ABSTRACTS FROM: The Prolongation of the Lifespan of Rats by Repeated Oral Administration of [60]Fullerene
Link to article: https://www.researchgate.net/publication/224004891_The_prolongation_of_the_lifespan_of_rats_by_repeated_oral_administration_of_60fullerene
the metabolic fate and the in vivo chronic effects of C60itself still remain unknown. fullerene is active only in soluble form. the extremely slow dissolution of C60in biological media prevents controlling accurately the active fraction .
the chronic toxicity of C60has never been investigated to our knowledge.C60is soluble in lipid droplets inside living cells as well as in fats in general [2. Moreover, C60can freely cross membrane barriers. C60interactions with living systems as well as its toxicity should be determined using solubleforms.
While C60solubility in vegetable oils solutions should be quite appropriate for studying its chronic toxicity at reiterated doses. the in vivo behaviour of soluble forms of C60 was unknown, we determined the in vivo fate of C60dissolved in olive oil before studying its chronic effects at reiterated doses.
Virgin olive oil is obtained from a Chemlali Boughrara cultivar from Tunisia planted in the Sahel area. C60(purity 99.98%) was obtained from SES ResearchCorporation (USA) and used without further purification. Fifty mg of C60were dissolved in 10 ml of olive oil by stirring for 2 weeks at ambient temperature in the dark. The resulting mixture was centrifuged at 5.000 g for1 h and the supernatant was filtered through a Millipore filter with 0.25mm porosity.
Chronic toxicity and effects of C60on survival of rats
The rats were housed three per cage and acclimated for 14 days, before dosing.Three groups of 6 rats (10 months old, weighing 465 31 g) were administered daily for one week, then weekly until the end of the second month and then every two weeks until the end of the 7th month, by gavages with 1 ml of water or olive oil orC60dissolved in olive oil (0.8 mg/ml), respectively.
The rats were weighed before each dosing. Routine observations following official recommendations [27] were made on all animals inside and outside the cageonce a day throughout the study for signs of departure from normal activity,morbidity and mortality.
Sixty rats randomly divided into 10 groups of 6 rats were pre-treated daily for 7days by oral gavages (og groups) or by i.p. injection (ip groups). Groups A (GAog andGAip), received 1 ml of water. Groups B and C (GBog, GCog and GBip, GCip) were pre-treated with 1 ml of olive oil while groups D and E (GDog, GEog and GDip, GEip)were pre-treated with 1 ml of C60-olive oil.
The resulting C60-olive oil solution is purple and contains0.80 0.02 mg/ml (n ¼ 6) The stability of both oily and control solutions stored at ambient temperature and in the dark was checked monthly during 48months. No change was recorded under our chromatographic conditions C60is well distributed in tissues.
Chronic toxicity and effects of C60on lifespan of rats
Fig. 3 shows the animal survival and growth. After five months of treatment (M15) one rat treated with water only exhibited some palpable tumours in the abdomen region. Due to the rapid development of tumours (about 4 cm of diameter) this rat died at M17. As rats are known to be sensitive to gavages, we decided to stop the treatment for all rats and to observe their behaviour and overall survival.
All remaining animals survived with no apparent sign of behavioural trouble until M25(Fig. 3a). At the end of M25 the animals of the control groups showed signs of ulcerative dermatitis with ageing while C60-treated animals remained normal. As the growths of all surviving animals showed no significant difference until M30(Fig. 3b) indicating that the treatment did not alter their food intake, we continued observing their survival.
At M38 all water-treated control rats were dead (Fig. 3a). This agrees with the expected lifespan of this animal species that is thirty to thirty six months. At this time 67% of olive-oil-treated rats and 100% of C60-treated rats were still alive.
The estimated median lifespan (EML) for the C60-treated rats was 42months while the EMLs for control rats and olive oil-treated ratswere 22 and 26 months, respectively. These are increases of 18 and 90% for the olive-oil and C60-treated rats, respectively, as compared to controls.
olive oil extends the lifespan of rats with respect to water with a probability of 0.99 while C60-olive oil extends the lifespan of C60 treated rats with a probability of 0.999 and 0.995 with respect to water and olive oil treatments, respectively.
. C60-olive oil solution preparation
The C60-olive oil solution used in this study can be considered as free of C60aggregates because its colour is purple that is characteristic of C60solutions AND it is freely and instantaneously soluble in toluene.
The stability of C60-olive oil solution determined under our experimental conditions agrees with recently published results showing that the addition of [60]fullerene significantly hampers the peroxide formation thus increasing the stability of the tested oils . The results of this pharmacokinetic study show for the first timethat C60is absorbed by the gastro-intestinal tract
The elimination process follows a non-urinary route because unmodified C60was not detected in urine samples C60is mainly eliminated through the bile ducts C60reacts inside the liver cells with vitamin A. As C60and some of its derivatives mainly accumulate in the livers and spleens we also studied the accumulation of C60in these organs after 7 successive days of administration.
C60molecules are eliminated from the organs in a few hours after both oral and i.p. administrations. the presence of significant amounts in the brain 24 hours after both oral and i.p. administrations under our experimental conditions confirms that solubilized C60can cross the bloodebrain barrier
Chronic toxicity and effects of C60on survival of treated rats
C60has no acute or sub-acute toxicity inrodents it can act as an antioxidant, we investigated its chronic toxicity concomitantly with its effects on the survival of rats. Ten-month old male rats (M10) were chosen instead of young ratsas officially recommended, in order to avoid possible compensatory effects that can occur during early development. we treated the rats daily only during 7 days and weekly during the first two months, then every two weeks until one control rat died.
Our results show that while olive oil treatment can lead to anincrease of 18% of lifespan of treated rats, C60-olive oil can increaseit up to 90%, as compared to controls. at M38 all C60-treated rats were still alive. C60should be the most efficient ever material for extending lifespan. C60itself is a powerful antioxidant
This is the first investigation of the in vivo chronic effects ofa soluble form of C60. C60is a powerful antioxidant .C60can scavenge large numbersof free radicals. C60isa powerful liver-protective agent. C60is ableto scavenge in vitro a large number of radicals per molecule and be this property can be involved in the mechanism of protection against toxicity,The results obtained for antioxidant enzyme activities also confirm the antioxidant effect of C60against toxicity.
the protective effect involves a free-radical scavenging mechanism. dissolved C60appears hundred times more active than when it is in suspension. In fact the action of soluble C60is immediate while that of suspended C60is delayed because it has to be dissolved to act. In all cases, based on C60-livercontent (Table 2), these results prove that this fullerene is active atthe nano molar level. However, the involved mechanism remains to be established.
For the time being the hypothesis of free-radical scavenging remains possible. C60could act as a free-radicalscavenger as it has been widely demonstrated in vitro, but up to now no resulting C60adduct has been observed in vivo.
Conclusion The effect of pristine C60on lifespan emphasizes the absence of chronic toxicity. In the present case, the treatment was stopped when a control rat died at M17, which proves that the effects of the C60treatment are long-lasting as the estimated median lifespan for C60-treated rats is 42 months. It can be thought that a longer treatment could have generated even longer lifespans. Anyway, this work should open the road towards the developmentof the considerable potential of C60in the biomedical field,including cancer therapy, neurodegenerative disorders and ageing.Furthermore, in the field of ageing, as C60can be administered orally and as it is now produced in tons,
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It is not known how C60 greatly extends the life of mammals. Hypotheses vary from extremely effective antioxidant action to DNA-demethylation (which would permanently rejuvenate an individual on the genetic-molecular level).
The latter hypothesis fits the observed test results of various Buckminsterfullerene animal trials better than the former, with old mice aquiring the problem-solving power and memory of young mice, and rats nearly doubling their expected lifespan with just two dozen administrations of C60.
It has been postulated that Buckminsterfullerene becomes a semi-permanent constituent of the mitochondrial lipid bilayer, where it functions as an extremely effective antioxidant, since it is not used up and keeps getting re-absorbed in the mitochondrial lifecycle.
The C60 molecule acts as an "electron reservoir" and can accept or donate multiple electrons and thus neutralize all types of free radicals indefinitely - until it leaves the body. There are indications that it takes weeks for all C60 to leave the lipid bilayers of the cells and mitochondria.
About short-term effects: Except for true athletes who reported increased strength and stamina as well as reduced recovery times and hence were able to achieve sustained gains, we do not think there will be any short-term beneficial effects for ordinary people.
A lot of what is mentioned online about short-term benefits is the Placebo effect, in our opinion. It is completely impossible to note that one ages say 15% slower, neither can one feel that tumors do not appear in the body.
"When the last rat in the control group had died,
all rats in the C60-olive oil group were still alive and they went on to live for an average of two more years."
Lipofullerene C60 semi-permanently embedded in the lipid bilayers of the cells and mitochondria, where it acts as a reusable, recyclable antioxidant:
The C60 molecule is not damaged when it accepts or donates electrons from/to free radicals. When a cell or mitochondrion dies, the lipofullerene C60 can be reused by other cells.
C60 IS EXTREMELY SAFE
1000 mg/kg/day of C60 has been proven to be non-toxic in this clinical study. The famous French C60 rat toxicity study that showed a 90% lifespan increase in rats used a few times times higher dose than our recommendation, 0.02 mg/kg/day.
THE LAST RAT WAS KILLED
The Baati study rats took only 24 doses during their entire lifetime, over a period of just 7 months and 24 doses only, starting when they were 10 months old. The increase in lifespan is officially 90% (nearly twice as old!) but in reality it would be even higher, because Dr. Fathi Moussa admitted recently that they killed the last surviving rat when it was 5.5 years old because they wanted to wrap up the study.
There even circulates a rumor that one more surviving rat was killed in order to be able to publish this groundbreaking study sooner, in fear other researchers may be first. There is no substance that prolongs the life of mammals longer than C60 in olive oil.
Not even the most expensive supplements such as Resveratrol come remotely close, and neither are those supplements supported by such solid evidence as the Baati study. Of course, humans can NOT expect a doubling of lifespan like rats but we estimate at most a 20% average increase - possibly cancer-free, like the rats.
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Aerobic- & strength gains, longevity, brain rejuvenation and tumor prevention with C60 olive oil
We found animal studies that documented brain rejuvenation in mice and the near-doubling of lifespan (PDF) and the prevention of cancer and the diseases of old age in rats.
We also found toxicological studies that show that C60 (Buckminsterfullerene) in olive oil is safe. C60 in olive oil also has an extremely effective liver-protective effect, as documented in the same publication, where rats were poisoned to death with liver-destroying chemicals but the rats that took C60 barely were affected at all.
Prof. Moussa mentions this in the interview: "The rats that took C60 did not develop any tumors, unlike the rats in the control group."
Professor Moussa, leader of the study team says in the interview that C60 (Buckminsterfullerene) is totally non-toxic. He has been researching Fullerene C60 for 18 years.
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And the rat study is not a fluke - a similar C60 study on mice reported not just an increased lifespan - also very greatly improved learning and memory. Very old mice (near end of life) performed as young mice (6 months old).
it is not even known whether the health benefits come purely from the C60, or whether the C60 merely acts as a vehicle to bring long chain fatty acids into the cells. Or whether there is a synergy at play
There must be something else hitherto undiscovered about this unique sphere-shaped molecule. C60, when dissolved in olive oil and ingested orally, gets absorbed and the C60 mulecules become distributed - albeit in small concentrations - throughout the body and intracellularly, even beyond the blood-brain barrier in minute amounts.
Big Pharma is working to patent a water-soluble designer-fullerene and sell it extremely expensively as a life-extension drug. They will have little trouble persuading the FDA, their enforcer, to ban all other natural fullerenes for being "toxic"
The cell walls are lipids and lipofullerene (C60 with adducts of the long-chain fatty acids of olive oil) therefore is readily absorbed by the cells and ends up where it is most needed - provenly in the mitochondrial bilayer membranes, where it will neutralize ROS (reactive oxygen species) like no other molecule, because C60 can function as an antioxidant in all perpetuity, it is not "used up" by performing its antioxidant action because it can capture high-energy electrons all the way up to Beta-radiation level, reduce their energy in the C60 matrix and drain them by electrical instead of chemical means.
Better absorption happens when smaller dosages are taken more frequently.