US3703591A - Triglyceride hydrolysis and assay - Google Patents

Triglyceride hydrolysis and assay Download PDF

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Publication number
US3703591A
US3703591A US98904A US3703591DA US3703591A US 3703591 A US3703591 A US 3703591A US 98904 A US98904 A US 98904A US 3703591D A US3703591D A US 3703591DA US 3703591 A US3703591 A US 3703591A
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Prior art keywords
glycerol
lipase
accordance
pyruvate
nadh
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Expired - Lifetime
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US98904A
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Giovanni Bucolo
Harold David
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CNA Holdings LLC
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Calbiochem Corp
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Assigned to AMERICAN HOECHST CORPORATION reassignment AMERICAN HOECHST CORPORATION MERGER (SEE DOCUMENT FOR DETAILS). EFFECTIVE DATE: 12/29/81 Assignors: CALBIOCHEM-BEHRING CORP.
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Classifications

    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12QMEASURING 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
    • C12Q1/00Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/61Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving triglycerides
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12QMEASURING 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
    • C12Q1/00Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/34Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving hydrolase
    • C12Q1/44Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving hydrolase involving esterase
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/02Food
    • G01N33/04Dairy products
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10STECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00Chemistry: molecular biology and microbiology
    • Y10S435/8215Microorganisms
    • Y10S435/822Microorganisms using bacteria or actinomycetales
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10STECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00Chemistry: molecular biology and microbiology
    • Y10S435/8215Microorganisms
    • Y10S435/822Microorganisms using bacteria or actinomycetales
    • Y10S435/886Streptomyces
    • Y10S435/897Streptomyces griseus
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10STECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00Chemistry: molecular biology and microbiology
    • Y10S435/8215Microorganisms
    • Y10S435/911Microorganisms using fungi
    • Y10S435/939Rhizopus

Definitions

  • a number of alternative enzymatic procedures are provided for the glycerol assay, all in the original aqueous medium, such as conversion to glycerol-l-phosphate with glycerol kinase; conversion of ATP to ADP by the same reaction followed by conversion of the ADP plus phosphoenolpyruvate using pyruvate kinase to ATP and pyruvate ion followed by determining the latter; and alternatively con verting the pyruvate ion so formed with NAD-H using lactate dehydrogenase to lactate ion and NAD and determining the latter by optical measurement.
  • This invention relates to a rapid method of determining fatty acid glycerol esters in various aqueous media, such as serum, milk, and the like.
  • An object of the present invention is to provide a rapid and accurate procedure which liberates glycerol from its esterified form as a fatty acid ester, for example when present in aqueous media such as serum, and which permits the glycerol to be assayed by a variety of alternative methods without the necessity of isolating the glycerol from the medium under test.
  • a lipase which may be of plant or animal origin, but among which we prefer and find best a microbial lipase, such as the lipase from Chromobacterium viscosum, variant paralipolyticum, crude or purified, the lipase from Rhizopus delemar, purified, for example as noted in Fukumoto et al., J. Gen. Appl. Microbiol., 10, 257-265 (1964); and lipases having similar activity; and second, a protease.
  • a lipase which may be of plant or animal origin, but among which we prefer and find best a microbial lipase, such as the lipase from Chromobacterium viscosum, variant paralipolyticum, crude or purified, the lipase from Rhizopus delemar, purified, for example as noted in Fukumoto et al., J. Gen. Appl. Microbiol., 10, 257-265 (1964); and lipases having similar activity; and second
  • Proteases in general maybe used, such as by way of example and not by limitation, chymotrypsin, trypsin, Streptomyces griseus protease (commercially available under the registered trademark Pronase), elastase, papain, and bromelin. Mixtures of these may also be employed. This simple enzyme mixture is in general sufficient, although the hydrolysis may generally be expedited somewhat by the simultaneous inclusion of a simple protein such as serum albumin, egg albumin, globulins, and the like.
  • the glycerol liberated by the action of the enzyme mixture just described may be assayed in a number of ways, although we prefer one in particular which will now be described.
  • a useful and preferred embodiment of our invention comprises an assay mixture containing the preferred constituents to carry out a single triglyceride assay in accordance with the invention.
  • the preferred procedure involves the enzymatic hydrolysis as already described using the enzyme combination disclOsed, followed by the conversion of the liberated glycerol as likewise described
  • Vial A may contain all the components needed for the assay except the glycerol kinase.
  • the latter may be placed in Vial B.
  • other distributions of the components are possible, any of those involved solely with glycerol conversion and subsequent steps being includable in Vial B if desired.
  • Bovine serum albumin 5.0 mg.
  • u-Chymotrypsin 1100 NLF. units Lipase, Rlzizopus delemar: 1200 lipase units (Total volume: 3 ml.)
  • One (1) lipase unit is the quantity of enzyme which will release fatty acids from a substrate of triolein to require 1 ml. of 0.05 N potassium hydroxide for neutrali zation after a 30-min. incubation period at 37 C.
  • N.F. unit of tar-chymotrypsin is that quantity of enzyme which will produce an absorbance change of 0.0075/minute at 237 nanometers when incubated with a substrate of N-acetyl-L-tyrosine ethyl ester under the conditions of assay.
  • Vial B Glycerol 'kinase 2 I.U. (International Units)
  • the assay in accordance with the invention is carried out by adding an aliquot of liquid containing the triglyceride to be assayed, which may be for example 50 #1. of serum, to the 3 ml. contents of Vial A. This is then incubated at between 25 C. and 37 C. for approximately ten minutes. The optical density is then determined at 340 nm. Then the 2 I.U. of glycerol kinase contained in Vial B is added, and the mixture allowed to stand for an additional minutes at the same temperature, whereupon the optical density is determined again. The difference is proportional to the triglyceride content of the aliquot.
  • the essential contents of the reaction mixture in accordance with the invention comprises a microbial lipase; a protease; pyruvate kinase; lactate dehydrogenase; NADH; ATP; Phosphoenol pyruvate; a magnesium ion source; a buffer; and glycerol kinase.
  • An alternative procedure consists in omitting the NADH and the lactate dehydrogenase from the mixture described above, so that only the first three reaction steps depicted hereinabove occur; and adding a sufiicient quantity of dinitrophenyl hydrazine to react with the pyruvate ion formed in the aforementioned third reaction step.
  • the reaction product is colored when alkalized, and may be readily measured by a colorimeter.
  • a further alternative procedure is to omit the phosphoenol pyruvate; the pyruvate kinase; and the lactate dehydrogenase from the reaction mixture previously described, and to add instead glycerol phosphate dehydrogenase, whereupon the a-glycerol phosphate and the NAD are converted respectively to dihydroxyacetone phosphate and NADH.
  • the amount of NADH formed which is proportional to the amount of triglyceride originally present, can conveniently be measured by the increase in fluorescence.
  • An alternative sub-procedure is to include also a sufficient quantity of dinitrophenyl hydrazine in this system as well which will result in the formation of a colored reaction product with the dihydroxyacetone phosphate liberated.
  • the amount of the reaction product can then readily be determined with a colorimeter and is likewise a measure of the triglyceride originally present.
  • a still further alternative procedure is to utilize a reaction mixture which provides only the enzymatic conversion to glycerol which is the first reaction step set forth in the tabulation hereinabove.
  • the reaction mixture also includes NAD and glycerol dehydrogenase.
  • NAD glycerol dehydrogenase
  • the glycerol which is liberated in the first step is converted to dihydroxyacetone with the simultaneous production of NADH.
  • the increase in optical density at 340 nm. then becomes a measure of the quantity of triglyceride originally present, as previously described.
  • a suitable glycerol dehydrogenase is that obtainable from Enterobacter aerogenes; this enzyme is commercially available.
  • An alternative subprocedure here is to add dinitrophenyl hydrazine, which forms a colored reaction product with the dihydroxy acetone, so that the latter may then be measured colorimetrically.
  • the NADH which is quantitatively formed from the triglyceride may be caused to transfer its hydrogen (becoming oxidized in the process) to the tetrazolium salt, again quantitatively, through the mediation of any of several known substances, among which we prefer especially diaphorase or, alternatively, phenazine methosulfate.
  • the amount of dye thus formed may be readily measured by colorimetry, i.e., by carrying out optical density change measurements in the visible region.
  • the lipase unit has been defined hereinabove.
  • the international unit of proteolytic activity is the amount of protease which causes a turnover of one micromol per minute of a substrate which is specific for the particular enzyme in question, under conditions approximating an optimum for the system considered.
  • the substrate is tyrosine ethyl ester
  • the turnover rate may be determined in any number of ways, as by the change in optical density at 237 nm., or by determining the amino acid liberated, as phenol reagent tyrosine equivalents or by formol titration.
  • the N.F. (National Formulary) unit is occasionally used for proteases, and appears in an example hereinabove. Since 1 I.U. unit is equivalent to 28 NP. units, it will be seen that the 1100 NR units of chymotrypsin in the example is equal to 39 I.U. Since 1200 lipase units were present in the exemplary mixture, it will be seen that for each 1000 lipase units, our example shows about 32 LU. of protease.
  • one of the vials e.g., Vial A
  • the second vial e.g., Vial B
  • the remaining components may be distributed as desired between the two vials.
  • Our preferred distribution has been set forth hereinabove.
  • the glycerol liberated is that quantity to be expected on the basis of complete hydrolysis of the triglycerides present, the latter being determined by standard procedures well known in the art.
  • a-glycerol phosphate may also be named as glycerol-l-phoshpate; and that the fatty acid glycerol esters in serum may be and generally are referred to simply as triglycerides.
  • one unit of lipase means the amount of lipase equivalent to one lipase unit as specified hereinabove.
  • lipase is chosen from the class. consisting of Rhizopus delemar and Chromobacferium viscoswm lipases, and mixtures thereof.
  • protease is chosen from the class consisting of chymotrypsin, trypsin, streptomyces griseus protease, elastase, papain, bromelin, and mixtures thereof.
  • a process in accordance with claim 4 wherein from about 5 to about 500 LU. of said protease is present for each 1000 units of said lipase.
  • a process in accordance with claim 2 wherein from about 5 to about 500 LU. of said protease is present for each 1000 units of said lipase.
  • protease is chosen from the class consisting of chymotrypsin, trypsin, streptomyces griseus protease, elastase, papain, bromelin, and mixtures thereof.
  • a process in accordance with claim 1 wherein, subsequent to said liberation of glycerol, adenosine triphosphate and glycerol kinase are added to said liquid in sufficient quantity to convert said glycerol to glycerol-l-phosphate and said adenosine triphosphate to adenosine diphosphate; and in which said adenosine diphosphate is assayed, whereby the original content of said fatty acid glycerol ester may be determined.
  • a process in accordance with claim wherein to said aqueous liquid containing said glycerol-l-phosphate, nicotinamide adenine dinucleotide and glycerol phosphate dehydrogenase are added in sufficient quantity to convert said glycerol-l-phosphate and said NAD to dihydroxyacetone phosphate and nicotinamide adenine dinucleotide, reduced (NADH) respectively.
  • said hydrogen transfer agent is selected from the class consisting of diaphorase and phenazine methosulfate.
  • a process in accordance with claim 10 wherein to said aqueous liquid containing said glyceroll-phosphate in said ADP, there are added phosphoenol pyruvate and pyruvate kinase in sufficient quantity to convert said phosphoenol pyruvate to pyruvate ion, and in which said pyruvate ion is assayed to give an indication of the amount of said glycerol ester originally present.
  • a process in accordance with claim 15 wherein to said aqueous liquid containing said pyruvate ion there are added NADH and lactate dehydrogenase in suflicient quantity to convert said pyruvate and said NADH to lactate ion and NAD respectively.
  • a reagent combination for the analysis of fatty acid glycerol esters which comprises a first vial containing a lipase and a protease; a second vial containing glycerol kinase; and pyruvate kinase; lactate dehydrogenase; NADH; ATP; phosphoenol pyruvate; a magnesium ion source; and a buffer in any preselected distribution in said vials.
  • a reagent combination in accordance with claim 28 wherein from about 5 to about 500 LU. of said protease is present for each 1000 units of said lipase.

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US98904A 1970-12-16 1970-12-16 Triglyceride hydrolysis and assay Expired - Lifetime US3703591A (en)

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JP (2) JPS549518B1 (fr)
AU (1) AU475955B2 (fr)
BE (1) BE776034A (fr)
BR (1) BR7108185D0 (fr)
CA (1) CA955161A (fr)
CH (2) CH563404A5 (fr)
CS (1) CS212729B2 (fr)
DE (1) DE2162325C3 (fr)
ES (1) ES395956A1 (fr)
FR (1) FR2118454A5 (fr)
GB (1) GB1373106A (fr)
IL (1) IL38235A (fr)
IT (1) IT972079B (fr)
NL (1) NL180523C (fr)
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ZA (1) ZA717957B (fr)

Cited By (48)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS49113695A (fr) * 1973-02-27 1974-10-30
US3862009A (en) * 1972-06-19 1975-01-21 Boehringer Mannheim Gmbh Determination of triglycerides
US3898130A (en) * 1974-03-18 1975-08-05 American Hospital Supply Corp Rapid enzymatic hydrolysis of triglycerides
US3964974A (en) * 1972-09-28 1976-06-22 Merck Patent Gesellschaft Mit Beschrankter Haftung Enzymatic determination of glucose
US4001089A (en) * 1970-01-21 1977-01-04 The Dow Chemical Company Method for determination of triglycerides and glycerol
US4011045A (en) * 1975-02-14 1977-03-08 Bonderman Dean P Turbidity reduction in triglyceride standards
US4012287A (en) * 1975-11-18 1977-03-15 Dr. Bruno Lange Gmbh Method and reagent for the quantitative analysis of triglycerides
US4014744A (en) * 1975-01-30 1977-03-29 Miles Laboratories Inc. Processes for measuring tri-, di- and monoglycerides
US4019961A (en) * 1974-03-14 1977-04-26 Boehringer Mannheim G.M.B.H. Analytical enzymatic determination
US4038146A (en) * 1974-06-07 1977-07-26 Latron Laboratories, Inc. Method of determination of serum triglycerides and reagents
US4045297A (en) * 1975-12-15 1977-08-30 Monsanto Company Triglycerides determination method
US4071413A (en) * 1975-07-30 1978-01-31 Ono Pharmaceutical Co., Ltd. Method for determining free fatty acids in blood serum using fatty acid activating enzymes
DE2737286A1 (de) * 1976-08-19 1978-02-23 Eastman Kodak Co Mehrschichtige analytische elemente fuer die bestimmung von triglyceriden und glycerin
US4142938A (en) * 1974-03-20 1979-03-06 The Dow Chemical Company Determination of triglycerides and glycerol
US4168203A (en) * 1974-04-30 1979-09-18 Fujisawa Pharmaceutical Co., Ltd. Quantitative analysis of neutral lipids and lecithin
US4178285A (en) * 1978-12-20 1979-12-11 Felts James M Separation of active α1 -acid glycoprotein and utilization in the lipoprotein lipase enzyme system
WO1980000260A1 (fr) * 1978-07-13 1980-02-21 American Hospital Supply Corp Determination de triglycerides et agents reactifs enzymatique
US4195126A (en) * 1977-10-04 1980-03-25 The Board Of Trustees Of The University Of Alabama Albumin-dye complex for fatty acid determination
US4241178A (en) * 1978-01-06 1980-12-23 Eastman Kodak Company Process and composition for the quantification of glycerol ATP and triglycerides
US4245041A (en) * 1977-12-07 1981-01-13 American Monitor Corporation Triglycerides assay and reagents therefor
US4259440A (en) * 1979-05-21 1981-03-31 Miles Laboratories, Inc. Hydrolysis and assay of triglycerides
US4264589A (en) * 1978-12-20 1981-04-28 Felts James M Separation of active α1 -acid glycoprotein and utilization in the lipoprotein lipase enzyme system
US4273870A (en) * 1978-07-18 1981-06-16 Boehringer Mannheim Gmbh Method and composition for the determination of glycerol
US4275151A (en) * 1977-02-03 1981-06-23 Eastman Kodak Company Hydrolysis of protein-bound cholesterol esters
US4275152A (en) * 1977-02-03 1981-06-23 Eastman Kodak Company Hydrolysis of protein-bound cholesterol esters
US4302536A (en) * 1978-08-15 1981-11-24 Longenecker Robert W Colorimetric immunoassay process
US4309502A (en) * 1980-06-30 1982-01-05 Beckman Instruments, Inc. Enzymatic assay for glycerol and triglycerides and a reagent for use therein
EP0044615A2 (fr) * 1980-07-21 1982-01-27 TECHNICON INSTRUMENTS CORPORATION (a New York corporation) Méthode pour l'analyse de triglycérides
EP0045031A1 (fr) * 1980-07-22 1982-02-03 Baker Instruments Corporation Réactif pour la détection de glycérol et son application
US4322496A (en) * 1980-04-17 1982-03-30 Eastman Kodak Company Inhibition of lactate oxidase
US4394444A (en) * 1982-06-21 1983-07-19 Miles Laboratories, Inc. Cofactor indicator compositions
US4394445A (en) * 1979-02-22 1983-07-19 Nix Paul T Enzymatic glyceride hydrolysis
US4626511A (en) * 1985-05-13 1986-12-02 Wayne State University Composition for reducing turbidity in samples of biological fluids
US4636465A (en) * 1983-04-21 1987-01-13 Amano Pharmaceutical Co., Ltd. Method and composition for determination of glycerol
US4693971A (en) * 1983-01-28 1987-09-15 Toyo Jozo Kabushiki Kaisha Highly sensitive enzyme assay method
US4923796A (en) * 1978-08-08 1990-05-08 Boehringer Mannheim Gmbh Method for the quantitative enzymatic determination of ADP
US5310679A (en) * 1985-05-13 1994-05-10 Artiss Joseph D Composition for reducing turbidity in samples of biological fluids
EP1329512A1 (fr) * 2000-09-28 2003-07-23 ARKRAY, Inc. Procede de production de produits de degradation de proteines
US20040137546A1 (en) * 2002-12-24 2004-07-15 Stepan Company Method for determination of free and combined glycerin in biodiesel
US20040157285A1 (en) * 2001-09-28 2004-08-12 Kaori Ishimaru Method of storing tetrazolium compound, stabilizer for use therein, and tetrazolium compound reagent solution stored by the method
US20080299543A1 (en) * 2007-05-31 2008-12-04 Uti Limited Partnership Method for Assessing Trace Element Related Disorders in Blood Plasma
US20110112186A1 (en) * 2008-02-29 2011-05-12 Isis Innovation Limited Diagnostic methods
WO2014066787A1 (fr) 2012-10-26 2014-05-01 Boston Heart Diagnostics Corporation Bilan diabétique
US9198890B2 (en) 2011-10-13 2015-12-01 Boston Heart Diagnostics Corporation Compositions and methods for treating and preventing coronary heart disease
US9696276B2 (en) 2008-09-27 2017-07-04 Boston Heart Diagnostics Corporation Methods for separation and immuno-detection of biomolecules, and apparatus related thereto
US9739790B2 (en) 2014-11-17 2017-08-22 Boston Heart Diagnostic Corporation Cardiovascular disease risk assessment
US9817001B2 (en) 2008-05-27 2017-11-14 Boston Heart Diagnostics Corporation Methods for determining LDL cholesterol treatment
US9828624B2 (en) 2013-07-24 2017-11-28 Boston Heart Diagnostics Corporation Driving patient compliance with therapy

Families Citing this family (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CA1056282A (fr) * 1974-03-25 1979-06-12 Charles T. Goodhue Elements multicouches pour le dosage du cholesterol
US3884764A (en) * 1974-03-25 1975-05-20 Eastman Kodak Co Method and composition for blood serum cholesterol analysis
CA1100023A (fr) * 1976-08-19 1981-04-28 Charles T. Goodhue Procede et composition pour le dosage du glycerol et des triglycerides
JPS6058746B2 (ja) * 1977-09-22 1985-12-21 中外製薬株式会社 高級脂肪酸エステル

Cited By (55)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4001089A (en) * 1970-01-21 1977-01-04 The Dow Chemical Company Method for determination of triglycerides and glycerol
US3862009A (en) * 1972-06-19 1975-01-21 Boehringer Mannheim Gmbh Determination of triglycerides
US3964974A (en) * 1972-09-28 1976-06-22 Merck Patent Gesellschaft Mit Beschrankter Haftung Enzymatic determination of glucose
JPS49113695A (fr) * 1973-02-27 1974-10-30
US4019961A (en) * 1974-03-14 1977-04-26 Boehringer Mannheim G.M.B.H. Analytical enzymatic determination
US3898130A (en) * 1974-03-18 1975-08-05 American Hospital Supply Corp Rapid enzymatic hydrolysis of triglycerides
US4142938A (en) * 1974-03-20 1979-03-06 The Dow Chemical Company Determination of triglycerides and glycerol
US4168203A (en) * 1974-04-30 1979-09-18 Fujisawa Pharmaceutical Co., Ltd. Quantitative analysis of neutral lipids and lecithin
US4038146A (en) * 1974-06-07 1977-07-26 Latron Laboratories, Inc. Method of determination of serum triglycerides and reagents
US4014744A (en) * 1975-01-30 1977-03-29 Miles Laboratories Inc. Processes for measuring tri-, di- and monoglycerides
US4011045A (en) * 1975-02-14 1977-03-08 Bonderman Dean P Turbidity reduction in triglyceride standards
US4071413A (en) * 1975-07-30 1978-01-31 Ono Pharmaceutical Co., Ltd. Method for determining free fatty acids in blood serum using fatty acid activating enzymes
US4012287A (en) * 1975-11-18 1977-03-15 Dr. Bruno Lange Gmbh Method and reagent for the quantitative analysis of triglycerides
US4045297A (en) * 1975-12-15 1977-08-30 Monsanto Company Triglycerides determination method
DE2737286A1 (de) * 1976-08-19 1978-02-23 Eastman Kodak Co Mehrschichtige analytische elemente fuer die bestimmung von triglyceriden und glycerin
US4275151A (en) * 1977-02-03 1981-06-23 Eastman Kodak Company Hydrolysis of protein-bound cholesterol esters
US4275152A (en) * 1977-02-03 1981-06-23 Eastman Kodak Company Hydrolysis of protein-bound cholesterol esters
US4195126A (en) * 1977-10-04 1980-03-25 The Board Of Trustees Of The University Of Alabama Albumin-dye complex for fatty acid determination
US4245041A (en) * 1977-12-07 1981-01-13 American Monitor Corporation Triglycerides assay and reagents therefor
US4241178A (en) * 1978-01-06 1980-12-23 Eastman Kodak Company Process and composition for the quantification of glycerol ATP and triglycerides
WO1980000260A1 (fr) * 1978-07-13 1980-02-21 American Hospital Supply Corp Determination de triglycerides et agents reactifs enzymatique
US4223090A (en) * 1978-07-13 1980-09-16 American Hospital Supply Corporation Reagents for the enzymatic determination of triglycerides
US4273870A (en) * 1978-07-18 1981-06-16 Boehringer Mannheim Gmbh Method and composition for the determination of glycerol
US4923796A (en) * 1978-08-08 1990-05-08 Boehringer Mannheim Gmbh Method for the quantitative enzymatic determination of ADP
US4302536A (en) * 1978-08-15 1981-11-24 Longenecker Robert W Colorimetric immunoassay process
US4178285A (en) * 1978-12-20 1979-12-11 Felts James M Separation of active α1 -acid glycoprotein and utilization in the lipoprotein lipase enzyme system
US4264589A (en) * 1978-12-20 1981-04-28 Felts James M Separation of active α1 -acid glycoprotein and utilization in the lipoprotein lipase enzyme system
US4394445A (en) * 1979-02-22 1983-07-19 Nix Paul T Enzymatic glyceride hydrolysis
US4259440A (en) * 1979-05-21 1981-03-31 Miles Laboratories, Inc. Hydrolysis and assay of triglycerides
US4322496A (en) * 1980-04-17 1982-03-30 Eastman Kodak Company Inhibition of lactate oxidase
US4309502A (en) * 1980-06-30 1982-01-05 Beckman Instruments, Inc. Enzymatic assay for glycerol and triglycerides and a reagent for use therein
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BE776034A (fr) 1972-03-16
IT972079B (it) 1974-05-20
GB1373106A (en) 1974-11-06
SE389919B (sv) 1976-11-22
AU475955B2 (en) 1976-09-09
JPS549518B1 (fr) 1979-04-25
DE2162325C3 (de) 1980-12-11
JPS53114493A (en) 1978-10-05
NL180523C (nl) 1987-03-02
BR7108185D0 (pt) 1973-05-31
IL38235A (en) 1974-10-22
ZA717957B (en) 1972-08-30
CA955161A (en) 1974-09-24
CH566003A5 (fr) 1975-08-29
NL7117275A (fr) 1972-06-20
CS212729B2 (en) 1982-03-26
NL180523B (nl) 1986-10-01
CH563404A5 (fr) 1975-06-30
AU3641671A (en) 1973-06-07
DE2162325A1 (de) 1972-06-22
ES395956A1 (es) 1974-09-01
IL38235A0 (en) 1972-01-27
DE2162325B2 (de) 1980-04-24
FR2118454A5 (fr) 1972-07-28

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