement l'ATP, qui sert à indiquer une salissure, mais aussi l'ADP, l'AMP et l'ARN difficilement mesurables au moyen de réactifs de luminescence à base de luciférine-luciférase utilisés dans les techniques précédentes. Ces réactifs permettent de détecter même une salissure peu importante et ont une sensibilité élevée qui permet d'évaluer la propreté avec beaucoup de précision. L'invention concerne également des procédés de contrôle de la propreté utilisant ces réactifs. Ces procédés consistent en ce qui suit: mettre en contact un support pour salissures et l'endroit à contrôler afin de récupérer le support comportant une salissure, qui y adhère; mettre ledit support en contact avec de l'eau aseptique ou avec un agent d'extraction pour composants microbiens intracellulaires; faire réagir la solution ainsi obtenue avec (1) un réactif de contrôle de la propreté comprenant orthophosphate dikinase de pyruvate, acide phosphoenolpyruvique, acide pyrophosphorique, luciférine, luciférase et un sel de métal ou (2) avec un réactif pour contrôler la propreté qui est obtenu par l'addition au réactif (1) d'au moins un élément sélectionné dans le groupe constitué d'enzymes catalysant la formation de l'ATP à partir de l'ADP et de ribonucléases et; mesurer l'intensité de luminescence.
PCT/JP1998/003761 1997-08-28 1998-08-25 Reactifs pour controler la proprete et procede de controle de la proprete Ceased WO1999011816A1 (fr)

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AU87503/98A AU8750398A (en) 1997-08-28 1998-08-25 Reagents for testing cleanness and methods for testing cleanness

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JP24598597A JPH1169997A (ja) 1997-08-28 1997-08-28 清浄度検査試薬およびその試薬を用いる清浄度検査法
JP9/245985 1997-08-28

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Cited By (3)

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WO2001081618A1 (fr) * 2000-04-26 2001-11-01 Kikkoman Corporation Procede de mesure de luminescence biologique
EP1264894A4 (fr) * 2000-01-17 2005-11-30 Satake Eng Co Ltd Systemes et procede de reaction de regeneration atp permettant d'examiner le nucleotide d'adenine, procede de detection d'arn et procede d'amplification d'atp
WO2021162123A1 (fr) * 2020-02-14 2021-08-19 キッコーマン株式会社 Composition liquide pour mesurer l'atp, et amp et/ou adp dans des échantillons

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JP2009172054A (ja) * 2008-01-22 2009-08-06 Olympus Medical Systems Corp 内視鏡管路清浄度検査装置
EP3184644A1 (fr) * 2015-12-22 2017-06-28 Omya International AG Dosage de la viabilité cellulaire microbienne pour détecter ou déterminer la contamination d'une pâte
JP7328763B2 (ja) * 2017-02-09 2023-08-17 キッコーマン株式会社 生体関連サンプル及び生体関連器具の清浄度測定キット及び方法
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JP7227008B2 (ja) * 2017-03-24 2023-02-21 キッコーマン株式会社 調理関連器具汚染度測定用キット及び調理関連器具洗浄プログラム評価方法

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