Abstract
A reversed-phase high performance liquid chromatographic(RP-HPLC) method for the determination of cytosine and 5-methylcytosine in flue-cured tobacco leaves were developed and optimized in mobile phase,detecting temperature and flow rate.The DNA was extracted by CTAB method and hydrolyzed with perchloric acid.Then the pH value was adjusted to between 3 and 5 with KOH.Cytosine and 5-methylcytosine were separated on a reversed-phase C18 column(250 mm×4.6 mm,5 μm),with the mobile phase of methanol+ 5 mmol/L pentane sodium(10︰90,V/V) at a flow rate of 0.7 ml/min.The ultraviolet detector was operated at 273 nm at 30℃.The injection volume was 20 μl.The recoveries of cytosine and 5-methylcytosine in flue-cured tobacco leaves were 103.50% and 96.67% respectively with the relative standard deviation of 2.35% and 2.80% respectively.This method was applied to determine the cytosine and 5-methylcytosine changes in flue-cured tobacco leaves during growth period and the satisfactory results were obtained.
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Claims 7
Claim · model-written summary
RP-HPLC separated cytosine in tobacco leaf DNA.
Not reportedComparatorValue
Verbatim
e0“Cytosine and 5-methylcytosine were separated on a reversed-phase C18 column”
t0“Flue-cured Tobacco Leaves”
Claim · model-written summary
RP-HPLC separated 5-methylcytosine in tobacco leaf DNA.
Not reportedComparatorValue
Verbatim
e0“Cytosine and 5-methylcytosine were separated on a reversed-phase C18 column”
t0“Flue-cured Tobacco Leaves”
Claim · model-written summary
Cytosine recovery in tobacco leaves was 103.50%.
Verbatim
e0“The recoveries of cytosine and 5-methylcytosine in flue-cured tobacco leaves were 103.50% and 96.67% respectively with the relative standard deviation of 2.35% and 2.80% respectively”
Claim · model-written summary
5-methylcytosine recovery in tobacco leaves was 96.67%.
Verbatim
e0“The recoveries of cytosine and 5-methylcytosine in flue-cured tobacco leaves were 103.50% and 96.67% respectively with the relative standard deviation of 2.35% and 2.80% respectively”
Claim · model-written summary
The cytosine assay had relative standard deviation 2.35%.
Verbatim
e0“The recoveries of cytosine and 5-methylcytosine in flue-cured tobacco leaves were 103.50% and 96.67% respectively with the relative standard deviation of 2.35% and 2.80% respectively”
Claim · model-written summary
The 5-methylcytosine assay had relative standard deviation 2.80%.
Verbatim
e0“The recoveries of cytosine and 5-methylcytosine in flue-cured tobacco leaves were 103.50% and 96.67% respectively with the relative standard deviation of 2.35% and 2.80% respectively”
Claim · model-written summary
The method gave satisfactory results when applied across tobacco leaf growth.
Not reportedComparatorValue
Verbatim
e0“This method was applied to determine the cytosine and 5-methylcytosine changes in flue-cured tobacco leaves during growth period and the satisfactory results were obtained”