urinary pyrrole (mauve factor): marker for oxidative stress in behavioral disorders

Urinary pyrrole (Mauve Factor): marker for oxidative stress in

behavioral disorders

Woody McGinnis MDmcginnis@mind.net

Seattle, 6 November 2004

CH3 C2H5

The Mauve Factor

OHHPL (hydroxyhemopyrrolin-2-one)

OHHPL (Mauve Factor)

• In human urine, blood and CSF

• Mistakenly identified as kryptopyrrole, a persistent erroneous term

• Chemically similar to kryptopyrrole, which can be used for OHHPL assay

CH3 C2H5 C2H5 CH3

HP KP

CH3 C2H5 C2H5 CH3

OHHPL OHKPL

Mauve history• Discovered in urine in 1957

• Named for lilac-colored appearance on paper chromatograms developed

with Erhlich's reagent

• Labile and elusive

• Abram Hoffer is the father of Mauve

Mauve Hall of FameHofferIrvineOsmondPfeifferSohlerCutlerO'Reilly

GrahamRiordanJacksonWalshAudhya

Europe

Hoffer J Neuropscyh 1961

• Qualitative Mauve assay

• All normals mauve-negative

• 27/39 early schizophrenics positive

• All 7 who recovered on niacinamide converted to negative

Hoffer 1961

• Relapses associated with reappearance of Mauve

• Apparent role in other behaviors: ETOH, depression.

• A "mentally retarded" mauve-positive child responded dramatically to

niacinamide

Hoffer and Mauve • Heat and light sensitive

• Relatives should be tested• Preventive potential

• 10/14 criminal / deviant positives• Report on 740 patients in 1966• All recovered schizophrenics

negative, unrecovered 50% positive

O'Reilly 1965

• Report on 850 behavioral patients

• 25% of "disturbed children" mauve-positive, vs 12% of well children

• First documented observation of Mauve association with stress

Mauve in schizophrenia

• Hoffer 1961, 1963, 1966

• Yutwiller 1962

• O'Reilly 1965

• Sohler 1967 x 2

High-Mauve and behavior

• Down syndrome 70%

• Schizophrenia 40-70%

• Autism 50%

• ADHD 30%

• ETOH 20-80%

Carl Pfeiffer 1972• "Sara" 15 y.o. with four years of unreality

spells, insomnia, seizures, attempted suicides, knee problems; quite well on

B6 1000 mg, Zn 160 mg, Mn 8 mg.

• Signs, symptoms, and clinical response imply high B6 / zinc need in high-Mauves.

• B6 and zinc quickly recognized by clinicians as main-stay treatment.

Mauve levels• Clinicians: behavioral symptoms in

individuals correlate with level• Irvine 1972: likelihood of depressive

reactions correlate with level• Cutler 1974: B6 dose needed to

normalize Mauve proportional to level• McCabe 1983: Mauve can be normalized with high-dose B6 only

Pfeiffer 1983• Symptoms may improve in 24 hours,

usually within 1 week

• May need months for full recovery

• Relapse within days or weeks if no nutrients

• Changing needs

OHHPL Levels and B6 (10mg/kg/day) + Zn (25mg) + Mg (400mg) in Autism

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Pfeiffer correlates

• Nail spots• Stretch marks• Pale skin• Poor tanning• Knees and joints• Constipation

• Dream recall• Morning nausea• Light and sound• Odor intolerance• Migraines• Stitch-in-side

Walsh• Low stress

tolerance• Anxious, overly

pessimistic• Explosive anger• Hyperactivity

Kruesi• Social

withdrawal• Emotionally

labile• Loss of appetite• Fatiguability

Mauve and stress

• Mauve is associated with stress, including, emotional stress.

Audhya 1992. Cold-immersion increased Mauve <1 hour

• The correlation is well-documented over decades

Mauve and stressO'Reilly 1965Sohler 1971

Pfeiffer "stress-dosing" 1973Ward says across all diagnoses 1975

Hippchem 1978McCabe 1983

Jaffe and Kruesi 1992

Non-behavioral Mauve• Acute Intermittent Porphyria

• Cutler 1974: High-mauve obesity and abnormal glucose tolerance

• Hoffer 1966: 33/99 Cancer patients, 7/8 lung cancer patients

• Riordan and Jackson: 43% of general medical patients: arthritis, chronic fatigue,

heart disease, hypertension, irritable bowel, migraine. Range 20-40 mcg%.

Mauve bumps in the road• U Michigan 1962: no pathological

importance in schizophrenia because found 34% Mauve-negative.

• U Cal 1969: their "simplified" assay produced phenothiazine false-positive

• India 1971: No Mauve in 120 psych patients--used HCl, 24° collection.

More bumps in the road..• Am J Psychiatr 1978: "Pyroluria a

poor marker in chronic schizophrenia" (based on 2/9 Mauve-positives)

• J Nutr 1979: "..urinary kryptopyrrole..proved invalid as screening test for vitamin dependent disorders.." (based on 6/20 Mauve-positives, all borderline. And no zinc.)

Bumps• U Cal 1975: "Non-occurrence of

kryptopyrrole and hemopyrrole in urine of schizophrenics by GC-MS"

• UC Berkeley 1978: GC/MS shows no kryptopyrrole or hemopyrrole in schizophrenics or controls

• Irvine 1977-78 confirms Mauve is OHHPL by synthesis.

Bumps..• Irvine, Nature 1969: Mauve identified

unequivocally as kryptopyrrole

• Irvine, in landmark Orthomolecular Psychiatry, "Mauve is kryptopyrrole"

• Irvine 1974: lactam of kryptopyrrole is the "identity of the natural kryptopyrrole"

Was Pfeiffer right about Mauve and low zinc?

Walsh. 1148 ADHD patients:

Plasma Zn vs colorimetric MauveStrong negative correlation

0.974 significance (F test)

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OHHPL

RB

C Z

inc

OHHPL vs.RBC Zinc

Correlation Coefficient -.985

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Mauve

WB

C Z

inc

Correlation Coefficient -.743

Mauve vsWBC Zinc

Mauve as clinical tool• Careful specimen collection

• Mild 20-30, moderate 30-40, severe over 40 mcg%

• Elevations imply zinc and B6 need

• Titrate nutrients to suppress Mauve

• Individualize adjunctive nutrients

Mauve is OHHPL• Graham, Univ Glasgow 1978 quantified

normal range by GLC• Audhya 1994-present: commercial OHHPL

by HPLC/MS and synthetic standard.• Strong logical imperative to cease

"kryptopyrrole" terminology

OHHPL facts• Irvine 1977: levels correlate with

emotional withdrawal, motor retardation, blocked affect and severe depression; IP to rats: ptosis, locomotor aberration, hypothermia

• Cutler 1990: IP to mice increased backward locomotion and head-twitching (as with psychotomimetics)

OHHPL facts• Photo, heat, and acid-labile• Urinary half-life 10-12 hours

• Nearly 100% urinary clearance intact after IP administration

• Daily excretion up to 1 mg • Urine, Blood, CSF; animal brain

• Graham: similarity to kainic acid and pyroglutamate suggests excitoxicity

Why do they both work?

Niacinamide (B3)OR

Vitamin B6 (P5P) and Zinc

Thinking points• B3, Zn and B6 are anti-oxidant

• Strong stress / Mauve association• Emotional stress clearly causes

oxidative stress• The behavioral and somatic high-

Mauve disorders feature high oxidative stress

High Oxidative Biomarkers• Down Syndrome• Schizophrenia

• Autism• ADHD

• Emotional Stress• Cancer and Inflammatory Disease

• Hyperglycemia

Zinc is anti-oxidant• Shields -SH groups

• Blocks lipid peroxidation and PLA2

• Induces metallothionein

• Constituent of SOD

• Maintains vitamin A

• Deficiency increases intestinal NO˙

Zinc deficiency increases oxidative stress

• Lower glutathione, vitamin E, GST, GSHPx and SOD

• Increased reactive species and lipid peroxides in tissue, membranes and

mitochondria

Oxidants mobilize zinc

• Oxidants release complexed zinc from zinc-binding proteins, including

metallothionein and albumin

• It is likely--but unproven--that zinc retention is reduced in direct

relationship to oxidative stress

Oxidative stress

Low zinc

B6 is anti-oxidant

• P5P for Glutathione, Metallothionein, CoQ10 and Heme synthesis

• With Zn, cofactor for GAD

• P5P protects vulnerable lysinyl groups, as in GSHPx

Marginal B6 deficiency:

Lowers GSHPx

Lowers glutathione reductase

Promotes mitochondrial decay

Raises lipid peroxide levels

B6 and oxidative stress• Binding of P5P-dependent enzymes

is subject to carbonyl inhibition

• Binding of key P5P-dependent enzymes such as GAD impaired by

oxidants generally

• OH˙ and 1O2 attack B6 vitamers

B6 and Mauve• B6 levels are normal

• Pfeiffer alluded to lower P5P and EGOT activity in high-Mauves

• Lower zinc may impair B6 activation• Oxidative stress affects activation of

B6 and binding of B6-dependent enzymes

B3 is anti-oxidant

• NADPH for reduction of glutathione

• Potent free-radical quencher: protects both lipids and proteins from oxidation

• Blocks NO˙ neurotoxicity

• High tissue levels: better lipoxidation prevention than ascorbate

B3 is anti-oxidant

• Niacin antagonists increase lipoxidation

• Low B3 decreases MT and increases apoptosis in brain cells

• Neuroprotective in experimental mitochondrial toxicity

Reciprocal relationships

Oxidative stress

Poor energetics Excitotoxicty

Require heme

• Cystathionine synthase

• Catalase• Heme-hemopexin

for MT translation• Pyrrolase

• Guanylate cyclase

• Cytochromes• Sulfite

reductase• NOS

OHHPL and heme• Durko 1970. Oxidized kryptopyrrole

binds heme in vitro

• Graham 1979. Microsomal levels 48 hours after IP OHHP to rats:

Heme down 42% Cytochrome p450 down 50%

Regulatory heme

• Sustains zinc, vitamin A and melatonin levels

• Differentiation, response to growth factors and resistance to viruses

• Levels lowered by toxins: gasoline, benzene, arsenic and cadmium

Low Heme is pro-oxidant• Ames 2002. Experimental heme

depression in cultured brain cells:

Decreased Complex IV 50% reduction in intracellular Zn

Increased intracellular FeIncreased NOS (x 7), NO2 and NO3

Heme depression

• Neuroblastoma cells fail to differentiate, axons degenerate

• Astrocytoma cells fail to proliferate

• Heme synthesis itself depends on sufficient B6 and Zn

HypothesisMauve may be a significant contributer

to oxidative stress. So, it may be a good biomarker for oxidative stress.

• Heme depression, with impaired energetics, zinc, and detox

• Mauve excitotoxicity

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Mauve

Glu

tath

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Mauve vs. RBC Glutathione

CorrelationCoefficient -.408

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Mauve

GS

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Mauve vs. GST

CorrelationCoefficient -.65087

p<.02

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OHHPL

Bio

tin

OHHPL vs. Biotin

CorrelationCoefficient -.973

A second hypothesis• Mauve may form as a result of

oxidative stress.• Sequential lipid oxidation, protein

adduction, oxidative side-chain scission and dissociation.

• Models: Isolevuglandin pyrroles and urinary pyrroles from hexane

Proposed mechanism

• Sequential lipid oxidation, protein adduction, oxidative side-chain

scission and dissociation.

• Models: Isolevuglandin pyrroles and urinary pyrroles from hexane

Oxidative by-product?Some consistencies:

• In the high-Mauve schizophrenics, membrane arachidonate, presumed substrate, is lower.

• Formation of the pyrrolic adduct occurs at lysinyl groups, so may impair pyridoxal kinase and the B6-dependent enzymes.

More Mauve research!• Controlled clinical trials

• Other therapeutic and toxic modulators

• Pro-oxidant and excitotoxic properties

• Prove origin

• Genetic predisposers

The Oxidative Stress in Autism Study

Principle Investigator: William WalshNIH Author: Robert Salomon

Collaborators: Bruce Ames, Allen Lewis, George Perry, Domenico

Pratico, Aristo VojdaniSponsors: Alexander and Bo MacInnis