1
Two xan hones and wo o ame ic (3⟶8) bi la onoids om he Came oonian medicinal
plan Allanblackia lo ibunda Oli . (Gu i e ae)
Bel Youssou G. Moun essoua,b, Joseph Tchamgouea, Jean Paul Dzoyemc, Roland T.
Tchuenguemc, F ank Su upd, Muhammad I. Choudha yb, I an R. G eene, Simeon F. Kouama,*
aDepa men o Chemis y, Highe Teache T aining College, Uni e si y o Yaoundé I, P.O. Box
47, Yaoundé, Came oon
bH.E.J. Resea ch Ins i u e o Chemis y, In e na ional Cen e o Chemical and Biological
Sciences (ICCBS), Uni e si y o Ka achi, Ka achi-75270, Pakis an
cDepa men o Biochemis y, Facul y o Science, Uni e si y o Dschang, P.O. Box 67, Dschang,
Came oon
dDepa men o Mic obial D ugs, Helmhol z Cen e o In ec ion Resea ch and Ge man Cen e
o In ec ion Resea ch, pa ne si e Hanno e /B aunschweig, Inho ens asse 7, D-31824
B aunschweig, Ge many
eDepa men o Chemis y and Polyme Science, Uni e si y o S ellenbosch, P/Bag X1,
Ma ieland, S ellenbosch, 7602, Sou h A ica
*Co esponding au ho : Tel.: +237 694 464 535; e-mail add ess: k [email protected] (S. F.
Kouam).
2
Abs ac
Two xan hones, 2-(3-hyd oxy-3,3-dime hyldihyd oallyl)-dihyd o-6-deoxyisojaca eubin
(1) and dihyd o-6-deoxyjaca eubin (2), and wo 3⟶8 o ame ic bi la onoids, (2R,3S)-
olkensi la one-7-O-𝛽-ace ylglucopy anoside (3) and (2S,3S)-mo ello la one-7-O-𝛽-
ace ylglucopy anoside (4), oge he wi h i een known compounds, we e isola ed om a
dichlo ome hane/me hanol (1/1, / ) ex ac o he ba k o he plan Allanblackia lo ibunda. The
s uc u es o he new compounds we e elucida ed by NMR spec oscopy and mass spec oscopic
echniques and hose o he known ones we e deduced by compa ison wi h da a epo ed in he
li e a u e. The isola ed bi la onoids we e ob ained as mix u es o con o me s exhibi ing
duplica e NMR signals in solu ion a 25 °C and hei espec i e absolu e con igu a ions we e
assigned using ci cula dich oism spec oscopy. Some o he isola ed compounds we e assessed
o hei an ibac e ial and an ioxidan p ope ies.
3
1. In oduc ion
Allanblackia lo ibunda Oli e belongs o he Gu i e ae amily and is widely dis ibu ed
along he coas al egions o Wes A ica. In olk medicine, di e en pa s o his plan a e ei he
used alone o in combina ion wi h o he plan s o he ea men o se e al human ailmen s
including uppe espi a o y ac in ec ions, dysen e y, dia hoea, and oo hache.1 Biological and
pha macological in es iga ions o ex ac s o his plan ha e enabled a be e unde s anding o
hei use in adi ional medicine. Fo ins ance, he an i umo , adical sca enging,
an imycobac e ial, an ibac e ial and an i ungal ac i i ies o he oo ba k ex ac o A. lo ibunda
ha e been epo ed.2 In es iga ions o his plan ha e led o he isola ion o a se ies o seconda y
me aboli es belonging o di e en classes including i e penoids,3 benzophenones,4 xan hones5
and bi la onoids.2,4 Some o hese compounds exhibi a wide ange o biological and
pha macological ac i i ies including an ioxidan , an i umo ,2 cy o oxic,5 an i-in lamma o y,
an imic obial and an i ungal ac i i ies,6,7 as well as HIV inhibi o y ac i i y.8 As pa o ou
con inuing sea ch o biologically ac i e compounds om Came oonian medicinal plan s,9–12 we
ha e in es iga ed he ba k o A. lo ibunda o i s mino seconda y me aboli es and he ein epo
he isola ion, s uc u al elucida ion and biological ac i i ies o wo new xan hones (1 and 2) and
wo new o ame ic bi la onoids (3 and 4).
2. Resul s and Discussion
The dichlo ome hane/me hanol ex ac o he s em ba k o A. lo ibunda was subjec ed o
epea ed column ch oma og aphy o gi e se e al ac ions, which we e u he pu ified o e
sephadex LH–20 and p epa a i e HPLC o yield a o al o nine een compounds o which
compounds 1–4 we e unknown.
4
3
OO
O
OH
OH
2
R1R2
H
6OH
7H
OH
O
OH
O
O
O
16
O
OH
O
O
OH OH
1
12
3
4
4a
5
6
7
8
4b
8a 9
1'
2'
3'
4''
5'
9a 1''
2''
3''
5''
4' TFA/
O
O
OH
O
O
OH
HO
OH
I-A
I-B
I-C
II-D
II-E
II-F
1
2
3
5
6
4
71'
8
9
10
2'
3'
4'
5'
6'
1
2
3
4
5
6
78
1''
2''
3''
4''
5''
6''
9
10
OH
OH
R2
O
O
OH
O
O
OH
HO
OH
I-A
I-B
I-C
II-D
II-E
II-F
OH
R1
O
OH
2''
4''
5''
6''
8'' 7''
1''
3''
4
5H OH
8
O
OH
OH
OH
OH
HO
9
O
O
OH
O
O
OH
HO
OH
OH
OH
O
HO
HO
OAc
O
OH
O
HO
HO
OH
O
OH
O
HO
HO
OH
O
OH
O
HO
HO
O
O
Figu e 1. Chemical s uc u es o compounds 1−9 and 16.
Compound 1 was ob ained as a yellow solid (m.p. 186–187 °C), which eac ed posi i ely
o FeCl3 eagen . I s molecula o mula C23H26O6 was es ablished om he posi i e ion mode
HRESIMS, which showed ion clus e s [M+H]+ a m/z 399.1802 (calcd. o C23H27O6: 399.1808).
The IR spec um showed s ong abso p ions o hyd oxyl and conjuga ed ca bonyl g oups a
3400 and 1640 cm–1 espec i ely, while he UV spec um exhibi ed cha ac e is ic abso p ion
bands o xan hones a 𝜆max 234 and 250 nm.13 The 1H NMR spec um o compound 1 displayed
an ABC spin sys em a 𝛿H 7.73 (dd, 1.8; 7.9 Hz), 7.27 (dd, 1.8; 7.9 Hz) and 7.22 (pseudo , 7.9
Hz) assignable o a 1,2,3- isubs i u ed benzene ing. The 2,2-dime hyldihyd opy an moie y was
deduced om he signals a 𝛿H 2.93 and 1.76 (2H each, , 7.9 Hz) and 𝛿H 1.37 (6H, s) which was
u he con i med by he 13C NMR spec um wi h esonances a 𝛿C 72.7, 41.3, 29.8 and 16.4.
This 2,2-dime hyldihyd opy an g oup was a ached a C-3 and C-4 o he xan hone skele on o
compound 1 as illus a ed by HMBC co ela ions (Fig. 2) obse ed be ween he me hylene
p o ons a 𝛿H 2.93 (H-1ꞌ) wi h he ca bon signals a 𝛿C 158.8 (C-3); 152.0 (C-4a); 107.9 (C-4);
72.7 (C-3′) and 41.3 (C-2′). On he o he hand, he p esence o a 3-hyd oxy-3-me hylbu yl g oup
was also deduced by he p esence o signals a 𝛿H 2.77 and 1.77 (2H each, , 7.0 Hz) and 1.40
5
(6H, s) which we e u he con i med in he 13C NMR spec um wi h esonances a 𝛿C 72.7, 41.3,
29.8 and 16.4 espec i ely. Thus, HMBC co ela ions be ween he me hylene p o ons a 𝛿H 2.77
(H-1ꞌꞌ) and he ca bon signals a 𝛿C 158.8 (C-3); 103.7 (C-2); 76.2 (C-3′′) and 31.7 (C-2′′) clea ly
iden i ied he poin o a achmen a C-2 o he xan hone skele on. Based on he abo e
in es iga ions, s uc u e 1 was named 2-(3-hyd oxy-3,3-dime hyldihyd oallyl)-dihyd o-6-
deoxyisojaca eubin (Fig. 1). The p oposed s uc u e was ully suppo ed by HMBC, DEPT and
COSY spec a. Key HMBC co ela ions o compounds 1 a e illus a ed in Fig. 2. Fu he mo e,
compound 1 was hea ed a e lux in TFA o 30 min. The esul ing p oduc , bispy anoxan hone
(16) allowed addi ional con i ma ion o he p oposed s uc u e o 1.
3
O
O
HO
HO OH
O
O
H3C
O
O
OH
O
O
OH
HO
OH
OH
H
H
O O
O
OH
OH
2
OH
O O
O
OH
OH
1
Figu e 2. Key HMBC co ela ions o compounds 1–3.

6
Table 1. 13C and 1H NMR da a o compounds 1 and 16 (CDCl3) a 25 °C.
C and H
no.
1
16
13C (176 MHz)
1H (500 MHz)
13C (176 MHz)
1H (700 MHz)
𝛿C (𝑚)
𝛿H (𝑛H, J in Hz)
𝛿C (m)
𝛿H (𝑛H, J in Hz)
1
158.2 (C)
-
154.2 (C)
-
2
103.7 (C)
6.19 (1H, s)
105.8 (C)
6.19 (1H, s)
3
158.8 (C)
-
157.8 (C)
-
4
107.9 (C)
-
99.6 (C)
-
4a
152.0 (C)
-
153.9 (C)
-
4b
144.6 (C)
-
143.5 (C)
-
5
145.5 (C)
-
144.1 (C)
-
6
119.9 (CH)
7.27 (dd, 1.8; 7.9)
119.0 (CH)
7.28 (1H, dd, 1.5, 8.0)
7
123.6 (CH)
7.22 (pseudo , 7.9)
123.8 (CH)
7.23 (1H, pseudo , 8.0)
8
116.0 (CH)
7.73 (dd, 1.8; 7.9)
117.9 (CH)
7.64 (1H, dd, 1.5, 8.0)
8a
121.0 (C)
-
106.2 (C)
-
9
181.3 (C)
-
176.1 (C)
-
9a
102.6 (C)
-
102.0 (C)
-
1
16.4 (CH2)
2.93 (2H, , 7.9)
16.7 (CH2)
2.93 (2H, , 6.7)
2
41.3 (CH2)
1.76 (2H, , 7.9)
31.7 (CH2)
1.93 (2H, , 6.7)
3
72.7 (C)
-
76.3 (C)
-
4
29.8 (CH3)
1.37 (3H, s)
26.9 (CH3)
1.42 (3H, s)
5
29.8 (CH3)
1.37 (3H, s)
26.9 (CH3)
1.42 (3H, s)
1
16.2 (CH2)
2.77 (2H, , 7.0)
17.0 (CH2)
2.65 (2H, , 6.7)
2
31.7 (CH2)
1.87 (2H, , 7.0)
31.5 (CH2)
1.84 (2H, , 6.7)
3
76.2 (C)
-
76.0 (C)
-
4
26.9 (CH3)
1.40 (3H, s)
26.6 (CH3)
1.45 (3H, s)
5
26.9 (CH3)
1.40 (3H, s)
26.6 (CH3)
1.45 (3H, s)
7
Compound 2, named dihyd o-6-deoxyjaca eubin, was ob ained as an o ange solid (m.p. 190–192
°C) which also eac ed posi i ely o FeCl3 eagen . I s molecula o mula C18H16O5 was
es ablished om he posi i e ion mode HRESIMS, which showed ion clus e s [M+H]+ a m/z
313.1072 (calcd. o C18H17O5: 313.1076). The IR spec um showed s ong abso p ions o
hyd oxyl and conjuga ed ca bonyl g oups a 3400 and 1640 cm–1 espec i ely, while he UV
spec um exhibi ed abso p ion maxima cha ac e is ic o xan hones a 𝜆max 234 and 250 nm.13 The
1H and 13C NMR spec a o compound 2 showed simila i ies wi h hose o compound 1 and
addi ionally displayed wo me hylene signals a 𝛿H 2.75 and 1.87 (2H each, , 6.7 Hz) oge he
wi h a 6-p o on single a 𝛿H 1.40 sugges ing he p esence o a 2,2-dime hyldihyd opy an moie y.
Fu he mo e, an ABC spin sys em a 𝛿H 7.78 (1H, dd, 1.5; 8.0 Hz), 7.32 (dd, 1.5; 8.0 Hz) and
7.25 (pseudo , 8.0 Hz) was sugges i e o a 1,2,3- isubs i u ed benzene ing. The 13C NMR
spec um o 2 displayed 18 ca bon signals which we e assigned in combina ion wi h DEPT and
HSQC expe imen s o wo me hyls, wo me hylenes, ou me hines and en qua e na y ca bons,
including a ca bonyl signal a 𝛿C 180.7 ppm. The p esence o a 2,2-dime hyldihyd opy an moie y
was u he con i med by signals in he 13C NMR spec um a 𝛿C 76.6 (C-3), 31.7 (C-2), 26.8
(C-4/C-5) and 16.0 (C-1). The HMBC spec um p o ed pi o al in o de o a ach his 2,2-
dime hyldihyd opy an moie y o he xan hone skele on. Thus, co ela ions be ween he
me hylene p o ons a 𝛿H 2.75 (H-1′) wi h he ca bon signals a 𝛿C 160.9 (C-1); 102.6 (C-9a); 76.6
(C-3′) and 31.7 (C-2′) sugges ed ha C-1′ (𝛿C 16.0) was a ached o C-2 (𝛿C 104.6) o he
xan hone skele on. In addi ion, co ela ions o he a oma ic p o on signal a 𝛿H 6.38 (1H, s, H-4)
wi h ca bon signals a 𝛿C 161.8 (C-3); 154.7 (C-4a); 104.6 (C-2) and 102.6 (C-9a) we e also
c ucial in con i ming he 2,2-dime hyldihyd opy an moie y being a ached a posi ions 2 and 3 o
he xan hone skele on. Mo eo e , he posi ion o he chela ed hyd oxyl (𝛿H 13.22) g oup a C-5
was assigned based on he co ela ions obse ed be ween he a oma ic p o on a 𝛿H 7.78 (1H, dd,
1.5; 8.0, H-8) wi h he ca bonyl g oup signal a 𝛿C 180.7. The p oposed s uc u e o compound 2
(Fig. 2) was ully suppo ed by HMBC, DEPT and COSY spec a. I s physical and spec oscopic
da a we e consis en wi h hose o a known syn he ic compound.14 To he bes o ou knowledge,
compound 2 is he ein epo ed o he i s ime om a na u al sou ce.
8
Table 2. 13C and 1H NMR da a o compound 2 (CDCl3) a 25 °C.
C and H no.
13C (176 MHz)
1H (500 MHz)
𝛿C (m)
𝛿H (𝑛H, J in Hz)
1
160.9 (C)
-
2
104.6 (C)
-
3
161.8 (C)
-
4
94.9 (CH)
6.38 (1H, s)
4a
154.7 (C)
-
4b
144.1 (C)
-
5
144.2 (C)
-
6
119.9 (CH)
7.32 (1H, dd, 1.5, 8.0)
7
123.7 (CH)
7.25 (1H, pseudo , 8.0)
8
116.9 (CH)
7.78 (1H, dd, 1.5, 8.0)
8a
121.1 (C)
-
9
180.7 (C)
-
9a
102.6 (C)
-
1′
16.0 (CH2)
2.75 (2H, , 6.7)
2′
31.7 (CH2)
1.87 (2H, , 6.7)
3′
79.6 (C)
-
4′
26.8 (CH3)
1.40 (3H, s)
5′
26.8 (CH3)
1.40 (3H, s)
Compound 3 was ob ained as a yellow solid, and had he molecula o mula C38H32O16 as
es ablished om low esolu ion FAB-MS (nega i e mode) and HRFAB-MS (posi i e mode)
echniques in which he molecula ions appea ed a m/z 743.3 [M–H]+ and 745.1781 [M+H]+
(calcd. o C38H33O16: 745.1769), espec i ely. The IR spec um o compound 3 showed s ong
abso p ion bands a ound 3400 and 1700 cm–1 indica ing he p esence o hyd oxyl and ca bonyl
g oups, espec i ely. Al hough he p ep. HPLC and LC-MS ch oma og ams indica ed his
compound o be pu e, bo h i s 1H and 13C NMR spec a exhibi ed doubled se s o signals,
sugges ing he p esence o wo con o me s. The 1H NMR spec um displayed cha ac e is ic
signals o a suga moie y (ace ylglucopy anosyl) wi h he anome ic p o on signal a 𝛿H 5.24 (H-
1‴) and a mul iple o non-anome ic p o ons in he up ield egion (𝛿H 3.70–3.48) along wi h a
wo-p o on iple o an oxyme hylene en i y a 𝛿H 4.36 ppm. The 13C NMR spec um displayed
signals o 38 ca bons, which we e so ed by DEPT and HSQC echniques in o one me hyl, one
me hylene, nine een me hines and se en een qua e na y ca bons, including signals o h ee
ca bonyls a 𝛿C 172.6, 184.0 and 197.3 ppm (Table 4). Compounds wi h simila spec al da a
9
ha e been p e iously epo ed om he plan s o he genus Ga cinia o he Gu i e ae amily.15–
18 The e o e, compa ison o he spec oscopic da a o compound 3 wi h hose desc ibed in he
li e a u e enabled us o deduce ha i was a monoglycosyla ed bi la onoid de i a i e p esen ing
doubled NMR signals. I is no ewo hy ha he doubled signals in he NMR spec a o
bi la onoids a oom empe a u e may be due o he ac ha he molecules adop di e en
con o ma ions16 which a ise om he es ic ed ee o a ion o he 3→8 in e la anyl bond.17
Ne e heless, hese NMR signals can be me ged in o a single se o signals a highe
empe a u es because unde hese condi ions, he molecules do no exis in a p e e ed s able
con o ma ion.19 Howe e , he s uc u al elucida ion o compound 3 based on i s NMR da a
eco ded a ambien empe a u e was no s aigh o wa d.
In he 1H NMR spec um, excep o a pai o double s a 𝛿H 5.96 and 5.97 (1H each, d,
1.6 Hz, H-6, H-8) assignable o wo me a-coupled p o ons o ing I-A o he la anone moie y o
bo h he majo and mino con o me s, he 1H NMR esonances o he majo con o me we e
mos ly obse ed in he deshielded a oma ic egion (𝛿H 6.40–7.80) whe eas, signals appea ing in
he up- ield egions we e assigned o he suga moie y. The 1H NMR spec um o compound 3
also displayed cha ac e is ic signals o la ones a 𝛿H 6.45 (1H, s, H-II-3) and 𝛿H 5.71 and 5.33
(1H each, d, 12.0 Hz, H-I-2, H-I-3) indica ing he p esence o bo h moie ies in he molecule. The
coupling cons an o 12.0 Hz indica es a biaxial ( ans) con igu a ion o he wo p o ons o ing
I-C. In addi ion, cha ac e is ic signals o wo sepa a e AA'BB' sys ems we e obse ed a 𝛿H 7.73
and 6.91 (2H each, d, 8.8 Hz, H-3″/5″, H-2″/6″) o one sys em and 𝛿H 7.02 and 6.34 (2H each,
d, 8.4 Hz, H-2′/6′, H-3′/5′) assignable o he wo 1,4-disubs i u ed a oma ic ing sys ems o he
la one ( ing II-E) and la anone ( ing I-B) moie y, espec i ely. The co ela ions o hese se s o
p o ons iz. H-2″/6″ wi h H-3″/5″and H-2′/6′wi h H-3′/5′ we e ully suppo ed by hei 1H–1H
COSY co ela ions (see ESI) bo h o he majo and he mino con o me s. A one p o on single
assignable o H-6 o each o he la one uni s was e iden in he 1H NMR spec um.
In e es ingly, signals a ibu able o an ace yl g oup we e obse ed in he 1H and 13C NMR
spec a a 𝛿H/C 1.95/20.7 (CH3) and 172.7 (CH3C=O). The comple e assignmen o he wo
con o me s o compound 3, as p esen ed in Tables 3 and 4 espec i ely, was acili a ed by he
HSQC and HMBC spec a. The ace ylglucopy anosyl moie y was linked a posi ion C-7 o he
la one uni and deduced by co ela ions obse ed in he HMBC spec um be ween he anome ic
p o on a 𝛿H 5.24 wi h ca bon signals a 𝛿C 161.6 (C-7) and 𝛿C 100.0 (C-6) o ing D (Fig. 2).
16
Silica gel 60 (0.230–0.400 mm) and (0.040–0.063 mm) was used as adso ben s o lash
and column ch oma og aphy espec i ely. The semi-pu e compounds we e inally pu i ied
successi ely o e Sephadex LH-20 (bead size 25–100 μm, Sigma-Ald ich) wi h CH2Cl2–MeOH
(1/1, / ), e e se (JAIGEL–ODS H80, se ial no. 209963) and no mal (JAIGEL–SIL, D-60-10,
se ial no. 051300228) phase p epa a i e HPLC. The speci ica ions o hese columns we e 250
mm leng h×20 mm inne diame e ; 4𝜇m pa icle size and 80 Å po e size. Me ck TLC pla es
(silica gel 60 F254) we e used o he de ec ion o he pu i y o compounds. Mel ing poin s we e
de e mined wi h Gallekamp appa a us. UV spec a we e eco ded using a Pe kinElme Lambda
25 UV/Vis spec ome e . IR spec a we e eco ded in KB on a Pe kinElme 2000 FT-IR
spec opho ome e . EI/ESI mass spec a we e eco ded on an Agilen 5975C MSD and The mo
Finnigan MAT95XL mass spec ome e s a he Hussein Eb ahim Jamal Resea ch Ins i u e o
Chemis y (HEJ–RIC) o he In e na ional Cen e o Chemical and Biological Sciences
(ICCBS), Uni e si y o Ka achi (UOK), Ka achi Pakis an. The NMR expe imen s o pu e
compounds we e pe o med in di e en deu e a ed sol en s depending on hei solubili y, using
B uke Ascend 400, 500 and 600 MHz (1H and 13C) spec ome e s equipped wi h a B uke 5 mm
B oadband p obe (depending on he amoun ). Chemical shi s (𝛿) in ppm a e e e enced o
e ame hylsilane (TMS) a 0.00 ppm o 1H and 13C. Coupling cons an s a e exp essed in he z
(Hz).
4.2. Plan ma e ial
The s em ba k o A. lo ibunda Oli e was collec ed in No embe 2014 in he Kye-Ossi,
N em alley Di ision in he Sou h egion o Came oon. The sample was iden i ied by M . Vic o
Nana, a e i ed bo anis a he Na ional He ba ium o Came oun, Cen e egion, whe e a ouche
specimen was deposi ed as ouche No: 52904/HNC.
4.3. Ex ac ion and isola ion
The ai -d ied and g ound s em ba k (2.2 kg) o A. lo ibunda was ex ac ed a oom
empe a u e wi h a mix u e o CH2Cl2/MeOH (1/1, / ) o 48 h and concen a ed o a iscous
black esidue (556 g). A pa o his esidue (550.0 g) was hen subjec ed o lash
ch oma og aphic sepa a ion o e a silica gel (230–400 mesh) column using a s epwise g adien
o 𝑛-Hex/E OAc ( anging om 0 o 100% o E OAc, / ), ollowed by a g adien o
E OAc/MeOH ( anging om 9/1 o 8/2, / ), o a o d a o al o 58 ac ions (𝑓 1–𝑓 58) o ca.

17
1000 mL pe ac ion. F ac ion 𝑓 15 (elu ed wi h 𝑛-Hex/E OAc 9/1) p ecipi a ed o gi e
compound 10 (100 mg, yellow solid). Fu he p ecipi a ion o he emaining aliquo o his same
ac ion ga e an insepa able mix u e o phy os e ols (180 mg, whi e powde ). F ac ion 𝑓 22 (𝑛-
Hex/E OAc 8/2, / ) p ecipi a ed o a o d compound 12 (6 mg, yellow solid). F ac ions 𝑓 48,
𝑓 50, 𝑓 51 and 𝑓 53 ob ained wi h pu e E OAc, we e mass-p ecipi a ed o gi e ou di e en
yellow amo phous powde s labelled Y5 (0.5 g), Y6 (3.4 g), Y7 (2.1 g) and Y8 (0.4 g) espec i ely.
All ou impu e powde s we e subsequen ly sepa a ely pu i ied o e e e se phase p epa a i e
HPLC (column JAIGEL–ODS H80) using a mix u e o ace oni ile/wa e : 1/1 + 0.08% o TFA
as eluen , wi h he ollowing se ings: UV sensi i i y 0.05; RI sensi i i y 50; ini ial & inal low
a es: 3 & 4 mL/min; ini ial & inal p essu es: 47 & 64 psi. Consequen ly, compounds 5 (75 mg),
6 (100 mg), 7 (25 mg) and 8 (105 mg) we e elu ed wi h e en ion imes o 22, 18, 14 and 12 min
espec i ely. All 58 ac ions (𝑓 1–𝑓 58) om he lash ch oma og aphy we e combined in o 4
main ac ions (A–D) on he basis o hei TLC analyses. F ac ion A (𝑓 1–𝑓 7: 850 mg) ob ained
wi h pu e 𝑛-Hex as eluen , consis ed o a y acids and was no in es iga ed u he . F ac ion B
(𝑓 8–𝑓 33: 38.7 g) ob ained wi h 𝑛-Hex/E OAc (8/2–4/6, / ), was subjec ed o u he column
ch oma og aphy o e silica gel (0.040–0.063 mm) and elu ed wi h a g adien o 𝑛-Hex/E OAc
(9.5/0.5–0/10, / ) o p oduce 100 ac ions (B1–B100) o ca. 500 mL each which we e combined
on he basis o TLC analysis. The i s 28 ac ions om ac ion B (B1–B28), we e elu ed wi h a
mix u e o 𝑛-Hex/E OAc (9.5/0.5, / ) and combined on he basis o hei TLC p o iles in o wo
sub- ac ions. The i s sub- ac ion b15–b20 (200 mg) was ech oma og aphed o e Sephadex
LH-20 (CH2Cl2/MeOH (1/1, / ) and hen o e no mal phase p ep. HPLC, using an isoc a ic
mode sol en o 𝑛-Hex/E OAc (8.8/1.2, / ), o a o d compound 14 (2.5 mg, o ange oil) wi h a
e en ion ime o 44 min. The second sub- ac ion b21–b28 (150 mg) showed a complex mix u e
o oils and was no u he in es iga ed. The second se ies o ac ions om ac ion B (B29-B40)
was elu ed wi h 𝑛-Hex/E OAc (9/1, / ) and combined, hen was u he ch oma og aphed and
elu ed on no mal phase p epa a i e HPLC, using an isoc a ic mode o sol en 𝑛-Hex/E OAc
(8.5/1.5, / ) o a o d compound 2 (0.7 mg, o ange solid), a a e en ion ime o 20 min. The
hi d se ies o ac ions om ac ion B (B41-B43) (30 mg) was elu ed wi h 𝑛-Hex/E OAc (8/2,
/ ), hen ech oma og aphed and elu ed on no mal phase p epa a i e HPLC, using an isoc a ic
mode o sol en 𝑛-Hex/E OAc (7/3, / ) o a o d compound 1 (12 mg, yellow solid), a a
e en ion ime o 25 min. The ou h se ies o ac ions om ac ion B (B44-B45) (88 mg) was
18
ech oma og aphed o e Sephadex LH-20 (CH2Cl2/MeOH (1/1, / ), o a o d compound 11 (18
mg, yellow solid). The i h se ies o ac ions om ac ion B (B52–B59) (10 mg) was u he
ch oma og aphed and elu ed on no mal phase p epa a i e HPLC, using an isoc a ic mode o
sol en 𝑛-Hex/E OAc (7.5/2.5, / ) o a o d compound 13 (1.8 mg, yellow solid), a a e en ion
ime o 44 min. F om he six h se ies o ac ions om ac ion B (B60–B77), a whi e solid
compound 15 (60 mg, whi e nea solid) was ob ained. The las se ies o ac ions ob ained om
ac ion B (B78–B100) elu ed wi h 𝑛-Hex/E OAc (7/3–2/8, / ) was ound o be a complex
mix u e o compounds and was no u he in es iga ed.
F ac ion C (𝑓 34–𝑓 57: 137 g), elu ed wi h 𝑛-Hex/E OAc 3/7–1/9, / ) om he lash
ch oma og aphy was shown o con ain he al eady pu i ied compounds 5; 6 and 8 and he e o e
was no u he in es iga ed.
A pa o ac ion D (𝑓 58: 80 g; pu e E OAc) was subjec ed o u he ch oma og aphic
sepa a ions o e sephadex LH-20 using CH2Cl2/MeOH (1/1) as eluen o a o d eigh een sub-
ac ions (D1–D18). Sub- ac ion D9 (1.8 g) was subjec ed o silica gel column ch oma og aphy
and elu ed wi h an isoc a ic sys em o 𝑛-Hex/E OAc (6/4, / ) o a o d a mix u e o
glycosyla ed s e ols (105 mg). Sub- ac ions D5 (150 mg), D16 (200 mg) and D17 (300 mg) we e
sepa a ely pu i ied on a e e se phase p ep. HPLC using isoc a ic sol en s o a o d compounds
9 (5 mg, o ange oil), 3 (15 mg, yellow solid), and 4 (18.8 mg, yellow solid) espec i ely.
4.3.1. 2-(3-hyd oxy-3,3-dime hyldihyd oallyl)-dihyd o-6-deoxyisojaca eubin (1). Yellow solid
om Hex/E OAc 7/3, m.p. 186–187 °C; UV (MeOH) – 𝜆max nm (PDA): 234, 250, 269, 330 nm;
HRESIMS m/z 399.1802 (calcd. o C23H27O6: 399.1808). Fo 1H and 13C NMR da a, see Table
1.
4.3.2. dihyd o-6-deoxyjaca eubin (2). O ange solid om Hex/E OAc 8.5/1.5, m.p. 190–192 °C;
UV (MeOH) – 𝜆max nm (PDA): 234, 250, 269, 330 nm; HRESIMS [M+H]+ a m/z 313.1072
(calcd. o C18H17O5: 313.1076). Fo 1H and 13C NMR da a, see Table 2.
4.3.3. (2R,3S)- olkensi la one-7-O-𝛽-ace ylglucopy anoside (3). Yellow solid om E OAc
ac ion; m.p. 241–243 °C; [𝛼]25D = 0° (c = 0.067, MeOH); UV (MeOH) – 𝜆max nm (log ℰ): 229
(3.91), 292 (3.80), 324 (3.73); IR (KB ): 𝜈max = 3418, 2927, 1728, 1645, 1604, 1510, 1451, 1370,
1243, 1172, 1082, 834, 742, 622, 526 cm–1; HRFESIMS m/z 745.1781 [M+H]+ (calcd. o
C38H33O16: 745.1769). Fo 1H and 13C NMR da a, see Tables 3 & 4.
19
4.3.4. (2S,3S)-mo ello la one-7-O-𝛽-ace ylglucopy anoside (4). Yellow solid om E OAc; m.p.
245–247 °C; [𝛼]20D = 0° (c = 0.046, MeOH); UV (MeOH) – 𝜆max nm (log ℰ): 222 (4.31), 230
(4.39), 287 (4.36), 291 (4.36), 344 (4.18); IR (KB ): 𝜈max = 3384, 1726, 1643, 1603, 1515, 1452,
1369, 1262, 1169, 1082, 834, 741, 630, 559, 526 cm–1; HRESIMS m/z 761.1732 [M+H]+ (calcd.
o C38H33O17: 761.1718). Fo 1H and 13C NMR da a, see Tables 3 and 4.
4.3.5. Con e sion o (1) o (16)
Compound 1 (10 mg) was dissol ed in i luo oace ic acid (2 mL) and was hea ed unde
e lux o 30 min. The esul ing mix u e was sepa a ed and pu i ied by p ep. HPLC wi h 𝑛-
Hex/E OAc (7/3, / ) o a o d compound 16 (7.4 mg, 74%), a a e en ion ime o 22 min.
Compound 16 was ob ained as a pale yellow oil and iden i ied as 1,5-dihyd oxy-1,2,3,4-bis(2,2-
dime hyldihyd opy ano)xan hone. HRESIMS [M+H]+ a m/z 381.1702 (calcd. o C23H25O5:
381.1702). Fo 1H and 13C NMR da a, see Table 1.
4.3.6. Assays o an ibac e ial and an ioxidan ac i i ies
The an ibac e ial ac i i y was e alua ed using he b o h mic odilu ion me hod by de e mining he
minimum inhibi o y concen a ion (MIC) alues agains he i e bac e ia s ains iz.,
Esche ichia coli (ATCC 25922); Pseudomonas ae uginosa (ATCC 27853); S aphylococcus
au eus (ATCC BAA1026); En e ococcus aecalis (ATCC 29212); P o eus mi abilis (isola e).
The an ioxidan capaci y o he compounds was e alua ed based on he p inciple o sca enging
he DPPH (2,2-diphenyl-1-pic ylhyd azyl) adical.
Acknowledgemen s
BYGM is g a e ul o The Wo ld Academy o Sciences (TWAS) o an eigh -mon h ellowship
unde he auspices o he ICCBS-TWAS p og amme o wo k a he H.E.J. Resea ch Ins i u e o
Chemis y, Uni e si y o Ka achi, Ka achi-75270, Pakis an. This s udy was also suppo ed by
he Alexande on Humbold Founda ion and he Ge man Academic Exchange Se ice (DAAD)
h ough he equipmen subsidies o SFK.
Con lic o In e es
The au ho s decla e no con lic o in e es .
20
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