Data on LC–MS profile of Brucea javanica (L.) Merr. and the NMR data of its major indole alkaloids

This article presents two types of phytochemical data obtained from Brucea javanica (L.) Merr. roots, a medicinal plant belonging to the Simaroubaceae family. The high-resolution LC–MS dataset comprised the chemical profile of dichloromethane extract, which was utilised to annotate 35 chemical constituents. For annotations, the measured spectral data were compared with the in-silico spectral data generated from 920 molecular structures previously reported in Simaroubaceae. Indole alkaloids, quassinoids, aliphatics and lignan were the chemical groups identified in the root extract. The second dataset provides NMR spectra (1H, 13C, COSY, HMQC and HMBC) for the six indole alkaloids previously detected in LC–MS analysis and isolated through centrifugal partition chromatography. The chemical structures of all compounds were confirmed based on NMR data as bruceolline J (compound 7), canthin-6-one-N-oxide (compound 10), bruceolline E (compound 15), 5-methoxycanthin-6-one (compound 16), canthin-6-one (compound 20), and 1‑hydroxy-11-methoxycanthin-6-one (compound 22). This phytochemical data was generated to support an ongoing anti-cancer and anti-dengue study.


a b s t r a c t
This article presents two types of phytochemical data obtained from Brucea javanica (L.) Merr.roots, a medicinal plant belonging to the Simaroubaceae family.The highresolution LC-MS dataset comprised the chemical profile of dichloromethane extract, which was utilised to annotate 35 chemical constituents.For annotations, the measured spectral data were compared with the in-silico spectral data generated from 920 molecular structures previously reported in Simaroubaceae.Indole alkaloids, quassinoids, aliphatics and lignan were the chemical groups identified in the root extract.The second dataset provides NMR spectra ( 1 H, 13 C, COSY, HMQC and HMBC) for the six indole alkaloids previously detected in LC-MS analysis and isolated through centrifugal partition chromatography.The chemical structures of all compounds were confirmed based on NMR data as bruceolline J (compound 7 ), canthin-6-one-N -oxide (com-pound 10 ), bruceolline E (compound 15 ), 5-methoxycanthin-6-one (compound 16 ), canthin-6-one (compound 20 ), and 1hydroxy-11-methoxycanthin-6-one (compound 22 ).This phytochemical data was generated to support an ongoing anticancer and anti-dengue study.
© • The reported chemical constituents could be a valuable resource for chemical marker selection during the development of standardised extract and quality control.• The NMR spectral data provide updated references and spectra for indole alkaloids, which is useful in elucidating structurally related indole alkaloids.

Objective
B. javanica has been widely studied for various bioactivities including anti-cancer, antiplasmodial, anti-inflammatory, and anti-viral [1][2][3][4] .While many phytochemical data described the seed extract [ 5 , 6 ], this article aimed to provide comprehensive and updated references for the chemical constituents in root extract.

Plant collection, authentication, and extraction
The roots of B. javanica, known in Malay as "melada pahit" were harvested in August 2019 from Jitra, Kedah, Malaysia.The plant was authenticated by a botanist from Forest Research In-  stitute Malaysia (FRIM) as Brucea javanica (L.) Merr (Voucher number: PID 461119-24).The dried roots were ground into powder (1 kg), defatted with hexane and extracted with DCM successively for 24 hr each.This step was repeated 3 times with fresh solvent with an approximately 8 L solvent used.The extract yield was 4.68 g (0.50 % yield, wt/wt).

Chemical profiling of Brucea javanica roots extract
Chemical profiling of the DCM extract was obtained from LC-MS.Briefly, 1 μL of the crude extract (5 mg/mL in methanol) was flowed (0.3 mL/min) through ACQUITY UPLC ® HSS T3 1.8 μm, 2.1 mm X 100 mm column (Waters Corporation, Ireland).Throughout analysis, column and sample compartment temperatures were maintained at 40 °C and 10 °C, respectively.The mobile phase comprised 0.1 % formic acid in water (A) and 0.1 % formic acid in acetonitrile (B).The LC gradient was as follows: 5 % B for 2 min; 5 % B -100 % B from 2-30 min; 100 % B from 30 to 35 min; 100 % B to 5 % B from 35 to 35.5 min; and finally, 4.5 min column equilibration with 5 % B.
The high-resolution mass spectrometer was operated at 70,0 0 0 orbitrap resolutions with a maximum IT of 250 ms for full scan.Data-dependent MS 2 (dd-MS 2 ) events were performed on the top five most abundant ions detected in the MS full-scan.The MS 2 data was obtained at 17,0 0 0 resolutions with maximum IT of 60 ms.The normalised stepped collision energy (NCE) was set to 15, 30, and 45 V. Positive and negative ionisation modes were applied to acquire a comprehensive list of chemical compounds.The optimised HESI parameters were set as follows: spray voltage, 4.0 kV (PI) and 3.5 kV (NI); capillary temperature, 350 °C; auxiliary gas flow rate, 11 L/min; sheath gas flow, 4 kV; S-lens RF level, 60 %.The mass range was obtained from m/z 100 to 1500.

LC-MS data acquisition and metabolite annotation
The raw scans of Q Exactive TM Orbitrap Mass Spectrometer data were imported into Compound Discoverer TM 3.1 software (CD) (Thermo Fisher Scientific, Waltham, MA, USA) for preprocessing.The parameters for spectra selection, alignment of the retention time nodes, compound detections, compound grouping to specify the preferred adducts and blank removal  1 H and (B) 13 CNMR spectra of bruceolline J ( 7 ).were optimised accordingly.B. javanica metabolite annotation was predefined using "Untargeted Metabolomics with Statistics Detect Unknowns with ID using Online Databases and mz-Logic" workflow template.A customised database containing 920 compounds previously reported from Simaroubaceae family was retrieved from Dictionary of Natural Product (DNP) and loaded into CD as "mass lists".The mass lists include chemical names, molecular formulas, molecular weights, and chemical structures.

Centrifugal partition chromatography isolation procedure
Indole alkaloids isolation was performed using a centrifugal partition chromatography CPC-250 system (Armen Instrument, Saint-Ave, France) fitted with a 250 mL hydrostatic column and UV/VIS detector.A ternary biphasic system consisting of a mixture of hexane:ethyl acetate:methanol:water, 1:2:1:2 (Arizona K) was prepared to obtain two immiscible liquid phases; upper and lower phases.By selecting ascending as the elution mode, the column was loaded with the aqueous stationary phase (lower phase) at a flow rate of 30 mL/min with 600 rpm rotation speed for 10 min.Subsequently, the organic mobile phase (upper phase) was pumped into the column at 10 mL/min with 1500 rpm rotation speed for 20 min to achieve column equilibration.The DCM extract (83 mg/mL in a mixture of upper and lower phase (5:1, v:v )) was then injected through the injection loop and monitored at UV 254 and 320 nm.During the 90 min elution, fractions were collected in every 10 mL/ tube.All 65 fractions were screened by HPTLC and fractions containing the same component were combined to afford 5 fractions (FrA, FrB, FrC, FrD and FrE).Further purification of these fractions was performed using JAI LC-200NEXT preparative HPLC system (Japan Analytical Industry, Co., Ltd, Meguro Tokyo, Japan).The purification of fraction FrA on a silica column (JAIGEL-SIL, SH-043-15 (150 × 21.2 mm)) and an isocratic solvent system (DCM:methanol (97:3)) yielded compounds 7 and 10 .For FrC and FrD purification, isocratic mobile phase of DCM:methanol (98:2) was applied.This purification yielded compounds 15, 16 and 20, 22, respectively.

Characterisation of indole alkaloids by NMR
The one-dimensional ( 1 H, 13 C) and two-dimensional (COSY, HMQC, HMBC) NMR analyses were recorded on a JNM-ECZ 600 MHz NMR spectrometer (JEOL Ltd., Tokyo, Japan) system.This system is equipped with a 5-mm digital auto-tune Royal probe in a variable temperature (VT) at 298 K and controlled by JEOL Delta NMR Software. 1 H NMR analysis was carried out with 16 scans, 5 s delay, spectral width −2.0 to 15 ppm, spectral resolution 0.5496 Hz, data point 16,384, and receiver gains at 50.The 13 C NMR analysis was carried out with more than 10 0,0 0 scans (depending on samples), 2 s delay, 1.1514 Hz spectral resolution, and 32,768 data point.The receiver gain was adjusted automatically for each sample before acquisition to avoid receiver overload.The default JEOL pulse sequence methods were used for all 2D NMR experiments.Data for 1 H NMR are reported as follows: chemical shift ( δ ppm), multiplicity (singlet (s), doublet (d), triplet (t), quartet (q), quintet (p), multiplet (m), doublet of doublets (dd), doublet of triplets (dt), broad (br)), and coupling constant ( J in Hz).The chemical shift for 13 C NMR data is reported in ppm.All spectra were referenced using solvent peak.The NMR data were processed in MestReNova ver.14 software (Mestrelab Research, SL, Spain).The structures of all compounds were determined based on 1 H, 13 C NMR, HMQC, HMBC, and COSY data.In addition, the proposed structures were verified by two approaches: comparison with reported values [ 1 , 8-13 ] and verification with > 95 % confidence using ACD/Labs NMR Structure elucidator (ACD/Labs, Ontario, Canada).

Fig. 1 .
Fig. 1. ( A ) PI and NI base peak chromatograms obtained from LC-MS analysis of B. javanica roots extract and B ) the zoom-in chromatograms at 8-14 min.The numbered peaks are corresponding to Table1.All unlabelled peaks are unknown.
2023 The Author(s).Published by Elsevier Inc.This is an open access article under the CC BY license ( http://creativecommons.org/licenses/by/4.0/ ) Mass Spectroscopy (LC-MS) profile of the dichloromethane (DCM) extract obtained from B. javanica was recorded on UPLC system coupled with Orbitrap Q Exactive TM Mass Spectrometer.A full scan with data-dependent MS 2 (dd-MS 2 ) acquisitions was applied to obtain the LC-MS data.
For compound isolation, the centrifugal partition chromatography (CPC) was fitted with 250 mL column volume rotor and UV-Vis detector.The 1 H,13C and 2D (COSY, HMQC, HMBC) NMR spectra for all compounds were recorded on JNM-ECZ 600 MHz NMR spectrometer.Data format Raw and analysed Description of data collection The roots of Brucea javanica (L.) Merr.(Simaroubaceae) were harvested from Jitra, Kedah, Malaysia at Latitude: 6 °15 15.9732 N and Longitude: 100 °26 7.6 84 8 E. Plant authentication was performed by Forest Research Institute Malaysia (Voucher number: PID 461119-24).An LC gradient elution of 40 min total run time in positive and negative HESI modes was used to acquire two LC-MS profiles of the dichloromethane extract.The raw data was processed using Compound Discoverer TM ver.3.1 software.The same software annotated 35 of the most intense peaks through in-silico spectral matching.For this purpose, a customised compound database containing 920 molecular structures was generated from the online Dictionary of Natural Products Database (Keyword: Simaroubaceae).All compounds isolated through a CPC procedure were used as reference standards to confirm the identification of 6 peaks (from the 35 peaks).The solvent system utilised for isolation consisted of hexane:ethyl acetate:methanol:water (1:2:1:2; v/v/v/v) in ascending mode.The structures of six indole alkaloids were determined based on 1D ( 1 H,13C) and 2D (HMBC, HMQC, COSY) NMR and by comparing the spectral data with reported values.

Table 1
Metabolite annotations of indole alkaloids, quassinoids, lignans and aliphatics in DCM extract of B. javanica roots by LC-MS analysis.

Table 2 1
H NMR spectroscopic data of all isolated compounds (