Effect of combination of extracts of ginseng and ginkgo biloba on acetylcholine in amyloid beta-protein-treated rats determined by an improved HPLC.
OBJECTIVE
To determine the concentration of acetylcholine (ACh) in amyloid beta-protein (Abeta) treated rats and offer a method determining ACh as well.


METHODS
A 1-month combination of extrats of ginseng and ginkgo biloba (Naoweikang) ig administration to rats was performed daily after bilateral injection of Abeta(1-40) (4 g/L, 1 microL for each side) into hippocampus. After decollation, homogenizing, and centrifuging and extracting, a high pressure liquid chromatographic (HPLC) method using electrochemical detection (ECD) combined with two immobilized enzyme reactors was used to determine ACh in rat whole brain.


RESULTS
With a mobile phase consisting of disodium hydrogen orthophosphate, tetramethylammonium chloride (TMACl), octanesulfonic acid sodium salt (OSA) and "Reagent MB" at a final pH of 8.0, ACh was determined while removing the interfering choline in less than 10 min at a flow rate of 0.35 mL/min on a platinum (Pt) working electrode at a potential of +300 mV vs a solid-state palladium (Pd) reference electrode. Linear regression analysis of peak area vs concentration demonstrated linearity in the 28.01 to 1400.06 microg/L injection range. The r-value was 0.9978. The limit of detection (LOD) is 0.28 ng on column. ACh in whole brain decreased by 20.34 % (from 162.1+/-32.7 to 134.7+/-14.0 microg/L, P<0.05) after bilateral injection of Abeta into rat hippocampus. After Naoweikang administration (31 and 15.5 mg/kg, respectively), ACh increased by 19.97 % (from 134.7+/-14.0 to 161.6+/-26.2 microg/L, P<0.05) and 18.56 % (from 134.7+/-14.0 to 159.7+/-22.9 microg/L, P<0.05), respectively.


CONCLUSIONS
Naoweikang significantly increased the level of ACh in whole brain of Abeta treated rats. And a sensitive, selective and reliable method for routinely determining ACh in rat whole brain was established in this study.
Liu
Jian-Xun
JX
Research Center, Xiyuan Hospital, China Academy of Traditional Chinese Medicine, Beijing 100091, China.
Cong
Wei-Hong
WH
Xu
Li
L
Wang
Jian-Nong
JN
D000109:Acetylcholine / Q000378:metabolism*; D016229:Amyloid beta-Peptides; D000818:Animals; D001921:Brain / Q000378:metabolism*; D001930:Brain Injuries / Q000139:chemically induced / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004338:Drug Combinations; D004365:Drugs, Chinese Herbal / Q000302:isolation & purification / Q000494:pharmacology*; D020441:Ginkgo biloba / Q000737:chemistry; D008297:Male; D005894:Panax / Q000737:chemistry; D051381:Rats; D017207:Rats, Sprague-Dawley
D016229:Amyloid beta-Peptides; D004338:Drug Combinations; D004365:Drugs, Chinese Herbal; D000109:Acetylcholine