Isolation and Purification of Coenzyme Q10 and its dietary effects on mice.

1
Isolation and Purification of Coenzyme Q10 and
its dietary effects on mice.

• Presented By
• Afroza Khatun
•                          Animal Reproductive
  Physiology Lab.
•         17 May 2007

2
What is Coenzyme Q10?

• fat soluble vitamin-like substance
• present in every cell of the body
• serves as a coenzyme
• It is a biologically active quinone
• Its structure related to vitamin K and E.
• Chemically, CoQ10 is known as
• 2,3-dimethyoxy-5-methyl-6-decaprenyl
• -1,4- benzoquinone,

3
Synonyms of Coenzyme Q10

• Coenzyme Q, Co-enzyme Q10, Coenzyme Q(50), CoQ
• Idebenone (synthetic Q10),
• Ubiquinone-10, Ubiquinone 50, Ubidecarenone,
  Neuquinone, Ubiquinone 10, Ubiquinone-Q10,
  Udekinon, vitamin q10

4
Deficiency of CoQ10
-Impairment in CoQ10 biosynthesis -Excessive
utilization of CoQ10 by the body -Insufficient
dietary CoQ10 (chronic malnutrition) -Any
combination of the three factors
5
Sources of CoQ10 in the body

• heart, liver, kidneys and pancreas (highest)
• lungs (lowest)
• Moleclar formulaC59H90O4
• Molucalar weight 863.34

6
Objectives

• i) Isolation and purification of Coenzyme Q10 in
  different organ of mice.
• ii) To know the dietary effect of Coenzyme Q10 on
  the different organs and  blood.

7
Materials and Method

• Preparation of Diet and animals
• 3 diets
• Diet 1 (Control)
• Diet 2 (100 IU VE)
• Diet 3 (500 mg CQ10/kg 100 IU VE)
• Animals Male ICR mice (n 48 Male
• 8 weeks old), n8 per diet

8
Preparation of Blood sample
Collected blood from mice by cardiac puncture

•  

Centrifuged at 6000X g at room temp.for 15 min
100µl plasma was collected
20µl IS 500µl distilled water 10µl 1 FeCl3
9

Vortexed for 5s
added 2.0 ml methanol and 3.0 ml hexane
Centrifugation at 1870?g for 10min
Hexane layer was collected
further extracted with 3.0 ml of hexane
Collected the hexane layer
10
Two hexane layers were mixed and evaporated
Reconstituted in 200µl of ethanol
100µl of the solution is injected
11
Preparation of tissue sample
Collected organs
Homogenized with d H2O
Added 20µl IS 2ml ethanol and 5ml n-hexane
Vortexed for 10mins
Centrifugation
Hexane layer was collected
12
Add 2ml of ethanol and 0.5 ml of 5 potassium
hexacyanoferrate solution
Reaction for 10 mins
Then add 2ml ethanol and 5ml n-hexane
Collected the hexane layer
Two hexane layers were mixed and evaporated
Reconstituted in 200µl of ethanol
100µl injected into an LC
13

• Analysis of Liquid Chromatography

Wash the column with Methanol Hexane (82)
solution
Put 100µl sample on the top of the column
1st elute the column with methanol
Collect the 10 bottles elute
2nd elute the column with hexane
14
Then collect the 10 bottles elute
Cheque the absorbtion rate by spectrophotometer
at 275 nm 
15
Results
16
Table 1 Concentration of CoQ10 in the organs of
male and female mice.
17
Table 2 Coenzyme Q10 concentrations in the
tissues of mice after VE and CoQ10
supplementation for 28 days.
18
Conclution

• Ovary contains higher CoQ10 than testis.
• Dietary administration of CoQ10 enhanced the
  concentration in testis, liver and blood.

19

• Thank you