Method for the determination of the reduced form of nicotinamide adenine dinucleotide
First Claim
1. A method for the determination of NAD(P)H comprising the steps of:
- reacting a chromogen represented by formula (I);
##STR8## wherein Y is hydrogen or hydroxyl;
R1, R2 and R3 are independently selected from the group consisting of (II), (III) and (IV);
##STR9## wherein Z is independently selected from the group consisting of hydrogen, hydroxyl, optionally substituted amino, optionally substituted alkyl, alkenyl, optionally substituted aryl, acyl, halogen, nitro, suflo, carboxyl and alkoxy, wheren n is 1, 2 or 3 and the optional substituent is selected from the group consisting of alkyl, alkenyl, aryl, alkoxy, hydroxyl, carboxyl, sulfo, sulfonyl, halogen, amino, alkoxycarbonylamino, alkoxycarbonyl, aminoalkyl, acyl and nitro;
provided that at least one Z in R1, R2 and R3 is hydroxyl or optionally substituted amino;
with NAD(P)H in the presence of (1) peroxidase or thiol oxide reductase and (2) diaphorase or an electron carrier;
determining the pigment thus formed, said pigment being determined in the absence of Mn2+ or Co2+ ion; and
determining NAD(P)H from a calibration curve.
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Abstract
The present invention provides a method for the determination of NAD(P)H which comprises reacting a chromogen represented by the following general formula (I): ##STR1## wherein Y is hydrogen or hydroxyl; R1, R2 and R3 may be the same or different, and are groups represented by the following general formula (II), (III) or (IV): ##STR2## wherein Z is hydrogen, hydroxyl, amino, substituted amino, alkyl, substituted alkyl, alkenyl, aryl, substituted aryl, acyl, halogen, nitro, sulfo, carboxyl or alkoxy; n is 1, 2, or 3; provided that at least one Z in R1, R2 and R3 is hydroxyl, amino, or substituted amino; and Zs in Zn may be the same or different, with NAD(P)H in the presence of (1) peroxidase or thiol oxide reductase and (2) diaphorase or an electron carrier, and determining the pigment thus formed.
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Citations
15 Claims
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1. A method for the determination of NAD(P)H comprising the steps of:
- reacting a chromogen represented by formula (I);
##STR8## wherein Y is hydrogen or hydroxyl;
R1, R2 and R3 are independently selected from the group consisting of (II), (III) and (IV);
##STR9## wherein Z is independently selected from the group consisting of hydrogen, hydroxyl, optionally substituted amino, optionally substituted alkyl, alkenyl, optionally substituted aryl, acyl, halogen, nitro, suflo, carboxyl and alkoxy, wheren n is 1, 2 or 3 and the optional substituent is selected from the group consisting of alkyl, alkenyl, aryl, alkoxy, hydroxyl, carboxyl, sulfo, sulfonyl, halogen, amino, alkoxycarbonylamino, alkoxycarbonyl, aminoalkyl, acyl and nitro;
provided that at least one Z in R1, R2 and R3 is hydroxyl or optionally substituted amino;
with NAD(P)H in the presence of (1) peroxidase or thiol oxide reductase and (2) diaphorase or an electron carrier;
determining the pigment thus formed, said pigment being determined in the absence of Mn2+ or Co2+ ion; and
determining NAD(P)H from a calibration curve. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15)
- reacting a chromogen represented by formula (I);
Specification
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- Resources
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Current AssigneeKyowa Medex Co., Ltd. (Resonac Holdings Corp.)
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Original AssigneeKyowa Medex Co., Ltd. (Resonac Holdings Corp.)
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InventorsShimizu, Yoshiaki, Aoyama, Norihito, Miike, Akira, Tatano, Toshio
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Primary Examiner(s)NOT, DEFINED
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Assistant Examiner(s)NOT, DEFINED
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Application NumberUS06/706,404Time in Patent Office1,517 DaysField of Search435/25, 435/28, 435/26, 260/386, 260/388, 260/389US Class Current435/25CPC Class CodesC12Q 1/008 for determining co-enzymes ...C12Q 1/28 involving peroxidaseC12Q 2326/00 Chromogens for determinatio...
