Effects of curcumin supplementation on liver metabolism in rats fed a high-fat diet: A proteomic approach

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_POSTERS

M361

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E. McClure, M. Jarvis

AB

SCIEX, Concord, ON, Canada

Background:Micro-flow liquid chromatography has become a compelling alternative to conventional HPLC for many analyses given

its

cost-reduction and time-saving potential. Lower solvent consumption resulting from the use of micro-flow rates

(5-60 pl/min), as compared to a typical flow rate of 500 pl/min using conventional HPLC, significantly reduces solvent and waste disposal

costs.

ln

addition, high on-column linear velocities and low mixing and delay volumes a|low for fast chromatography and thus higher sample throughput. These

benefits are realized while sensitivity is maintained or enhanced as compared to conventional HPLC, making micro-flow LC an exceilent fit for the analysis of Vitamin D given the sensitivity and throughput requirements of this assay.

Methods: Here we present the details of the analysis of 25-monohydroxy Vitamin D2 and 2S-monohydroxy Vitamin D3

using

the

Eksigent

ekspertTM

microlC

200

system

in

combination with the

AB SClEX

QTRAP@ 4500 LC/MS/MS system. Chromatographic separation of the two compounds is achieved using a Halo C18 column (0.5x50 mm,2.7 pm) and a water/methanol/formic acid gradient. Total runtime for the ana|ysis is 3 minutes, which is achieved at a flow rate of 20

pl/min. To avoid introducing early-eluting matrix to the system,

the first part of the gradient is diveıled to waste thus improving robustness and column lifetime.

Results: The Limit of Quantitation (LOQ) for the analysis was determined using 'blank'stripped serum and spiked serum and found to be below 4 nglmL, and 5 ng/ml for 2S-monohydroxy Vitamin D3 and 2S-monohydroxy Vitamin D2, respectively, Accuracies were within 92-109o/o across the calibration range (1-"l00 ng/ml), with CVs below 15% based on n=5 for each calibration concentration. Serum samples from donors were also analyzed using the method described above, and the results indicate that method peı,formance is comparable to that

of a conventional HPLC method.

Conclusions: An LC-MS/MS method has been developed for the analysis of 25-hydroxyvitamin D2 and D3 in human serum

using

micro-flow

,LC,

resulting

in

a

reduction

of

solvent consumption and faster analytical run-times.

M362

EFFECTS

OF CURCUMlN SUPPLEMENTATION ON

LıVER METABoLıSM

lN RATS FED A HIGH-FAT D|ET

A

PROTEOMİC

APPROACH

H. Kocak(1), Y. Oner-lyidogan(2), A.T. Bayka|(3), M.

Seyidhanoglu(2), A. Cevik(a), F. Gurdol(2), M. Uys21(2)

1Biochemistry, _{Faculty of Medicine, Istanbul Bilim lJniversity,} Esentepe. lstanbul. Turkey

'Biochemistry. lstanbul Faculty of Medicine, lstanbul University, _Çapa, Turkey

" Genetic Engineering and Biotechnology _lnstitute,

rÜ BİTeX-Uarmara Research Center, Kocaeli, Turkey aLaboratory _{Animal Biology and Applied Biomedical}

Techniques, lnstitute for Experimental Research and Applied Medical Research, lstanbul _{University, Çapa,}

Turkey

Background: ln the modern world, many individuals routinely consume diets rich in bnimal fat, leading to a high incidence of obesity. Curcumin (d iferuloylmethane, iu rmeric) extracted from

the dried root of the rhizome Curcuma longa, is a popularAsian

dietary

spice used in

curry. Curcumin

is the

most active component of turmeric, comprising 2-8olo by weight. Proteomics is an important tool for elucidating the cellular response to

various treatments including dietary factors.The aim of the study was to investigate the effect of curcumin treatment on

hepatic

proteome

in

rats

fed

high-fat

diet

(HFD). Methods: Male Sprague-Dawley rats were divided into six groups. Group ,1 _{was fed control diet ('10% of total calories from}

fat). Groups

2and3

were given curcumin (100 and 400 mg/kg

bw/day, respectively ₎ by gavage for B weeks and were fed control diet. Group 4 was fed HFD (60% of total calories from

fat). Groups 5 and 6 received HFD together with the two doses

of curcumin, respectively. ln order to understand the molecular changes

in

liver ceIls,

we

performed

_a

nano LC-MS/MS analyses.

Results: Our results identified 539 proteins and of these, differential expression of 122were calculated to be statistically significant (P <0,05). Proteomic analysis revealed '11 _proteins

which

were

associated

with lipid

metabolism. Curcumin treatment caused changes in the expressions of three hepatic proteins such as carnitine O palmitoyltransferase 1, ATP citrate synthase and acyl CoA binding protein in HFD fed rats. The results were confirmed by western blot.

Conclusions: The most outstanding effects of the curcumin

treatment in HFD may be considered as the inhibition of hepatic lipid synthesis and the enhancement of fatty acid oxidation. Curcumin treatment

is

assumed

to

affect

liver

protein expressions. To the best of our knowiedge, this is the first study reflecting the beneficial effects of curcumin treatment in fatty

liver metabolism by performing a proteomic