Posted: 11 June 2005
Five recent abstracts on formaldehyde damage to cells -- vitamin E and selenium protect, Yoshiro Saito and Yasukazu Yoshida et al, O'Brien PJ et al, Costa M et al: Murray 2005.06.07
The abstract below shows that even when the formaldehyde levels alone are not enough to cause cell death, formaldehyde combined with a chemical to increase free radicals can dramatically reduce cell viability (and increase cell death). The researchers used AIPH to increase free radicals. As you may know, excitotoxins can also increase free radical production significantly.
As Dr. Blaylock says:
"Excitotoxins destroy neurons partly by stimulating the generation of large numbers of free radicals." See: http://www.dorway.com/blayart1.txt
[Rich Murray: Here is the abstract and four more that show that many groups are intensively studying cell damage and death from formaldehyde, and the role of chemicals that increase or decrease this damage from formaldehyde.]
"It is well known that formaldehyde (HCHO) and reactive oxygen species (ROS), such as free radicals, are cytotoxic as well as potentially carcinogenic. Although the individual effects of these reactants on cells have been investigated, the cytotoxicity exerted by the coexistence of HCHO and reactive radicals is poorly understood."
Toxicology. 2005 Jun 1; 210(2-3): 235-45.
Cytotoxic effect of formaldehyde with free radicals via increment of cellular reactive oxygen species.
Saito Y, Nishio K, Yoshida Y, Niki E.
It is well known that formaldehyde (HCHO) and reactive oxygen species (ROS), such as free radicals, are cytotoxic as well as potentially carcinogenic.
Although the individual effects of these reactants on cells have been investigated, the cytotoxicity exerted by the coexistence of HCHO and reactive radicals is poorly understood. The present study using Jurkat cells demonstrated that the coexistence of HCHO with water-soluble radical initiator, 2,2'-azobis-[2-(2-imidazolin-2-yl)propane] dihydrochloride (AIPH) dramatically decreased cell viability, and that under such conditions scant cell death was observable induced by either of the reactants alone.
Based on the results of phosphatidylserine exposure and caspase activation, this observed cell death, in fact, was apparently necrotic rather than apoptotic. To understand the mechanisms of the cell toxicity of HCHO and AIPH, we assessed two kinds of oxidative stress markers such as cellular glutathione (GSH) content and cellular ROS, and the DNA-protein cross-links, which formed as the result of HCHO treatment.
A marked decrease in total cellular GSH was observed not only in the case of the coexistence conditions but also with AIPH alone. Dichlorodihydrofluorescein (DCF) assay revealed that cellular ROS were synergistically increased before cell death. The formation of DNA-protein cross-links was observed in the presence of HCHO and AIPH, and the extent was similar to HCHO alone.
Co-incubation with semicarbazide, which inactivates HCHO, prevented this cell death induced by a combination of HCHO and AIPH. Semicarbazide also exhibited an inhibitory effect on the synergistic increment of cellular ROS and the formation of DNA-protein cross-links. These results suggest that the free radicals from AIPH induced GSH reduction, while HCHO resulted in the formation of DNA-protein cross-links, eventuating in a synergistic, incremental increase of cellular ROS and cell death brought about by this combination.
Free Radic Res. 2004 Apr; 38(4): 375-84.
Application of water-soluble radical initiator, 2,2'-azobis[2-(2-imidazolin-2-yl)propane] dihydrochloride, to a study of oxidative stress.
Yoshida Y, Itoh N, Saito Y, Hayakawa M, Niki E.
Human Stress Signal Research Center, National Institute of Advanced Industrial Science and Technology, 1-8-31 Midorigaoka, Ikeda, Osaka 563-8577, Japan. firstname.lastname@example.org
It is essential to generate free radicals at a controlled and constant rate for specific duration and at specific site to study the dynamics of oxidation and also antioxidation. Both hydrophilic and lipophilic azo compounds have been used for such purpose.
In the present work, the action of 2,2'-azobis[2-(2-imidazolin-2-yl)propane] dihydrochloride (AIPH) was examined and compared with those of 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) and 2,2'-azobis[2-methyl-N-(2-hydroxyethyl)-propionamide] (AMHP). The rate constant of free radical formation (ek(d)) for AIPH was 2.6 x 10(-6)/s at 37 degrees C in PBS (pH 7.4) solution, indicating that AIPH gives 3.8 times more free radicals than AAPH under the same conditions.
It was found that the dynamics of oxidation and antioxidation induced by AIPH can be studied satisfactorily in the oxidation in micelles, LDL and erythrocyte suspensions, plasma, and cultured cells. The extent of cell death induced by AIPH and AAPH was directly proportional to the total free radicals formed.
Interestingly, it was found that rats would not drink water containing AAPH, but they drank water containing AIPH.
The levels of 8-iso-prostaglandin F2alpha (8-isoPs), 7-hydroxycholesterol (FCOH), lysophosphatidylcholine in the plasma of rats given water containing 50 mM AIPH for 1 month increased compared with those of control rats which drank water without AIPH.
It may be concluded that AIPH is useful for kinetic and mechanistic studies on oxidative stress to membranes, lipoproteins, cells, and even animal models. PMID: 15190934
"Many reports have shown significant correlations between selenium deficiency and the incidence of cancer (51, 52). It has been also reported that selenium supplementation prevents the generation of cancer (2, 53). As shown in this study, selenium deficiency caused a significant increase in ROS and peroxidation inside cells. It is therefore considered that selenium-deficient conditions cause oxidative DNA damage that eventually leads to cancer formation. In conclusion, the present study clearly shows that selenium deficiency decreased the activities of cGPx, PH-GPx, and TR, increased lipid peroxidation in the membranes, and eventually induced cell death. The cell death was inhibited by other types of antioxidants with different functions, such as tocopherols and deferoxamine, which inhibit lipid peroxidation in the membranes and sequester redox-active iron, respectively. These results strongly indicate that the lipid hydroperoxides play a causative role in the oxidative damage to cells induced by selenium deficiency."
http://www.jbc.org/cgi/content/full/278/41/39428 Free full text
Originally published In Press as doi:10.1074/jbc.M305542200 on July 29, 2003
J. Biol. Chem., Vol. 278, Issue 41, 39428-39434, October 10, 2003
Cell Death Caused by Selenium Deficiency and Protective Effect of Antioxidants*
Yoshiro Saito * , email@example.com
Yasukazu Yoshida * , firstname.lastname@example.org
Takashi Akazawa ¶
Kazuhiko Takahashi ¶
Etsuo Niki *
* From the Human Stress Signal Research Center (HSSRC), National Institute of Advanced Industrial Science and Technology (AIST), 1-8-31 Midorigaoka, Ikeda, Osaka 563-8577, Japan and the ¶ Department of Immunology, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita 12 Nishi 6, Kita-ku, Sapporo, 060-0812, Japan
Received for publication, May 27, 2003, and in revised form, July 28, 2003.
Selenium is an essential trace element and it is well known that selenium is necessary for cell culture.
However, the mechanism underlying the role of selenium in cellular proliferation and survival is still unknown. The present study using Jurkat cells showed that selenium deficiency in a serum-free medium decreased the selenium-dependent enzyme activity (glutathione peroxidases and thioredoxin reductase) within cells and cell viability.
To understand the mechanism of this effect of selenium, we examined the effect of other antioxidants, which act by different mechanisms. Vitamin E, a lipid-soluble radical-scavenging antioxidant, completely blocked selenium deficiency-induced cell death, although -tocopherol (biologically the most active form of vitamin E) could not preserve selenium-dependent enzyme activity. Other antioxidants, such as different isoforms and derivatives of vitamin E, BO-653 and deferoxamine mesylate, also exerted an inhibitory effect.
Dichlorodihydrofluorescein (DCF) assay revealed that cellular reactive oxygen species (ROS) increased before cell death, and sodium selenite and -tocopherol inhibited ROS increase in a dose-dependent manner. The generation of lipid hydroperoxides was observed by fluorescence probe diphenyl-1-pyrenylphosphine (DPPP) and HPLC chemiluminescence only in selenium-deficient cells. These results suggest that the ROS, especially lipid hydroperoxides, are involved in the cell death caused by selenium deficiency and that selenium and vitamin E cooperate in the defense against oxidative stress upon cells by detoxifying and inhibiting the formation of lipid hydroperoxides.
Selenium is an essential trace element for humans and many other forms of life, and a deficiency of this element induces some pathological conditions, such as cancer, coronary heart disease, and liver necrosis (1-5). Selenium deficiency is also accompanied by a loss of immunocompetence (6), and both cell-mediated immunity and B-cell function can be impaired (7). Supplementation with selenium has marked immunostimulant effects, including an enhancement of activated T-cell proliferation (8).
Selenium is an essential component of several enzymes such as glutathione peroxidase (GPx)1 (9), thioredoxin reductase (TR) (10), and selenoprotein P (SeP) (11), which contain selenium as selenocysteine. It is also well known that selenium is essential for cell culture when a serum-free medium is used (12).
Serum-free media, especially for immune cells and neurons, contain insulin, transferrin, and sodium selenite. Without selenium, cells can neither proliferate nor survive. However, the underlying mechanism for the role of selenium in cell proliferation is still unknown.
Vitamin E, a generic term for tocopherols and tocotrienols, is one of the most potent lipid-soluble antioxidants (13). Vitamin E occurs in nature in at least eight different isoforms: -, -, -, and -tocopherols and -, -, -, and -tocotrienols (14).
Tocotrienols differ from the corresponding tocopherols only in their aliphatic tail. Vitamin E deficiencies have been implicated in some pathologic conditions, such as cancer, coronary heart disease, and liver necrosis (15, 16) and are also accompanied by a loss of immunocompetence (17).
It is well known that selenium and vitamin E show compensative effects and that a deficiency of both elements causes massive injury in some cases (18-20). In the present study, we characterize the nature of cell death caused by selenium deficiency and the cell death inhibitory effect of antioxidants including vitamin E. We also demonstrate the involvement of reactive oxygen species (ROS), especially lipid hydroperoxides, on the cell death.
The essential role of selenium in nutrition has been well established. It is also well known that selenium is necessary for cell culture when using a serum-free medium. Insulin (as a growth factor), transferrin (as an iron source), and selenite are added to the serum-free media for immune and neuronal cells. Although the effects of selenium on cell viability and the cellcycle progression has been reported (12, 32), the underlying mechanism of the protective effect of this element has not yet been elucidated. In the present study using Jurkat cells, a model of proliferating T lymphoma cells, the decrease in cell viability was observed when applying a serum-free medium without selenium.
This cell death was completely blocked by selenium-containing materials, except for ebselen, in a dose-dependent manner. It has been reported that these selenium-containing materials are incorporated and can be the cellular source of selenium used for synthesis of selenoprotein (22). SeP, which is a selenium-rich extracellular glycoprotein (11, 33, 34) that functions as selenium transport protein (22, 35, 36), was the most effective of the materials tested (Table I). Although we observed a loss of cell viability after 24 h, Jurkat cells duplicated for 20 h under the serum-free culture conditions. This observation suggests that cell death occurred after a single division. We speculate that the proliferating cells became selenium-deficient, and the divided cells contained almost half of the selenoenzyme activities, such as cGPx, PH-GPx, and TR.
In the presence of sodium selenite, these enzyme activies were retained or up-regulated in cells.
The decrease in TR activity was lower than that of cGPx and PH-GPx activities in the Jurkat cells cultured with selenium-deficient medium for 72 h (Fig. 5). Selenoproteins have been proposed to follow a hierarchy for selenium supply in that the amounts of certain selenoproteins decrease more rapidly under selenium-deficient conditions (37).
A previous study suggested that this was due in part to differences in the SECIS elements. Gasdaska et al. (38) demonstrated that the element of TR was highly active; therefore, TR levels would be better preserved when the selenium supply was limited as in a selenium-deficient medium.
It has been reported that these selenoproteins play an important role in the defense against oxidative stress (1, 39).
In the case of lipid hydroperoxides, it is known that PH-GPx, but not cGPx, is able to reduce lipid hydroperoxides, including phospholipid hydroperoxide and cholesterol hydroperoxide, directly (40, 41).
It has also been proved that overexpression of PH-GPx suppresses cell death due to oxidative damage induced by radical initiator and lipid hydroperoxide (42, 43). Moreover, Lewin et al. (44) reported that TR also plays a role in preventing oxidative damage induced by tert-butyl hydroperoxide and oxidized LDL, but the mechanism of the protection afforded by this selenoprotein against oxidative stress induced by lipid hydroperoxides is still unclear.
At present, these selenoproteins are assumed to play an important role in the defense against oxidative stress related to lipid hydroperoxides. Under selenium-deficient conditions, the decrease in these selenoproteins is speculated to cause peroxidation in the lipid layer inside cells. It is noteworthy that radical-scavenging antioxidants, such as -tocopherol, completely blocked the cell death caused by selenium deficiency, although -tocopherol did not affect the enzyme activity of selenoproteins.
-Tocopherol-supplemented Jurkat cells did not show any loss of cell viability despite the undetectable levels of cGPx and PH-GPx activity. Other isoforms of vitamin E, such as -, -, and -tocopherols and -, -, -, and -tocotrienols, were also effective.
Tocotrienols were more effective than the corresponding tocopherols (Table I), which may be ascribed primarily, if not solely, to the differences in the rate of cellular uptake.
A higher uptake of -tocotrienol than -tocopherol into culture cells has been reported (45, 46).
Such a difference was also observed for liposomal membranes (47). Vitamin E derivatives, such as PMC and its water-soluble analogue Trolox, also showed an inhibitory effect on cell death, but their higher ED50 values (Table I) suggest that the antioxidant incorporated into cell membranes is more effective than that localized outside the membranes.
The fact that selenium deficiency, which results in a decrease in the capacity to reduce lipid hydroperoxides, induced cell death and that this cell death could be inhibited by radical-scavenging antioxidants that suppress the formation of lipid hydroperoxides strongly indicates the causative role of lipid hydroperoxides in cell death.
The ED50 value of -tocopherol was as low as 36 nM (Table I). One may argue that this concentration is quite low compared with the physiological concentration; for example, 30 µM in human plasma. It should be pointed out, however, that the concentration of -tocopherol in the membrane is of more importance than that in the bulk phase.
In the present study, the lipid concentrations were measured as follows; FC, 4.4; PC, 12; PE, 5.2 nmol/106 cells.
Thus, the molar ratio of -tocopherol to total lipids in the cell culture system (106 cells/ml) was 36 x 10-9/22 x 10-6 M = 1/610 mol/mol, which is similar to that in human plasma; that is, 30 x 10-6/11 x 10-3 M = 1/370 mol/ mol.
It may be noted that the micromolar ranges of -tocopherol applied to many cell culture systems are not always physiological, but that the concentration in the membranes should be considered. Deferoxamine mesylate, a well-known iron chelator, was also found to be a potent inhibitor of cell death induced by selenium deficiency. It has been suggested that iron plays an important role in oxidative damage to cells (48, 49), by reacting with hydrogen peroxide or lipid hydroperoxides to form reactive oxygen radicals.
It was found that the removal of selenium from the culture medium induced ROS production as measured by DCF fluorescence and also lipid hydroperoxides as measured by DPPP fluorescence and HPLC chemiluminescence.
We are aware of the inherent drawbacks of DCFH, but it can be a useful probe for estimating semi-quantitatively the generation of ROS under specific conditions (50). Both selenium and -tocopherol suppressed ROS by apparently different mechanisms, the former by enhancing the reduction of hydroperoxides, while the latter by inhibiting their formation. FC-OOH was detected as a major lipid hydroperoxide, which was unexpected since polyunsaturated lipids in PC and PE are more susceptible to oxidation than free cholesterol.
One possible reason could be that the reduction of FC-OOH by PH-GPx is at least six times slower than that of phospholipid hydroperoxides (41). Many reports have shown significant correlations between selenium deficiency and the incidence of cancer (51, 52).
It has been also reported that selenium supplementation prevents the generation of cancer (2, 53).
As shown in this study, selenium deficiency caused a significant increase in ROS and peroxidation inside cells.
It is therefore considered that selenium-deficient conditions cause oxidative DNA damage that eventually leads to cancer formation. In conclusion, the present study clearly shows that selenium deficiency decreased the activities of cGPx, PH-GPx, and TR, increased lipid peroxidation in the membranes, and eventually induced cell death. The cell death was inhibited by other types of antioxidants with different functions, such as tocopherols and deferoxamine, which inhibit lipid peroxidation in the membranes and sequester redox-active iron, respectively.
These results strongly indicate that the lipid hydroperoxides play a causative role in the oxidative damage to cells induced by selenium deficiency.
* The costs of publication of this article were defrayed in part by the payment of page charges.
This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
To whom correspondence should be addressed: Human Stress Signal Research Center (HSSRC), National Institute of Advanced Industrial Science and Technology (AIST), 1-8-31 Midorigaoka, Ikeda, Osaka 563-8577, Japan. Tel.: 81-72-751-8293; Fax: 81-72-751-9964; E-mail: email@example.com
1 The abbreviations used are: GPx, glutathione peroxidase; ROS, reactive oxygen species; PBS, phosphate-buffered saline; HPLC, high pressure liquid chromatography; DCFH-DA, dichlorofluorescin diacetate; TR, thioredoxin reductase; pNA, p-nitroanilide; FITC, fluorescein isothiocyanate; SeP, selenoprotein P.
We express cordial thanks to Dr. Shuichi Shimakawa, Nanako Itoh, and Mieko Hayakawa in HSSRC, AIST for the total lipid analysis, and also to Dr. Kohji Ichimori in HSSRC, AIST for technical support.
Y. SAITO, Y. YOSHIDA, K. NISHIO, M. HAYAKAWA, and E. NIKI Characterization of Cellular Uptake and Distribution of Vitamin E Ann. N.Y. Acad. Sci., December 1, 2004; 1031(1): 368 - 375. [Abstract] [Full Text] [PDF]
C.-S. You, T. J. Sontag, J. E. Swanson, and R. S. Parker Long-Chain Carboxychromanols Are the Major Metabolites of Tocopherols and Tocotrienols in A549 Lung Epithelial Cells but Not HepG2 Cells J. Nutr., February 1, 2005; 135(2): 227 - 232. [Abstract] [Full Text] [PDF]
C. C. McCormick and R. S. Parker The Cytotoxicity of Vitamin E Is Both Vitamer- and Cell-Specific and Involves a Selectable Trait J. Nutr., December 1, 2004; 134(12): 3335 - 3342. [Abstract] [Full Text] [PDF] This Article
All ASBMB Journals Molecular and Cellular Proteomics Journal of Lipid Research Biochemistry and Molecular Biology Education
Copyright (c) 2003 by the American Society for Biochemistry and Molecular Biology.
"The toxicity and carcinogenicity of formaldehyde (HCHO) has been attributed to its ability to form adducts with DNA and proteins."
"This work demonstrates that DNA-protein cross-links can be formed in vitro following exposure to a variety of industrial compounds and that most cross-links are formed at cytotoxic concentrations."
"At higher HCHO concentrations, lipid peroxidation ensued followed by cell death."
"Antioxidants and iron chelators protected against HCHO cytotoxicity."
Chem Biol Interact. 2001 Jan 30; 130-132(1-3): 285-96.
The formaldehyde metabolic detoxification enzyme systems and molecular cytotoxic mechanism in isolated rat hepatocytes. Teng S, Beard K, Pourahmad J, Moridani M, Easson E, Poon R, O'Brien PJ. Faculty of Pharmacy, University of Toronto, 19 Russell St., Ont., M5S 2S2, Toronto, Canada.
[ O'Brien Prof Peter J THE LESLIE DAN FACULTY OF PHARMACY 416-978-2716 fax 416-978-8511 firstname.lastname@example.org ]
The toxicity and carcinogenicity of formaldehyde (HCHO) has been attributed to its ability to form adducts with DNA and proteins.
A marked decrease in mitochondrial membrane potential and inhibition of mitochondrial respiration that was accompanied by reactive oxygen species formation occurred when isolated rat hepatocytes were incubated with low concentrations of HCHO in a dose-dependent manner.
Hepatocyte GSH was also depleted by HCHO in a dose-dependent manner. At higher HCHO concentrations, lipid peroxidation ensued followed by cell death. Cytotoxicity studies were conducted in which isolated hepatocytes exposed to HCHO were treated with inhibitors of HCHO metabolising enzymes. There was a marked increase in HCHO cytotoxicity when either alcohol dehydrogenase or aldehyde dehydrogenase was inhibited.
Inhibition of GSH-dependent HCHO dehydrogenase activity by prior depletion of GSH markedly increased hepatocyte susceptibility to HCHO.
In each case, cytotoxicity was dose-dependent and corresponded with a decrease in hepatocyte HCHO metabolism and increased lipid peroxidation.
Cytotoxicity was also prevented, when cyclosporine or carnitine was added to prevent the opening of the mitochondrial permeability transition pore which further suggests that HCHO targets the mitochondria.
Thus, HCHO-metabolising gene polymorphisms would be expected to have toxicological consequences on an individual's susceptibility to HCHO toxicity and carcinogenesis. PMID: 11306052
"Chemicals such as cis-platinum, formaldehyde, chromate, copper, and certain arsenic compounds have been shown to produce DNA-protein cross-links in human in vitro cell systems at high doses, such as those in the cytotoxic range."
J Toxicol Environ Health. 1997 Apr 11; 50(5): 433-49.
DNA-protein cross-links produced by various chemicals in cultured human lymphoma cells.
Costa M, Zhitkovich A, Harris M, Paustenbach D, Gargas M. Institute of Environmental Medicine, New York University Medical Center, New York 10016, USA.
Chemicals such as cis-platinum, formaldehyde, chromate, copper, and certain arsenic compounds have been shown to produce DNA-protein cross-links in human in vitro cell systems at high doses, such as those in the cytotoxic range.
Thus far there have only been a limited number of other chemicals evaluated for their ability to produce cross-links.
The purpose of the work described here was to evaluate whether select industrial chemicals can form DNA-protein cross-links in human cells in vitro.
We evaluated acetaldehyde, acrolein, diepoxybutane, paraformaldehyde, 2-furaldehyde, propionaldehyde, chloroacetaldehyde, sodium arsenite, and a deodorant tablet [Mega Blue; hazardous component listed as tris (hydroxymethyl) nitromethane].
Short- and long-term cytotoxicity was evaluated and used to select appropriate doses for in vitro testing. DNA-protein cross-linking was evaluated at no fewer than three doses and two cell lysate washing temperatures (45 and 65 degrees C) in Epstein-Barr virus (EBV) human Burkitt's lymphoma cells.
The two washing temperatures were used to assess the heat stability of the DNA-protein cross-link, 2-Furaldehyde, acetaldehyde, and propionaldehyde produced statistically significant increases in DNA-protein cross-links at washing temperatures of 45 degrees C, but not 65 degrees C, and at or above concentrations of 5, 17.5, and 75 mM, respectively.
Acrolein, diepoxybutane, paraformaldehyde, and Mega Blue produced statistically significant increases in DNA-protein cross-links washed at 45 and 65 degrees C at or above concentrations of 0.15 mM, 12.5 mM, 0.003%, and 0.1%, respectively. Sodium arsenite and chloroacetaldehyde did not produce significantly increased DNA-protein cross-links at either temperature nor at any dose tested.
Excluding paraformaldehyde and 2-furaldehyde treatments, significant increases in DNA-protein cross-links were observed only at doses that resulted in complete cell death within 4 d following dosing. This work demonstrates that DNA-protein cross-links can be formed in vitro following exposure to a variety of industrial compounds and that most cross-links are formed at cytotoxic concentrations.
Short review: research on aspartame (methanol, formaldehyde, formic acid) toxicity
Research on aspartame (methanol, formaldehyde, formic acid) toxicity
Fully 11% of aspartame is methanol-- 1,120 mg aspartame in 2 L diet soda, almost six 12-oz cans, gives 123 mg methanol (wood alcohol). If 30% of the methanol is turned into formaldehyde, the amount of formaldehyde is 18 times the USA EPA limit for daily formaldehyde in drinking water, 2 mg in 2 L water.
Methanol (formaldehyde, formic acid) disposition: Bouchard M et al, full plain text, 2001: Substantial sources are degradation of fruit pectins, liquors, aspartame, smoke
"That substantial amounts of methanol metabolites or by-products are retained for a long time is verified by Horton et al. (1992) who estimated that 18 h following an iv injection of 100 mg/kg of 14C-methanol in male Fischer-344 rats, only 57% of the dose was eliminated from the body.
From the data of Dorman et al. (1994) and Medinsky et al. (1997), it can further be calculated that 48 h following the start of a 2-h inhalation exposure to 900 ppm of 14C-methanol vapors in female cynomolgus monkeys, only 23% of the absorbed 14C-methanol was eliminated from the body.
These findings are corroborated by the data of Heck et al. (1983) showing that 40% of a 14C-formaldehyde inhalation dose remained in the body 70 h postexposure."
"Exposure to methanol also results from the consumption of certain foodstuffs (fruits, fruit juices, certain vegetables, aspartame sweetener, roasted coffee, honey) and alcoholic beverages (Health Effects Institute, 1987; Jacobsen et al., 1988)."
"However, the severe toxic effects are usually associated with the production and accumulation of formic acid, which causes metabolic acidosis and visual impairment that can lead to blindness and death at blood concentrations of methanol above 31 mmol/l (Røe, 1982; Tephly and McMartin, 1984; U.S. DHHS, 1993).
Although the acute toxic effects of methanol in humans are well documented, little is known about the chronic effects of low exposure doses, which are of interest in view of the potential use of methanol as an engine fuel and current use as a solvent and chemical intermediate.
Gestational exposure studies in pregnant rodents (mice and rats) have also shown that high methanol inhalation exposures (5000 or 10,000 ppm and more, 7 h/day during days 6 or 7 to 15 of gestation) can induce birth defects (Bolon et al., 1993; IPCS, 1997; Nelson et al., 1985)."
"The corresponding average elimination half-life of absorbed methanol through metabolism to formaldehyde was estimated to be 1.3, 0.7-3.2, and 1.7 h."
"Inversely, in monkeys and in humans, a larger fraction of body burden of formaldehyde is rapidly transferred to a long-term component. The latter represents the formaldehyde that (directly or after oxidation to formate) binds to various endogenous molecules..."
"Animal studies have reported that systemic methanol is eliminated mainly by metabolism (70 to 97% of absorbed dose) and only a small fraction is eliminated as unchanged methanol in urine and in the expired air (< 3-4%) (Dorman et al., 1994; Horton et al., 1992).
Systemic methanol is extensively metabolized by liver alcohol dehydrogenase and catalase-peroxidase enzymes to formaldehyde, which is in turn rapidly oxidized to formic acid by formaldehyde dehydrogenase enzymes (Goodman and Tephly, 1968; Heck et al., 1983; Røe, 1982; Tephly and McMartin, 1984).
Under physiological conditions, formic acid dissociates to formate and hydrogen ions.
Current evidence indicates that, in rodents, methanol is converted mainly by the catalase-peroxidase system whereas monkeys and humans metabolize methanol mainly through the alcohol dehydrogenase system (Goodman and Tephly, 1968; Tephly and McMartin, 1984).
Formaldehyde, as it is highly reactive, forms relatively stable adducts with cellular constituents (Heck et al., 1983; Røe, 1982)."
"The whole body loads of methanol, formaldehyde, formate, and unobserved by-products of formaldehyde metabolism were followed.
Since methanol distributes quite evenly in the total body water, detailed compartmental representation of body tissue loads was not deemed necessary."
"According to model predictions, congruent with the data in the literature (Dorman et al., 1994; Horton et al., 1992), a certain fraction of formaldehyde is readily oxidized to formate, a major fraction of which is rapidly converted to CO2 and exhaled, whereas a small fraction is excreted as formic acid in urine.
However, fits to the available data in rats and monkeys of Horton et al. (1992) and Dorman et al. (1994) show that, once formed, a substantial fraction of formaldehyde is converted to unobserved forms.
This pathway contributes to a long-term unobserved compartment.
The latter, most plausibly, represents either the formaldehyde that (directly or after oxidation to formate) binds to various endogenous molecules (Heck et al., 1983; Røe, 1982) or is incorporated in the tetrahydrofolic-acid-dependent one-carbon pathway to become the building block of a number of synthetic pathways (Røe, 1982; Tephly and McMartin, 1984).
That substantial amounts of methanol metabolites or by-products are retained for a long time is verified by Horton et al. (1992) who estimated that 18 h following an iv injection of 100 mg/kg of 14C-methanol in male Fischer-344 rats, only 57% of the dose was eliminated from the body.
From the data of Dorman et al. (1994) and Medinsky et al. (1997), it can further be calculated that 48 h following the start of a 2-h inhalation exposure to 900 ppm of 14C-methanol vapors in female cynomolgus monkeys, only 23% of the absorbed 14C-methanol was eliminated from the body.Mp< These findings are corroborated by the data of Heck et al. (1983) showing that 40% of a 14C-formaldehyde inhalation dose remained in the body 70 h postexposure.
In the present study, the model proposed rests on acute exposure data, where the time profiles of methanol and its metabolites were determined only over short time periods (a maximum of 6 h of exposure and a maximum of 48 h postexposure).
This does not allow observation of the slow release from the long-term components.
It is to be noted that most of the published studies on the detailed disposition kinetics of methanol regard controlled short-term (iv injection or continuous inhalation exposure over a few hours) methanol exposures in rats, primates, and humans (Batterman et al., 1998; Damian and Raabe, 1996; Dorman et al., 1994; Ferry et al., 1980; Fisher et al., 2000; Franzblau et al., 1995; Horton et al., 1992; Jacobsen et al., 1988; Osterloh et al., 1996; Pollack et al., 1993; Sedivec et al., 1981; Ward et al., 1995; Ward and Pollack, 1996).
Experimental studies on the detailed time profiles following controlled repeated exposures to methanol are lacking."
"Thus, in monkeys and plausibly humans, a much larger fraction of body formaldehyde is rapidly converted to unobserved forms rather than passed on to formate and eventually CO2."
"However, the volume of distribution of formate was larger than that of methanol, which strongly suggests that formate distributes in body constituents other than water, such as proteins.
The closeness of our simulations to the available experimental data on the time course of formate blood concentrations is consistent with the volume of distribution concept (i.e., rapid exchanges between the nonblood pool of formate and blood formate)."
"Also, background concentrations of formate are subject to wide interindividual variations (Baumann and Angerer, 1979; D'Alessandro et al., 1994; Franzblau et al., 1995; Heinrich and Angerer, 1982; Lee et al., 1992; Osterloh et al., 1996; Sedivec et al., 1981)."
Toxicological Sciences 64, 169-184 (2001)
Copyright (c) 2001 by the Society of Toxicology
BIOTRANSFORMATION AND TOXICOKINETIC
A Biologically Based Dynamic Model for Predicting the Disposition of Methanol and Its Metabolites in Animals and Humans
Michèle Bouchard *, #,1, email@example.com
Robert C. Brunet, # firstname.lastname@example.org
Pierre-Olivier Droz, #
Gaétan Carrier* email@example.com
* Department of Environmental and Occupational Health, Faculty of Medicine, Université de Montréal, P.O. Box 6128, Main Station, Montréal, Québec, Canada, H3C 3J7
# Institut Universitaire romand de Santé au Travail, rue du Bugnon 19, CH-1005, Lausanne, Switzerland, and
# Département de Mathématiques et de Statistique and Centre de Recherches Mathématiques, Faculté des arts et des sciences, Université de Montréal, P.O. Box 6128, Main Station, Montréal, Québec, Canada, H3C 3J7