Study On The Chemical Constituents Of The Mid-polar Part Of Curcuma Aromatica Salisb And Quantification Of The Sesquiterpenoids

Objective: To study the chemical constituents in the mid-polar part of Curcuma aromatica Salisb.To establish an ultra-performance liquid chromatography-mass spectrometry(UPLC-MS)method for the determination of sesquiterpenoids in C.Aromatica.Methods:The rhizomes of C.Aromatica were crushed into a oedema powder and heated under reflux with 70% aqueous ethanol.The extract was dissolved in distilled water and the solution was extracted with petroleum ether,ethyl acetate(EtOAc)and n-butanol(n /BuOH)saturated with water.The EtOAc fraction was separated and purified by silica gel column chromatography,Sephadex LH-20 column chromatography,ODS column chromatography,PTLC,half preparative HPLC,recrystallization,and so on.Their structures were identified by physicochemical means,spectral analysis,and comparison of NMR spectral data with those reported in the literature.To determine the content of the five compounds of curcumol(CAW-2),gweicurculactone(CAW-3),12-hydroxycurcumenol(CAW-6),aromaticain A(CAL-1),aerugidiol(CAW-7)in C.Aromatica by UPLC-MS.The determination was carried out by a Waters ACQUITY UPLC? HSS T3 column(2.1 mm×100 mm ID,1.8 ?m)with a gradient mobile phase of methanol(A)-water(addition of 5 mmol/L ammonium formate in aqueous phase)(B)solution(0~5.5 min,40% A;5.5~8.5 min,40% A?65% A;8.5~12.5 min,65% A;12.5~15.5 min,65% A?80% A;15.5~18.5 min,80% A?95% A).The flow rate was 0.3 mL/min.The column temperature was 40 ? and the injection volume was 10 ?L.We used the multi-reaction monitoring(MRM)scanning as detector.curcumol(CAW-2),gweicurculactone(CAW-3),12-hydroxycurcume nol(CAW-6),aromaticain A(CAL-1)was collected under the ESI positive ion modes.aerugidiol(CAW-7)was collected under the ESI negaitive ion modes.Results:Sixteen compounds were obtained from the mid-polar part of C.aromatica and twelve compounds were identified.The twelve compounds are ?-sitosterol(CAW-1),curcumol(CAW-2),gweicurculactone(CAW-3),methylzedoarondiol(CAW-5),12-hydroxycurcumenol(CAW-6),aerugidiol(CAW-7),2-hydroxygweicurculactone(CAW-8),zedoalactone B(CAW-11),wenyujinepoxide A(CAW-12),zedoalactone A(CAW-13),zedoalactone C(CAW-14),zedoarolide B(CAW-19).compounds of 12-hydroxycurcumenol(CAW-6),2-hydroxygweicurculactone(CAW-8),zedoalactone B(CAW-11),wenyujinepoxide A(CAW-12),zedoalactone A(CAW-13),zedoalactone C(CAW-14),zedoarolide B(CAW-19)are isolated from this plant for the first time.Five sesquiterpenoids,curcumol(CAW-2),gweicurculactone(CAW-3),12-hydroxycurcumenol(CAW-6),aromaticain A(CAL-1),aerugidiol(CAW-7)in C.Aromatica can be quantitatively analyzed within 18.5 min.The experimental results were satisfactory for the intra-day and inter-day precision and accuracy of the method for simultaneous.The linear range of curcumol was60~6000 ng/mL(r = 0.9994),The linear range of gweicurculactone was 0.8~80ng/mL(r = 0.9998),The linear range of 12-hydroxycurcumenol was 52~5200ng/mL(r = 0.9997),The linear range of aromaticain A was 340~17000 ng/mL(r= 0.9998),The linear range of aerugidiol was 236~11800 ng/mL(r = 0.9997).The recoveries of high,middle and low concentration of curcumol were 99.7%,100.1%,99.0% with RSDs were 0.69%,0.34%,0.62%,respectively.The recoveries of high,middle and low concentration of gweicurculactone were97.0%,96.8%,100.0% with RSDs were 0.42%,0.77%,0.14%,respectively.The recoveries of high,middle and low concentration of 12-hydroxycurcumenol were 100.8%,97.9%,99.0% with RSDs were 0.74%,2.71%,0.58%,respectively.The recoveries of high,middle and low concentration of aromaticain A were 99.4%,99.9%,100.3% with RSDs were 0.11%,0.36%,0.47%,respectively.The recoveries of high,middle and low concentration of aerugidiol were 99.7%,100.5%,100.2% with RSDs were 0.29%,0.83%,0.38%,respectively.The limits of detection of curcumol,gweicurculactone,12-hydroxycurcumenol,aromaticain A and aerugidiol were 0.04 ng/mL,2.40pg/mL,0.37 ng/mL,0.33 ng/mL,0.50 ng/mL,respectively.The limits of quantification of them were 0.15 ng/mL,0.01 ng/mL,1.24 ng/mL,1.09 ng/mL,1.66 ng/mL,respectively.This chromatographic method is reliable.Conclusions: Sixteen compounds are isolated from the mid-polar part of C.aromatica and twelve compounds of them are identified,seven compounds are isolated from this plant for the first time.An accurate and sensitive method isdeveloped for the determination of the sesquiterpenoids in C.Aromatica.