Abstract BACKGROUND: Several studies revealed that vitamin E supplementation results in an increase of AChE activity, whereas L-carnitine addition causes its partial restoration. L carnitine, a naturally occurring amino acid, is potentially effective in preventing peripheral neuropathy as well as lessening neuropathic symptoms. These actions account for the popular use of L carnitine and Vitamin E as an antiaging or memory-supportive nutrient and delaying progression of neurodegenerative diseases. SUBJECTS AND METHODS: A comparative invitro study was performed, in which blood samples were collected from 40 healthy individuals. Erythrocyte membranes were separated from blood of the different samples, and then erythrocyte membranes from each sample were subdivided into 4 groups: group (1) is considered as a control group, group (2) incubation of erythrocyte membranes with vitamin E, group (3) incubation of erythrocyte membranes with L carnitine and group (4) incubation of erythrocyte membranes with vitamin E and L carnitine. Acetylcholine esterase was measured spectrophotometrically, and its levels were evaluated in these groups. RESULTS: Comparison between the acetylcholine esterase levels in the control group and the other groups showed highest level of the enzyme in the (Vit. E + L carnitine) group without a statistically significant difference (p >0.05). CONCLUSIONS: Our study showed that the best protective role for erythrocyte membrane acetylcholine esterase was achieved when both L carnitine and vitamin E were added. Keywords: Acetylcholine esterase – erythrocyte membrane - Vitamin E – L Carnitine

1. 1
Protective Role of Vitamin E and L carnitine on Human
Erythrocyte Membrane Acetylcholine Esterase Activity
Nevine El Kady, Usama Mohamed El-Barrany, Khaled Bayoumi, Mohamed Adly
Department of Forensic Medicine and Clinical Toxicology, Faculty of Medicine, Cairo University
Abstract
BACKGROUND: Several studies revealed that vitamin E supplementation results in an
increase of AChE activity, whereas L-carnitine addition causes its partial restoration. L carnitine,
a naturally occurring amino acid, is potentially effective in preventing peripheral neuropathy as
well as lessening neuropathic symptoms. These actions account for the popular use of L carnitine
and Vitamin E as an antiaging or memory-supportive nutrient and delaying progression of
neurodegenerative diseases.
SUBJECTS AND METHODS: A comparative invitro study was performed, in which blood
samples were collected from 40 healthy individuals. Erythrocyte membranes were separated
from blood of the different samples, and then erythrocyte membranes from each sample were
subdivided into 4 groups: group (1) is considered as a control group, group (2) incubation of
erythrocyte membranes with vitamin E, group (3) incubation of erythrocyte membranes with L
carnitine and group (4) incubation of erythrocyte membranes with vitamin E and L carnitine.
Acetylcholine esterase was measured spectrophotometrically, and its levels were evaluated in
these groups.
RESULTS: Comparison between the acetylcholine esterase levels in the control group and the
other groups showed highest level of the enzyme in the (Vit. E + L carnitine) group without a
statistically significant difference (p >0.05).
CONCLUSIONS: Our study showed that the best protective role for erythrocyte membrane
acetylcholine esterase was achieved when both L carnitine and vitamin E were added.
Keywords: Acetylcholine esterase – erythrocyte membrane - Vitamin E – L Carnitine
Conflicts of interest: None declared.
INTRODUCTION
Acetylcholine esterases belong to a family of proteins that is widely distributed
throughout the body in both neuronal and non-neuronal tissues. Acetylcholine esterase, also

2. 2
known as AChE or acetyl hydrolase, is a hydrolase that hydrolyzes the neurotransmitter
acetylcholine (Ach). AChE is found at mainly neuromuscular junctions and cholinergic brain
synapses, where its activity serves to terminate synaptic transmission [1,2].
Vitamin E refers to a group of ten lipid-soluble compounds that include both
tocopherols and tocotrienols. Of the many different forms of vitamin E, γ-tocopherol is the
most common while α-tocopherol, the most biologically active form of vitamin E, is the second-
most common form of vitamin E in the diet [3].
Vitamin E has many biological functions, the antioxidant function being the most
important function. There are other functions including enzymatic activities, gene expression
e.g. in atherosclerosis [4], and neurological functions. Vitamin E is also used as a commercial
antioxidant in ultra-high molecular weight polyethylene used in hip and knee replacements, to
help resist oxidation [5].
Vitamin E also plays a role in neurological functions; increasing AChE activity, inhibiting
platelet aggregation, protecting lipids and preventing the oxidation of polyunsaturated fatty
acids [6 – 8].
Vitamin E can act as an anticoagulant, increasing the risk of bleeding problems. As a
result, many agencies have set a tolerable upper intake levels at 1,000 mg (1,500 IU) per day. In
combination with certain other drugs such as aspirin, hypervitaminosis E can be life-
threatening. Hypervitaminosis E may also counteract vitamin K, leading to a vitamin K
deficiency [8].
Carnitine is a quaternary ammonium compound biosynthesized from the amino acids
lysine and methionine. The highest concentrations of carnitine are found in red meat. Carnitine
can be found at significantly lower levels in many other foods including nuts and seeds as
sunflower, sesame, beans, peas; vegetables as artichokes, broccoli, sprouts and garlic; fruits as
apricots and bananas; cereals as corn, oatmeal, rice bran, wheat, and other foods as bee pollen,
yeast and carob [9].

3. 3
It is required for the transport of fatty acids from the intermembraneous space in the
mitochondria, into the mitochondrial matrix during the breakdown of lipids for the generation
of metabolic energy. It is widely available as a nutritional supplement. Carnitine exists in two
stereoisomers: its biologically active form is L-carnitine, whereas its enantiomer, D-carnitine, is
biologically inactive [10].
There may be a link between dietary consumption of carnitine and atherosclerosis, but
there is also evidence that it lowers the risk of mortality and arrhythmias after an acute
myocardial infarction. In the course of human aging, carnitine concentration in cells diminishes,
affecting fatty acid metabolism in various tissues. Particularly adversely affected are bones,
which require continuous reconstructive and metabolic functions of osteoblasts for
maintenance of bone mass [11].
Carnitine has been proposed as a supplement to treat a variety of health conditions
including heart attack, heart failure, angina, narcolepsy, and diabetic neuropathy, but not
fatigue, improving exercise performance, nor wasting syndrome (weight loss). There is also
some suggestion that use of carnitine may improve sperm motility in men with sperm problems
[12 – 15].
L carnitine, a naturally occurring amino acid, is potentially effective at preventing
peripheral neuropathy as well as lessening neuropathic symptoms. It is also involved in fatty
acid beta-oxidation in mitochondria, with influencing neurotransmitters, including acetylcholine
and dopamine. L carnitine may also prevent neural degeneration related to aging in the brain
through the preservation of the neurotrophin and nerve growth factor. These actions have
been known for decades and account for the popular use of L carnitine as an antiaging or
memory-supportive nutrient [16].
Vitamin E (α-tocopherol) and L carnitine were found to reduce lipid peroxidation. A
study was done to investigate the erythrocyte membrane acetylcholine esterase (AChE) in
basketball players with or without vitamin E supplementation, before and after training. Also
the study aimed to find out any additional effect of L carnitine on the enzyme activities [6, 17].

4. 4
AChE activity was found increased after the exercises and decreased after the game. In
vitro incubation of erythrocyte membranes from players after the game with L carnitine
resulted in a partially restoration of the membrane AChE activity. As a conclusion, vitamin E
supplementation results in an increase of AChE activity, whereas L carnitine addition may
restore AChE activity [6].
The aim of this study is to evaluate the protective role of vitamin E as well as L
carnitine either alone or in combination on human erythrocyte membranes through its effect
on acetylcholine esterase (AChE) activity.
MATERIALS & METHODS
This was a prospective comparative invitro study, where blood samples were obtained
from 40 healthy individuals above 12 years of age. Consent was taken from each one before
sampling extraction.
Erythrocyte membranes will be separated from blood of the different samples, and
then erythrocyte membranes from each sample will be subdivided into the following 4 groups:
group (1) is considered as a control group in which erythrocyte membranes will be left alone,
group (2) incubation of erythrocyte membranes with vitamin E, group (3) incubation of
erythrocyte membranes with L carnitine and group (4) incubation of erythrocyte membranes
with vitamin E and L carnitine. Acetylcholine esterase was measured spectrophotometrically, in
these groups.
Data were statistically described in terms of mean ± standard deviation (± SD), median
and range. Comparison between the study groups was done using one way analysis of variance
(ANOVA) test with posthoc multiple 2-group comparisons. p values less than 0.05 were
considered statistically significant.
RESULTS

5. 5
Spectrophotometric assessment showed that the mean level of acetylcholine esterase
was highest in the vitamin E+ L carnitine groups then in the L- carnitine group (table 1, Fig. 1).
Table (1): The mean, standard deviation, minimum, maximum and median of the results of the
acetylcholine esterase results in all the groups:
Group *N Mean
(IU/L)
*Std. Deviation Minimum Maximum Median
Control 40 4.5770 0.61320 3.80 5.80 4.4000
*Vit.E 40 4.3560 0.57746 3.10 5.20 4.5000
*L.car. 40 4.6150 1.04291 2.90 6.80 4.4500
*Vit.E+
*L.car.
40 4.6645 0.63299 3.80 5.80 4.4500
* Vit.E: Vitamin E, L.car.: L carnitine, N: number, Std. Deviation: standard deviation.
Figure (1) Mean level of AChE in the different groups
Comparison between the acetylcholine esterase levels in the control group and the
other groups: the Vitamin E, L carnitine and Vitamin E+ L carnitine groups showed no
statistically significant difference (p >0.05). (table 2)
Control
Group
The other
Groups
Mean
Difference
*Std. Error *P value 95% Confidence
Interval
Upper Upper
4.2
4.25
4.3
4.35
4.4
4.45
4.5
4.55
4.6
4.65
4.7
Control Vit E L Carnitine Vit E + L
carnitine
Mean AchE (IU/L)

6. 6
Bound Bound
*Vit. E 0.22100 0.20250 1.000 -0.4229 0.8649
*L car. -0.03800 0.20250 1.000 -0.6819 0.6059
*Vit. E+ *L
car.
-0.08750 0.20250 1.000 -0.7314 0.5564
Table (2): Comparison between the acetyl cholinesterase level of the control group and the other
groups:
DISCUSSION
Vitamin E (α-tocopherol) and carnitine were found to reduce lipid peroxidation. A
study performed on basketball players revealed that vitamin E supplementation results in an
increase of AChE activity, whereas L-carnitine addition causes partially restoration AChE activity
[6].
Lee and co-workers (2001) study also agreed with our results, which studied the
effects of vitamins E and C on the levels of neurotransmitters and acetylcholine esterase activity
in the brains of rats treated with scopolamine, an inducer of dementia. They found that the
supplementation of vitamins E and C in the diet significantly increased (p <0.05) the reduced
brain acetyl cholinesterase activity up to the level of the scopolamine-untreated group [18].
A study was performed to evaluate the effect of L carnitine in association with
Acetylcholine esterase inhibitors in 23 patients with mild Alzheimer's disease. Clinical
evaluation of cognitive functions, functional status and behavioral symptoms showed the
response rate, which was 38% after Acetylcholine esterase inhibitors treatment, significantly
increased to 50% (p <0.05) after the addition of L carnitine [19].
A study performed by Ahmed and his colleagues (2012) explained the important role
of dietary supplementation (vitamin E and L carnitine) in delaying the progression of age-
related neurodegenerative diseases e.g. Alzheimer disease [20]. L carnitine may also prevent
neural degeneration related to aging in the brain through the preservation of the neurotrophin,
nerve growth factor [16].

7. 7
CONCLUSION
Several studies revealed the protective role of vitamin E and L carnitine on the
erythrocyte membrane acetylcholine esterase by lowering its levels. One of their clinical
applications is in delaying the progression of age-related neurodegenerative diseases e.g.
Alzheimer disease. Our study showed that the best protective role for erythrocyte membrane
acetylcholine esterase was achieved when both L carnitine and vitamin E were added.
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