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the first to dig the mine get to share the gold -- 7 recent similar animal research studies indicating much lower aspartame and methanol ADI levels: Rich Murray 2013.12.23

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  • Rich Murray
    the first to dig the mine get to share the gold -- 7 recent similar animal research studies indicating much lower aspartame and methanol ADI levels: Rich
    Message 1 of 1 , Dec 23, 2013
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      the first to dig the mine get to share the gold -- 7 recent similar animal research studies indicating much lower aspartame and methanol ADI levels: Rich Murray 2013.12.23


      My efforts since 1999, when the public Net was four years old, have served to blow gently on the sail of science, helping catalyze the exponential expansion  of science in a becalmed, neglected area.

      In fall, 2007 a far away faint breeze unexpectedly rose in New Zealand, the WC Monte methanol ADH1 enzyme formaldehyde toxicity paradigm MMAEFTP.

      It became a mild wind in 2010 with publication of a concise introduction.

      Then, in isolated gust in January 2012 with his textbook and website.

      Now, in fits and starts, still hardly visible to most, about to emerge in 2014 as a gale of progress, a publicly acknowledged breakthrough paradigm inspiring focused research by a spontaneously emerging global network of collaborators -- quickly easily  cheaply and safely serving the huge benefit of all humans -- motivating confused corporations and corrupt bureaucracies towards earnest, willing, mature and positive public service -- exemplifying new modes for positive handling of a panoply of global challenges.

      So, today, one more puff at the sail... 
       
      Studies are urgently needed on MMAEFTP to elucidate the inevitably complex details in humans, the panoply of methanol sources, the actual results in 20 tissues with high ADH1 levels over time in members of vulnerable groups, the discarding of outdated older models of methanol toxicity, the spread of cheap convenient diagnostic tools, the value of antidotes like ethanol and folate.

      World citizens need rapid, up to date information for making crucial changes about methanol.

      Corporations may choose to substantially compensate victims, in the tradition of past opportunities like tobacco.


      UK COT chronic methanol toxicity in diet, 2011 July, free full text 22 pages -- earnest critical comments, applying the WC Monte methanol formaldehyde ADH1 enzyme paradigm: Rich Murray 2013.12.09


      free full text, 245 page EFSA draft report on methanol toxicity 2013.01.08

      line 5826 of the EFSA draft aspartame review 2013 January
      COT (UK Committee on toxicity of chemicals in food, consumer products and the environment),
      2011. COT Statement on the effects of chronic dietary exposure to methanol.
      Available at:


      263 page EFSA aspartame assent -- some critical notes -- also 214 public comments and Sept. 2013 US-EPA methanol review 212 page: Rich Murray 2013.12.15


      I selected information from several of my long posts, but found I understood what I was doing better if I stayed interested by including a lot of details -- EFSA may have only looked at and, of course, rejected only the two 2012 studies by Kate Collison, which were confirmed by their more sophisticated 2013 epigenetic study.


      [ one study ]

      [ 50.4 mg daily aspartame per rat ]

      histological and genetic harm of oral aspartame in female rats and pups, Azza A. M. Abd Elfatah, Inas S. Ghaly, Safaa M. Hanafy, Al-Azhar University, Cairo,
      Pak J Biol Sci. 2012.10.01,15 pages free full text, color microphotos: Rich Murray 2013.12.09


      [ 3 studies -- EFSA rejected the two 2012 studies, and didn't include the 2013 epigenetic study -- all three studies use same dose of about 50 mg/kg bw aspartame, and confirm harm in mice. ]

      Kate S. Collison et al show prediabetic harm in gene expression in mice fed lifetime aspartame, MSG, trans fats -- reduce human aspartame ADI 1000 times: Rich Murray 2013.07.30



      [ 4 studies -- which confirm harm in rats at 75 mg/kg bw aspartame.

      So, that gives 7 recent rodent studies that show harm for 50 to 75 mg/kg bw aspartame, which, conservatively divided by 100, gives a human aspartame ADI of 0.5 to 0.75 mg/kg bw, 35 to 52.5 mg for a 70 kg human, about 2 to 3 oz aspartame drink...  ]

      Unpublished birth defect studies in animals by WC Monte and two aspartame industry labs found some birth defects from 220 and 300 mg/kg bw methanol. ]

      aspartame harm in rat brain from 75 mg/kg gives human ADI 0.75 mg/kg, 53 times less than EU ADI 40 mg/kg, Ashok Iyyaswamy, SheelaDevi Rathinasamy, U. Madras 2012.08.03 free full text -- main methanol toxin is formaldehyde, not formate: Rich Murray 2013.06.01



      [ All the studies from India use high levels of aspartame 500 to 1,000, or methanol 2,370 to 3,000 mg/kg bw. ]

      more lower aspartame and methanol ADIs from studies by RH Nair, SheelaDevi Rathinasamy, WC Monte, PS Jeganathan, A Namasivayam, Hazleton Labs, Searle Labs: Rich Murray 2013.06.01


      James McDonald to EFSA, outdated aspartame ADI gives methanol 35 times too high for human safety, ten minute talk at April 9 public sharing, Brussels: Rich Murray 2013.04.15


      California OEHHA sets methanol ingestion level 23 mg daily, same as from 1 can aspartame diet soda, 10 cigarettes, 3 tomatoes, or 4 cans green beans: Rich Murray 2013.07.03

      "However, the anticipated exposure to methanol from consumption of aspartame would not be considered an exposure within the meaning of Proposition 65 because aspartame is not listed under Proposition 65."

      [ Rich Murray: Many pregnant women drink one 12-oz can aspartame diet drink daily, with 200 mg aspartame that gives 11% methanol, 22 mg, which is just under the OEHHA limit of 23 mg daily.

      The smoke from 10 cigarettes gives 20 mg methanol, the same as from 1 can aspartame drink, 3 full size fresh tomatoes, or 4 cans of unfresh green beans. ]


      smoke from pack cigarettes gives 40 mg methanol for 20 gr tobacco, 6 tobacco methanol papers, Carl Neuberg 1926-1939, Berlin -- so methanol formaldehyde toxicity paradigm is co-factor in 18 tobacco diseases -- WC Monte gives 23 references: Rich Murray 2013.03.29


      [ 4 studies ]

      aspartame harm in rat brain from 75 mg/kg gives human ADI 0.75 mg/kg, 53 times less than EU ADI 40 mg/kg, Ashok Iyyaswamy, SheelaDevi Rathinasamy, U. Madras 2012.08.03 free full text -- main methanol toxin is formaldehyde, not formate: Rich Murray 2013.06.01


      [ not in 263 page EFSA 2013.12.10 aspartame review ]

      J Biosci. 2012 Sep;37(4):679-88.
      Effect of chronic exposure to aspartame on oxidative stress in the brain of albino rats.
      Iyyaswamy A, Rathinasamy S.
      Department of Physiology, University of Madras, 
      Sekkizhar campus, Taramani, Chennai 600 113, India.

      [ The Ashok Iyyaswamy and SheelaDevi Rathinasamy, 2012 September, study used 75 mg/kg aspartame, which gave 8.25 mg/kg methanol, finding harm in distinct brain regions:

      "To mimic the human methanol metabolism, methotrexate (MTX)-treated rats [ 0.2 mg/kg/day ] were included to study the aspartame effects.

      Wistar strain male albino rats were administered with aspartame orally and studied along with controls and MTX-treated controls."

      Methotrexate (MTX) was also used in their similar 2006 study to induce folate deficiency, making rats much more vulnerable to methanol toxicity, similar to the uniquely high vulnerability of humans -- so this showed that the other two breakdown products of aspartame, phenyalanine (50 %) and aspartic acid (39 %) are far less toxic in the rat brain -- a key finding, in light of the fact that methanol circulates with a half-life of 3 hours in the human bloodstream to all parts of the body and fetus every minute, to be made quickly by ADH1 enzyme into free floating formaldehyde right inside the cells of 20 tissues, including the inner linings of brain blood vessels, the rods and cones of the retina, and the Purkinje cells in the vermis of the cerebellum. It fits the picture that headaches, including migraines, are the first and most common symptoms for many methanol diseases. ]

      "Heavy users of aspartame-containing products consume as much as 250 mg of methanol daily, or 32 times the EPA limit."
      [ This would be supplied by a dozen cans of aspartame diet drink... ]

      "From the foregoing it is clear that the methanol, which is a by-product of aspartame, may be responsible for the alteration observed in the free-radical-scavenging system. 

      Since methanol is freely permeable through membranes and lipids, it also gets distributed in the brain tissues and may cause the damage.

      Increased production of free radicals and increased oxidative damage to proteins in distinct brain regions, retina and optic nerve after methanol administration (Rajamani et al. 2006) was also reported earlier lending support to the present findings. 

      The scientific reports on aspartame also revealed that aspartame consumption affects the brain. 

      The aspartame intake has been reported to be responsible for neurological and behavioural disturbances in sensitive individuals (Johns 1986).

      Moreover, Mourad and Noor (2011)  have observed that the daily ingestion of aspartame for 4 weeks induces significant increase in the LPO levels in the cerebral cortex, which is accompanied by significant decrease in GSH content and a significant increase in SOD activity, which is also in agreement with this study. 

      According to them, the decrease in GSH content and increase in SOD activity persisted until after 6 weeks of aspartame treatment.

      Moreover, they add that aspartame-induced oxidative stress may depend on the duration of aspartame administration even within the acceptable daily intake dose. 

      The present study reveals that aspartame administration in the body system persists for longer duration, which indicates the possible accumulation of methanol and its metabolite. 

      Since it is consumed more by diabetic people whose metabolism is already altered, it is essential to do more work on these lines and create awareness regarding the usage of this artificial sweetener.

      5. Conclusion

      After chronic exposure of aspartame, detectable methanol continues to circulate in the blood; methanol per se and its metabolites may be responsible for the generation of oxidative stress in brain regions.

      This study confirms the presence of toxic metabolite after aspartame administration and it emphasizes the need to caution the people who are using aspartame routinely."


      Abstract

      This study was aimed at investigating the chronic effect of the artificial sweetener aspartame on oxidative stress in brain regions of Wistar strain albino rats. 

      Many controversial reports are available on the use of aspartame as it releases methanol as one of its metabolite during metabolism. 

      The present study proposed to investigate whether chronic aspartame (75 mg/kg) administration could release methanol and induce oxidative stress in the rat brain.
      To mimic the human methanol metabolism, methotrexate (MTX)-treated rats [ 0.2 mg/kg/day ]  were included to study the aspartame effects.

      Wistar strain male albino rats were administered with aspartame orally and studied along with controls and MTX-treated controls.

      The blood methanol level was estimated, the animal was sacrificed and the free radical changes were observed in brain discrete regions by assessing the scavenging enzymes, reduced glutathione, lipid peroxidation (LPO) and protein thiol levels.

      It was observed that there was a significant increase in LPO levels, superoxide dismutase (SOD) activity, GPx levels and CAT activity with a significant decrease in GSH and protein thiol.

      Moreover, the increases in some of these enzymes were region specific.

      Chronic exposure of aspartame resulted in detectable methanol in blood.

      Methanol per se and its metabolites may be responsible for the generation of oxidative stress in brain regions.

      PMID: 22922192   Free full text


      pages 679-688

      Effect of chronic exposure to aspartame on oxidative stress in the brain of albino rats.
      ASHOK IYYASWAMY and SHEELADEVI RATHINASAMY*
      Ashok Iyyaswamy and SheelaDevi Rathinasamy <drsheeladevi@...>,
      Department of Physiology, Dr. ALM PG Institute of Basic Medical Sciences,
      University of Madras, Sekkizhar campus,
      Taramani, Chennai 600 113, India
      *Corresponding author (Fax, +91-44-24540709;


      [Iyyaswamy A and Rathinasamy S   2012 
      Effect of chronic exposure to aspartame on oxidative stress in the brain of albino rats. 
      J. Biosci. 37 679–688] 
      DOI 10.1007/s12038-012-9236-0

      J. Biosci. 37(4), September 2012, 679–688,
      * Indian Academy of Sciences
      Keywords. Aspartame; blood methanol; oxidative stress; rat folate-deficient model; free radical
      Published online: 3 August 2012

      MS received 18 April 2012; accepted 06 June 2012
      Corresponding editor: INDRANEEL MITTRA




      [  not in 263 page EFSA 2013.12.10 aspartame review ]

      [  Iman M. Mourad gives two studies in 2011, showing harm in albino rats from 40 mg/kg aspartame ( which gives 11 % = 4.4 mg/kg methanol ). ] 

      Mourad, I. M., and N. A. Noor,
      "Aspartame (A Widely used Artificial Sweetener) and Oxidative Stress In the Rat Cerebral Cortex",
      International Journal of Pharmacy and Biomedical Sciences, 2011.
      Int J Pharm Biomed Sci 2011, 2(1), 4-10
      ISSN No: 0976-5263

      Zoology Department, Faculty of Science,
      Cairo University, Cairo, Giza 12613,
      Egypt
      *Correspondence:
      Dr. Neveen A. Noor
      Tel: 00202-3567 6708
      Fax: 00202-3760 3735

      Abstract  CU-PDF.pdf  free full text 7 pages 70 references



      [  not in 263 page EFSA 2013.12.10 aspartame review ]

      [  aspartame 40 mg/kg gives methanol 4.4 mg/kg ]

      Mourad, I. M.,
      "Effect of Aspartame on some Oxidative Stress parameters in Liver and Kidney of Rats",
      African Journal of Pharmacy and Pharmacology, vol. 5, issue 6, pp. 678-682, 2011.

      African Journal of Pharmacy and Pharmacology Vol. 5(6), pp. 678-682, June 2011
      ISSN 1996-0816 ©2011 Academic Journals
      Full Length Research Paper
      Effect of aspartame on some oxidative stress parameters in liver and kidney of rats
      Iman, M. Mourad
      Department of Zoology, Faculty of Science, Cairo University, Egypt.

      Abstract CU-PDF.pdf  free full text 5 pages




      [  not in 263 page EFSA 2013.12.10 aspartame review ]

      Parthasarathy JN, Ramasundaram SK, Sundaramahalingam M,
      Pathinasamy SD, Devi RS (2006).
      Methanol induced oxidative stress in rat lymphoid organs.
      J. Occup. Health, 48: 20-27.

      8 pages free full text [ 2.37 g/kg = 287 times more than 8.25 mg/kg ]
      183.  Parthasarathy N, Kumar R, Manikandan S, Devi R.
      Methanol-Induced Oxidative Stress in Rat Lymphoid Organs.
      J Occup Health 2006;48:20-7. 





      [ All the studies from India use high levels of aspartame 500 to 1,000 or methanol, 2,370 to 3,000 mg/kg bw. ]

      more lower aspartame and methanol ADIs from studies by RH Nair, SheelaDevi Rathinasamy, WC Monte,  PS Jeganathan, A Namasivayam, Hazleton Labs, Searle Labs: Rich Murray 2013.06.01



      Drug Chem Toxicol. 2012 Mar 2. [Epub ahead of print]
      Long-term consumption of aspartame and brain antioxidant defense status.
      Abhilash M,
      Sauganth Paul MV,
      Varghese MV,
      Nair RH.        R. Harikumaran Nair <harinair@...>,
      School of Biosciences, Mahatma Gandhi University , Kottayam , India.

      Abstract

      The present study investigated the effect of long-term intake of aspartame, a widely used artificial sweetener, on antioxidant defense status in the rat brain.

      Male Wistar rats weighing 150-175 g were randomly divided into three groups as follows:
      The first group was given aspartame at a dose of 500 mg/kg body weight (b.w.);
      the second group was given aspartame at dose of 1,000 mg/kg b.w., respectively,
      in a total volume of 3 mL of water; and
      the control rats received 3 mL of distilled water.
      Oral intubations were done in the morning, daily for 180 days.

      The concentration of reduced glutathione (GSH) and the activity of glutathione reductase (GR) were significantly reduced in the brain of rats that had received the dose of 1,000 mg/kg b.w. of aspartame,
      whereas only a significant reduction in GSH concentration was observed
      in the 500-mg/kg b.w. aspartame-treated group.

      Histopathological examination revealed mild vascular congestion in the
      1,000 mg/kg b.w. group of aspartame-treated rats.

      [ The visible brain harm, " mild vascular congestion" in the 1,000 mg/kg aspartame, 110 mg/kg methanol, is similar to that found by WC Monte and students from 950 mg/kg methanol, a dose 9 times larger.

      Divided by 1,000 gives a human aspartame  ADI of 1 mg/kg body weight = 70 mg for a 70 kg person, the dose from  4 ounces of a 12-oz can diet drink with 200 mg aspartame. ]

      The results of this experiment indicate that long-term consumption of aspartame leads to an imbalance in the antioxidant/pro-oxidant status in the brain, mainly through the mechanism involving the glutathione-dependent system.
      PMID: 22385158





      Aspartame water in rats for 6 months causes liver harm, RH Nair et al, Mahatma Gandhi U, Food Chem Toxicol 2011.03.02: Rich Murray 2011.03.12

      "Histopathological examination revealed leukocyte infiltration in
      aspartame-treated rats (1000 mg/kg.b.wt)."


      Food Chem Toxicol. 2011 Mar 2. [Epub ahead of print]
      Effect of long term intake of aspartame on antioxidant defense status in liver.
      Abhilash M, Paul MV, Varghese MV, Nair RH.
      School of Biosciences, Mahatma Gandhi University,
      Kottayam, Kerala, India, 686560.

      Abstract

      The present study evaluates the effect of long term intake of aspartame, the artificial sweetener, on liver antioxidant system and hepatocellular injury in animal model.
      Eighteen adult male Wistar rats, weighing 150 - 175 g, were randomly divided into three groups as follows:
      first group was given aspartame dissolved in water in a dose of 500 mg/kg.b.wt;
      the second group was given a dose of 1000 mg/kg.b.wt;
      and controls were given water freely.
      Rats that had received aspartame (1000 mg/kg.b.wt) in the drinking water for 180 days showed a significant increase in activities of
      alanine aminotransferase (ALT),
      aspartate aminotransferase (AST),
      alkaline phosphatase (ALP) and
      y-glutamyl transferase (GGT).
      The concentration of reduced glutathione (GSH)
      and the activity of glutathione peroxidase (GPx), and
      glutathione reductase (GR)
      were significantly reduced in the liver of rats that had received aspartame (1000 mg/kg. b.wt).
      Glutathione was significantly decreased in both the experimental groups.

      Histopathological examination revealed leukocyte infiltration in
      aspartame-treated rats (1000 mg/kg.b.wt).

      It can be concluded from these observations that long term consumption of aspartame leads to hepatocellular injury and alterations in liver antioxidant status mainly through glutathione dependent system.
      Copyright 2011. Published by Elsevier Ltd. PMID: 21376768

      [ Again, divided by 1,000 gives a human aspartame  ADI of 1 mg/kg body weight = 70 mg for a 70 kg person, the dose from  4 ounces of a 12-oz can diet drink with 200 mg aspartame. ]

      Souganth Paul, MV - Research Fellow

      Abhilash, M - Research Fellow

      Varghese, Mathews V - Research Fellow



      School of Bio-Sciences

      Teacher Profiles

      Name: Dr. R. Harikumaran Nair [ R. Harikumaran Nair
      Designation: Assistant Professor
      Address: Room No. 5
      School of Biosciences,
      Mahatma Gandhi University
      Priyadarshini Hills. P O. , Kottayam-686 560
      Kerala, India
      Mobile Phone: +91-94472 60362

      Research Interest

      In our laboratory, we study the effect of food additives such as monosodium glutamate,
      aspartame, drug component arsenic trioxide and other toxins like organic solvents on different physiological mechanisms in rats and tissue culture model systems.
      Another area of interest is environmental and occupational stress.

      Academic Profile
      Ph. D-Physiology (2001), School of Biosciences, Mahatma Gandhi University, Kottayam, Kerala, India
      M. Sc- Zoology (1994) University of Kerala, Thiruvananthapuram, Kerala, India,
      B. Sc-Zoology (1992) University of Kerala, Thiruvananthapuram, Kerala, India,

      Professional Experience:

      *Lecturer in Physiology (Permanent service) - School of Biosciences,
      Mahatma Gandhi University, Kerala, India. 22nd May 2006 to present
      *Lecturer in Physiology (contract service) - School of Biosciences,
      Mahatma Gandhi University, Kerala, India.
      20th November 2003 to 21st May 2006
      *Postdoctoral Research fellow- Dept. of Physiology, St. John’s Medical College,
      Bangalore, India. 01st March 2002 to 15th July 2002
      *Research Fellow – School of Biosciences, Mahatma Gandhi University,
      Kottayam, Kerala, India. October 1995- April 2001




      free full text, 8 pages 49 references [ 2.37 g/kg methanol ]

      J Occup Health 2006; 48: 20-27
      Journal of Occupational Health
      Methanol-Induced Oxidative Stress in Rat Lymphoid Organs
      Narayanaperumal J. PARTHASARATHY,
      Ramasundaram S. KUMAR, 
      Sundaramahalingam MANIKANDAN
      and Rathinasamy S. DEVI
      Immunology Laboratory, Department of Physiology, Dr. ALM. PG.
      Institute of Basic Medical Sciences, University of Madras, 
      Taramani Campus, Chennai, Tamilnadu, India

      Received Apr 25, 2005; Accepted Oct 12, 2005

      Correspondence to: R.S. Devi, Immunology Laboratory,
      Department of Physiology, Dr. AIM. PG. Institute of Basic Medical Sciences,
      University of Madras, Taramani Campus, Chennai,
      Tamilnadu 600 113, India (e-mail: drsheeladevi@... )

      (J Occup Health 2006; 48: 20-27)
      Key words: Methanol, Antioxidants, Lipid peroxidation, Lymphoid organs

      Abstract: 

      Methanol-Induced Oxidative Stress In Rat Lymphoid Organs: 
      Narayanaperumal J. PARTHASARATHY, et al.
      Immunology Laboratory, Department of Physiology,
      Dr. ALM. PG. Institute of Basic Medical Sciences,
      University of Madras, Taramani Campus, India

      Methanol is primarily metabolized by oxidation to formaldehyde and then to formate. 

      These processes are accompanied by formation of superoxide anion and hydrogen peroxide.

      This paper reports data on the effect of methanol on antioxidant status and lipid peroxidation in lymphoid organs such as the spleen, thymus, lymph nodes and bone marrow of rats. 

      Male Wistar albino rats were intoxicated with methanol (2.37 g/kg b.w intraperitoneally) for detecting toxicity levels for one day, 15 d and 30 d, respectively. 

      Administration of methanol at 15 and 30 d significantly (p 0.05) increased lipid peroxidation and decreased the enzymatic (superoxide dismutase, catalase, glutathione peroxidase) and non-enzymatic antioxidants (reduced glutathione and vitamin C) in lymphoid organs.

      However, lipid peroxidation and enzymatic and nonenzymatic antioxidants in the acute methanol exposed group animals were found to be significantly (p 0.05) increased.

      In one day methanol intoxication, the levels of free radicals initially increased, and to remove these free radicals, antioxidants levels were elevated, which generally prevented oxidative cell damage.

      But in longer periods of intoxication, when the generation of reactive free radicals overwhelmed the antioxidan defense, lipid peroxidation increased. 

      Further, decreased antioxidants in 15 and 30 d methanol intoxication may have been due to overutilization of non-enzymatic and enzymatic antioxidants to scavenge the products of lipid peroxidation.

      In addition, the liver and kidney markers of serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), urea and creatinine significantly increased.

      This study concludes that exposure to methanol causes oxidative stress by altering the oxidant/antioxidant balance in lymphoid organs of the rat.

      [ 29) NJ Parthasarathy, R Srikumar and RS Devi:
      Effect of methanol intoxication on rat neutrophil functions.
      J Immunotoxicol 2, 115-121 (2005)


      37) ME Paula, DC Mathangi and A Namasivayam:
      Free radical changes in methanol toxicity.
      Indian J Physiol Pharmacol 47, 207-211 (2003)  ]



      [ 29) NJ Parthasarathy, R Sri Kumar and RS Devi:
      Effect of methanol intoxication on rat neutrophil functions.
      J Immunotoxicol 2, 115-121 (2005) [ 2.37 g/kg b.wt., i.p. ] 

      J Immunotoxicol. 2005 Apr 1;2(2):115-21.
      doi: 10.1080/15476910500187425.
      Effect of methanol intoxication on rat neutrophil functions.
      Parthasarathy NJ, Kumar RS, Devi RS.
      N. Jeya Parthasarathy, R. Sri Kumar and R. Sheela Devi.
      Immunology Laboratory, Department of Physiology, 
      Dr. ALM.PG., Institute of Basic Medical Sciences,
      University of Madras, Taramani Campus,
      Chennai, Tamil Nadu, India.

      Abstract

      Methanol (MeOH) toxicity, a potential problem from accidental, intentional, as well as occupational and daily ingestion of the agent, receives attention only after severe signs of intoxication have set in or death is imminent.
       
      While accidental and intentional exposures involve high doses, the occupational and ingestion forms more often reflect small daily intakes.
       
      Still, even at these low levels, little is known about the potential immunotoxic implications from these recurring exposures. 

      As innate immunity confers a first-line of defense against infection, a study was designed to examine the effects of daily exposure to MeOH
      (at (1)/(4) LD(50) level, [ 2.37 g/kg b.wt., i.p. ] for up to 15 or 30 days)
      on neutrophil (PMN) functions using rats that were (or were not)
      injected with sheep red blood cells (SRBC) during the course of exposures.

      Blood samples were analyzed for total (TLC) and differential leucocyte counts (DLC), and isolated neutrophils (PMN) were assessed for changes in function
      by monitoring phagocytic (PI) and avidity indices (AI), nitroblue tetrazolium (NBT) reduction, and adherence.

      Body weights were monitored during exposures and weights of major immune system organs (i.e., spleen, thymus, lymph nodes) were assessed at sacrifice.

      Body and organ weight, TLC, blood PMN levels, PMN PI, and adherence were all significantly decreased in SRBC-untreated rats that received MeOH, although these cells did also display significant increases in AI and NBT reduction.

      With SRBC-treated rats, though the percentage of PMN in the blood increased with ongoing MeOH exposure, all the other parameters were markedly decreased in comparison to their controls. 

      Thus, this study showed that repeated exposures to MeOH modulates PMN functions, thereby potentially altering the first line of defense in a normal immune response in exposed hosts.
      PMID: 18958665 ]



      Effect of methanol-induced oxidative stress on the neuroimmune system of experimental rats.
      Parthasarathy NJ, Kumar RS, Manikandan S, Narayanan GS, Kumar RV, Devi RS.
      Chem Biol Interact. 2006 May 15;161(1):14-25.
      Epub 2006 Mar 6.
      PMID: 16564515   [ 2.37 g/kg b.wt., i.p. ]

      Chem Biol Interact. 2006 May 15;161(1):14-25. Epub 2006 Mar 6.
      Effect of methanol-induced oxidative stress on the neuroimmune system of experimental rats.
      Parthasarathy NJ, Kumar RS, Manikandan S, Narayanan GS, Kumar RV, Devi RS.
      Immunology Laboratory, Department of Physiology,
      Dr. ALM PG Institute of Basic Medical Sciences, 
      University of Madras, Taramani Campus,
      Chennai 600113, India.

      Abstract

      It is well known that the nervous system has increased susceptibility to methanol intoxication.

      The present study reveals the effect of methanol intoxication on antioxidant status, lipid peroxidation and DNA integrity in hypothalamic-pituitary-adrenal (HPA) axis organs and spleen. 

      Non-specific and specific immune functions were analyzed. In addition, open field behavior, plasma corticosterone level and blood methanol level were estimated.
       
      Male Wistar albino rats were intoxicated with methanol (2.37 g/kg b.wt., i.p.) for 1 day, 15 and 30 days.

      Administration of methanol showed significant increase in enzymatic (superoxide dismutase, catalase, glutathione peroxidase), non-enzymatic (reduced glutathione and Vitamin C) antioxidants and lipid peroxidation (LPO) in hypothalamus and adrenal gland of day 1 group.

      However, decrease in enzymatic and non-enzymatic antioxidants with concomitant increase in LPO level were observed in 15 and 30 days groups.

      Plasma corticosterone level was significantly increased in day 1 and 15 days groups
      whereas, 30 days methanol intoxication group showed considerable decrease in corticosterone level compared with control animals.

      Cell-mediated immune response of footpad thickness was significantly decreased with an increased leukocyte migration inhibition.

      Humoral immune response of antibody titers was elevated in methanol-intoxicated groups.

      Neutrophil functions, adherence and phagocytic index (PI) were found to be significantly decreased.

      Furthermore, significant increase in the avidity index and nitro blue tetrozolium reduction was observed in the methanol exposed animals.

      Day 1 methanol exposed group showed increased PI compared to the control ones.

      Methanol exposure for 30 days showed an increased DNA fragmentation in the hypothalamus, adrenal glands, and spleen.

      In conclusion, exposure to methanol-induced oxidative stress disturbs the HPA-axis function altering the level of corticosterone, which lead to varied non-specific and specific immune response in experimental rats.
      PMID: 16564515  



      37) ME Paula, DC Mathangi and A Namasivayam:
      Free radical changes in methanol toxicity.
      Indian J Physiol Pharmacol 47, 207-211 (2003)

      Indian J Physiol Pharmacol. 2003 Apr;47(2):207-11.
      Free radical changes in methanol toxicity.
      Paula EM, Mathangi DC, Namasivayam A.
      Department of Physiology,
      Dr. AML Post Graduate Institute of Basic Medical Sciences,
      University of Madras, Taramani, Chennai.

      Abstract

      Role of free radicals in methanol toxicity was evaluated in methanol treated albino rats.
      Methanol intoxication increased lipid peroxidation and depleted the free radical scavenging enzyme systems.

      The free radical quenching effect of vitamin E protected the animals from methanol induced free radical damage.

      PMID: 15255626    ]




      Their similar brain study 7 years ago used 364 times more daily methanol than their 2012 study with 75 mg/kg aspartame, which gave 8.25 mg/kg methanol:

      "We report on MeOH induced free radical changes and oxidative damages to proteins in the discrete regions of rat brain, retina and optic nerve.

      The present study used rats administered with methotrexate (MTX) to induce folate deficiency. 

      Three groups of animals, namely saline control, MTX control, MTX-MeOH group were tested."


      Toxicol Lett. 2006 Sep 10;165(3):265-73.  9 pages [ 3.0 g/kg ]
      Oxidative stress induced by methotrexate alone and in the presence of methanol in discrete regions of the rodent brain, retina and optic nerve.
      Rajamani R, Muthuvel A, Senthilvelan M, Sheeladevi R.
      Department of Physiology, Dr. ALM PG Institute of Basic Medical Sciences, University of Madras, Taramani Campus, Taramani, Chennai, India.

      Abstract

      The vulnerability of the nervous system due to methanol (MeOH) intoxication is a well known fact and reports on the production of free radicals due to MeOH exposure and their involvement in excitotoxicity and neuronal apoptosis are being increasingly reported.

      We report on MeOH induced free radical changes and oxidative damages to proteins in the discrete regions of rat brain, retina and optic nerve.

      The present study used rats administered with methotrexate (MTX) to induce folate deficiency. 

      Three groups of animals, namely saline control, MTX control, MTX-MeOH group were tested.

      The rats were injected intraperitoneally with MeOH (3 g/kg).
      [ minimum lethal dose for rats is 9.0 g/kg ]

      After 24 h of MeOH administration, the levels of free radical scavengers, superoxide dismutase, catalase, glutathione peroxidase and reduced glutathione levels were estimated in the six discrete regions of brain (cerebral cortex, cerebellum, midbrain, pons medulla, hippocampus and hypothalamus), retina and optic nerve specimens. 

      The levels of protein thiol, protein carbonyl and lipid peroxidation were also estimated and the expression of HSP70 in the hippocampus was analyzed by Western blot. 

      Marked reduction in the levels of glutathione in the MTX-MeOH group in relation to MTX control was observed and found to be increased in MTX control in relation to saline control.

      Increased protein carbonyls and decreased protein thiols were documented in all the specimens tested. 

      In addition, marked expression of HSP70 was observed in the hippocampus.

      The present investigation suggest that MeOH exposure results in increased generation of free radicals and significant protein oxidative damage and attempts to study the underlying mechanisms involved might reveal more insights to our existing knowledge on MeOH intoxication and related areas.

      PMID: 16766145  ]



       
      [ Prof. Woodrow C. Monte did expert unpublished research with his graduate students about 1989, using daily 0.95 g/kg oral methanol for 18-39 days each to prove brain damage in rats -- this dose served to overwhelm the rat biochemical defenses against this specific type of harm from methanol and its main toxic product formaldehyde.

      This methanol dose, producing visible brain damage within weeks in rats, would be given by 8,550 mg/kg daily aspartame, which, divided by the safety factor of 1,000, gives a human aspartame ADI of 8.6 mg/kg body weight, which for a 70-kg person is 600 mg aspartame, as much as from 3 12-oz cans diet drink at 200 mg aspartame each. 

      So, the many rat studies that find harm at 1-3 g/kg methanol imply human ADI levels from 8.6 to 25.8 mg/kg, 3 to 9 cans diet drink, below the existing official EU level of 40.0 mg/kg body weight.

      Thus, the doses used in many recent rat studies with 1 to 3 g/kg daily methanol doses are indeed likely to show many kinds of biochemical distress, along with visible damage in many small regions of many tissues with high levels of ADH1 enzyme, especially in highly vulnerable humans.

      The hazards are obvious for those people who use the highest level of aspartame and other methanol sources like cigarettes, and fruits juices vegetables cut up and preserved wet at room temperature in sealed cans jars plastic containers.

      Methyl alcohol ingestion as a model etiologic agent in multiple
      sclerosis, WC Monte, D Glanzman, C Johnston; Methanol induced
      neuropathology in the mammalian central nervous system, Woodrow C.
      Monte, Renee Ann Zeising, both reports 1989.12.04: Murray 2007.12.28
      2012.05.01
      posted again Tuesday, May 1, 2012

      Friday, December 28 2007 


      "METHANOL INDUCED NEUROPATHOLOGY IN THE MAMMALIAN CENTRAL NERVOUS SYSTEM
      Woodrow C. Monte Ph.D
      Renee Ann Zeising
      Department of Family Resources and Human Development
      Arizona State University, Tempe, AZ 85287 (U.S.A.)
      Key words: Methanol--Degeneration--Axon--Rat--
      Brain--Central Nervous System--Neuropathology
      Please address correspondence to:
      Woodrow C. Monte
      Department of Family Resources and Human Development
      Arizona State University, Tempe, Az. 85287


      SUMMARY

      The neurotoxic effect of methyl alcohol (methanol) was visualized in the rat central nervous system using reduced silver staining techniques.

      Following chronic administration of methanol (intubation with 0.95 gm/kg for 18, 25 or 33 days) all experimental animals showed massive axonal degeneration in multiple regions of brain, regardless of the duration of exposure.

      Histological processing yielded degeneration by-products of fibers with cells of origin lying in cerebellar cortex, deep cerebellar nuclei, cranial nerve nuclei and the red nucleus.

      Additional regions of axonal degeneration were found in the hippocampus, the flocculus, dorsal raphe nucleus, ventral cochlear nuclei, retrosplenium, the internal capsule of the corpus striatum and the optic chiasm.

      These results show that by using sufficiently sensitive neurohistological techniques, the neurotoxicity of methyl alcohol is revealed in the vertebrate central nervous system."




      "A ten-fold lower dose of 0.3 g/kg body weight was used with 272 fetuses in 21 litters, producing a much lower number of birth defects, compared to 296 fetuses in 22 control litters with no methanol."


      177. Hoque M., Monte WC., Black L., Johnston CS.
      Methanol Neuropathy and Teratology A Histological Study on Long-Evans Rats.
      FASEB 1988;25(2(6)):A513.

      many strong birth defects in 336 rat fetuses, 3 g/kg bw oral methanol (not toxic dose) in 26 rat litters, thesis done 1987, WC Monte, M Hoque, L Black, CS Johnson: Rich Murray 2013.04.13


      With years of expertise in testing toxins on rats, Prof. Woodrow C. Monte, Arizona State University, had his own lab, where he mentored graduate students.

      Vivid personal reports led him to test the possibility of birth defects from methanol, 11% of aspartame, 22 mg in the 200 mg aspartame in a 12-ounce can diet drink.

      He was unaware that the FDA and Searle Corp. had hidden 6 of their studies that showed birth defects from aspartame in rabbits and mice in 1975.

      This dose was planned to swamp the catalase enzyme defense system, which, effective in rats and all animals, except the uniquely vulnerable human species, protects against harm from methanol being made into free floating  formaldehyde inside cells with high levels of ADH1 enzyme, notably the inner walls of brain blood vessels and in retina rods and cones.

      As the smallest organic molecule, methanol readily enters all cells, while it is carried to all parts of the body and the fetus with the bloodstream -- in humans with a blood half-life of 3 hours, circulating through the whole body every minute.

      A ten-fold lower dose of 0.3 g/kg body weight was used with 272 fetuses in 21 litters, producing a much lower number of birth defects, compared to 296 fetuses in 22 control litters with no methanol.


      gives all photos and figures in "While Science Sleeps" textbook --
      Figure 12.5 shows a damaged rat pup brain...


      177. Hoque M., Monte WC., Black L., Johnston CS.
      Methanol Neuropathy and Teratology A Histological Study on Long-Evans Rats.
      FASEB 1988;25(2(6)):A513.


      [ See also:

      WC Monte finally got secret FDA memo 37 years after Searle Co. labs found birth defects in rabbits from aspartame (methanol, becomes formaldehyde via ADH1 enzyme within human cells) and its phenylalanine: Rich Murray 2012.06.02



      177.  Hoque M., Monte WC., Black L., Johnston CS.
      Methanol Neuropathy and Teratology A Histological Study on Long-Evans Rats.
      FASEB 1988;25(2(6)):A513.

      [ oral methanol 1.28 mg/kg in several rat experiments in 1988 by WC Monte and students with rats found severe birth defects, as well as brain damage in adult rats. ]

      [ Rich Murray: I organized the items in this reference, and corrected minor typos. ]


      TUESDAY, DECEMBER 8, 1987 deadline

      1988. FASEB ABSTRACT FORM

      Ms. Mahmuda Hoque
      1140 East Orange #116
      Tempe, AZ 85281
      Phone: Home/Holiday (602) 968-0400

      METHANOL NEUROPATHY: A HISTOLOGICAL STUDY ON LONG-EVANS RATS.
      [ Experiment I, II ]
      M. Hogue, W.C. Monte, and C.S. Johnston.
      Food and Nutrition laboratory, Arizona State University. Tempe AZ 85287


      Abstract

      Methyl alcohol (methanol), a highly toxic substance, is in food products containing the artificial sweetener, aspartame.

      Since aspartame is a very popular artificial sweetener, a large segment of our population is currently consuming chronic, low doses of methanol. 

      This study examined the neurotoxic effect of methanol on the central nervous system of rat pups.

      Methanol was administered daily by the gavage technique to 20 female Long-Evans rats (1.28 g/kg body weight) beginning two weeks prior to mating and throughout gestation and lactation.

      A control group of ten female rats received same volume of distilled water.

      Prenatal methanol exposure induced brain defects: hydrocephalus, eye defects, spina bifida occulta, and stillborn pups.

      All experimental, pups showed: massive axonal degeneration in multiple regions of brain.

      Histological examination of the brain, using reduced silver staining techniques, revealed massive fiber degeneration of the cerebellar cortex, deep cerebellar nuclei, and cranial nerve nuclei. 

      Additional regions of axonal degeneration were found in the hippocampus, corticospinal tract and optic chiasm. 

      These results show that by using sufficiently sensitive neurohistological technique, the neurotoxicity of methanol is revealed in the mammalian central nervous system.

      Although the daily dosage used in this study is high (approx. equivalent to 3 liters of an aspartame-sweetened beverage), pregnant or lactating women should limit consumption of aspartame-sweetened food products. [ "3 liters" -- this may be a typo or mistake ]



      Results


      Experiment I: Physical Growth, Malformation and External Deformity

      There were no significant difference in body weight gain among the two groups of pregnant dams.

      However, prenatal exposure to methanol significantly reduced body weight of the offspring at birth, relative to control group (Table 1).

      The incidence of malformation was significantly increased in the methanol treated group (Table 2).

      Fourteen of the 20 litters had one or more fetuses with malformation of external morphology (p 0.001, 55 of the 263 total fetuses).

      The external malformations included brain defects, eye defects and spina bifida occulta.

      The incidence of brain defects was significant (p 0.005),
      specifically, seven of the 20 litters had thirteen fetuses with hydrocephalus,
      and one little had two fetuses with aencephaly and one fetus with exencephaly.

      The incidence of eye defects was significant (p 0.01),
      specifically, four of the 20 litters had eleven fetuses with anophthalimia,
      and one litter had two fetuses with microopthalmia.

      The incidence of spina bifida occulta was significant (p 0.005),
      thirteen of the 20 litters had one of more fetuses with this spinal deformity.

      The incidence of still born was also significant (p 0.005),
      fourteen of the 20 litters had one or more still born fetuses (25 out of 263 total fetuses).

      The incidence of pups mortality rate was higher in the first week of nursing period,
      twelve of the 20 litters had one or more dead fetuses within first week of their birth.

      All of the dead pups had severe breathing problem for two/three days and lost weight before they passed away.

      This strongly suggests that methanol or its metabolic products, either formaldehyde or formic acid can cause severe problem in pups during nursing period.

      Overall, prenatal methanol exposure significantly affected mean fetal birth weight and also affected the incidence of brain defects, eye defects, spina bifida and stillborn, all significant.


      Histological Examination

      In this study, methanol was administered orally to rats during gestational and lactational period was neurotoxic at 1.28 g/kg dose level. 

      Histopathological examination indicates that methanol causes damage to the central nervous system of rats and their offspring.

      The effect of prenatal methanol exposure during lactation period were observed under light microscope.

      All pups showed massive degeneration in widely distributed regions of the central nervous system (See Table 3).

      Microscopic examination revealed that degenerating fibers in the different layers of cerebellar cortex, several cranial nerve nuclei, and in the brain stem.

      All experimental pups showed axonal damage, independent of the sex of pups.

      Microscopic examination indicates anterior magdola and periventicular gray have degenerating fiber.

      Inferior colliculus and autorhinal cortex of the pups brain have beaded axon.

      The normal nerve fibers of the control rat brain were photographed using light microscope (See Figure 6).

      All experimental pups showed massive degeneration in the different layer of cerebellar cortex (See Figure 7).

      Degeneration was also found in subliculum.

      Enthorhinal cortex also had some degeneration;
      and the hippocampus were filled with degenerating fibers throughout the region (See Figure 8).

      Methanol was toxic to the dams at the 1.28/kg dose level.

      All experimental animals showed massive degeneration in numerous regions of the brain (See Table 3).

      Microscopic analysis indicated massive degeneration in the white matter on the ventral-most aspect of the brain.

      In gray matter there also appears to be a number of broken axons and this region has degeneration throughout the entire ventral surface.

      Light microscopic examination showed complete absence of detectable neuronal cell body.

      All experimental animals showed massive degeneration throughout the medullary layer of the cerebellum.

      Microscopic analysis indicates beaded axons all over the cerebellur medulla.

      In the cerebellum cortex, axons found were beaded and degenerated.

      Degeneration also found in the cerebellum and brain stem, especially in the ventral and lateral most tracts.

      Degenerating fiber were found in motor nucleus of the trigeminal nerve (and particularly the nucleus of spinal tract of the trigeminal (5th) nerve), the trapezoid body, the trigeminal nerve, corticospinal tract and rubrospinal tract.

      Exceptionally heavy degeneration was observed in the flocculus and in the ventral-most aspect of the periventricular gray (dorsal raphe nucleus).

      There was also extensive damage to the dorsal and ventral cochlear nuclei.

      The optic chiasm showed areas of degeneration.

      All experimental animals exhibited beaded axons and massive degeneration throughout the hippocampus regions.

      The thalamus, hypothalamus, cingulate cortex, substantia nigra and the reticular formation showed no signs of degeneration in any animals (See Table 4).


      Experiment II

      In this study, methanol, administered orally to adult rats for six weeks, was neurotoxic to rats.

      Microscopic examination indicates extensive destruction of myelin throughout the cerebral hemispheres of the rat brain.

      Histological examination showed massive degeneration in the white matter on the ventral-most aspect of the brain.

      In the gray matter there also appeared lost of broken axons and this region has degeneration throughout the entire ventral surface.

      Beaded axons were found in cerebellar medulla, cerebellar cortex and brain stem, especially medulla, cerebellar cortex and brain stem, especially in the ventral and lateral most tracts
      .
      Microscopical examination showed degeneration in motor nucleus of the trigeminal nerve.

      The spinal tract, trigeminal, and trapezoid were filled with degenerating fiber.

      Optic chiasm had patchy fibers of degeneration.

      Hippocampus wer

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