Total synthesis of acylphloroglucinols and their antibacterial activities against clinical isolates of multi-drug resistant (MDR) and methicillin-resistant strains of Staphylococcus aureus

Article abstract of DOI:10.1016/j.ejmech.2018.05.038

Bioassay-directed drug discovery efforts focusing on various species of the genus Hypericum led to the discovery of a number of new acylphloroglucinols including (S,E)-1-(2-((3,7-dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)-2-methylbutan-1-one (6, olympicin A) from H. olympicum, with MICs ranging from 0.5 to 1 mg/L against a series of clinical isolates of multi-drug-resistant (MDR) and methicillin-resistant Staphylococcus aureus (MRSA) strains. The promising activity and interesting chemistry of olympicin A prompted us to carry out the total synthesis of 6 and a series of analogues in order to assess their structure-activity profile as a new group of antibacterial agents. Following the synthesis of 6 and structurally-related acylphloroglucinols 7–15 and 18–24, their antibacterial activities against a panel of S. aureus strains were evaluated. The presence of an alkyloxy group consisting of 8–10 carbon atoms ortho to a five-carbon acyl substituent on the phloroglucinol core are important structural features for promising anti-staphylococcal activity.

Full text of DOI:10.1016/j.ejmech.2018.05.038

Accepted Manuscript
Total synthesis of acylphloroglucinols and their antibacterial activities against
clinical isolates of multi-drug resistant (MDR) and methicillin-resistant strains of
Staphylococcus aureus
M. Mukhlesur Rahman, Winnie K.P. Shiu, Simon Gibbons, John P. Malkinson
PII:
S0223-5234(18)30451-3
10.1016/j.ejmech.2018.05.038
EJMECH 10449
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 29 March 2018
Revised Date: 22 May 2018
Accepted Date: 24 May 2018
Please cite this article as: M.M. Rahman, W.K.P. Shiu, S. Gibbons, J.P. Malkinson, Total synthesis of
acylphloroglucinols and their antibacterial activities against clinical isolates of multi-drug resistant (MDR)
and methicillin-resistant strains of Staphylococcus aureus, European Journal of Medicinal Chemistry
(2018), doi: 10.1016/j.ejmech.2018.05.038.
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ACCEPTED MANUSCRIPT
Graphical Abstract
A series of new structurally-related acylphloroglucinols were synthesized and
screened for their antibacterial activities against a panel of multi-drug resistant (MDR) and
methicillin-resistant Staphylococcus aureus (MRSA) strains.
Antibacterial activity
HO
O
O
12 showed strongest anti-
staphylococcal activity
(MIC 0.25-1 mg/L)
OH
6
HO
OR
O
HO
O
O
HO
OH
R
OH
OH
OH
6-15
18-24

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Total synthesis of acylphloroglucinols and their antibacterial activities
against clinical isolates of multi-drug resistant (MDR) and methicillin-
resistant strains of Staphylococcus aureus
M. Mukhlesur Rahman,a,b Winnie K. P. Shiu,a,c Simon Gibbonsa and John P. Malkinsona,*
aResearch Department of Pharmaceutical and Biological Chemistry, UCL School of
Pharmacy, 29-39 Brunswick Square, London WC1N 1AX, UK
bMedicine Research Group, School of Health, Sport and Bioscience, University of East
London, Stratford Campus, Water Lane, London E15 4LZ, UK
cPresent Address: 16 Royal Swan Quarter, Leret Way, Leatherhead, Surrey KT22 7JL, UK
____________
*Corresponding author.
E-mail address: j.malkinson@ucl.ac.uk (J. P. Malkinson)

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ABSTRACT
Bioassay-directed drug discovery efforts focusing on various species of the genus
Hypericum led to the discovery of a number of new acylphloroglucinols including (S,E)-1-(2-
((3,7-dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)-2-methylbutan-1-one
(6,
olympicin A) from H. olympicum, with MICs ranging from 0.5 to 1 mg/L against a series of
clinical isolates of multi-drug-resistant (MDR) and methicillin-resistant Staphylococcus
aureus (MRSA) strains. The promising activity and interesting chemistry of olympicin A
prompted us to carry out the total synthesis of 6 and a series of analogues in order to assess
their structure-activity profile as a new group of antibacterial agents. Following the synthesis
of 6 and structurally-related acylphloroglucinols 7-15 and 18-24, their antibacterial activities
against a panel of S. aureus strains were evaluated. The presence of an alkyloxy group
consisting of 8-10 carbon atoms ortho to a five-carbon acyl substituent on the phloroglucinol
core are important structural features for promising anti-staphylococcal activity.
Key words: Acylphloroglucinols; Total synthesis; Anti-staphylococcal; MRSA

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1. Introduction
In the era of antibiotic resistance, natural products notably from plants, microbes and
marine organisms continue to be an important source of lead compounds for drug discovery.
Since the discovery of penicillin from Penicillium notatum, researchers have focused on
natural resources, mostly microorganisms, for effective and safe antibiotics. Although plants
have been well documented for their production of biologically-active metabolites including
anticancer agents such as paclitaxel (Taxol®) [1], antimalarial drugs (artemisinin) [2],
narcotic analgesics (morphine) [3] and cardioactive agents such as digoxin [4], this area has
been under-exploited for antimicrobial drug discovery. However, there are numerous reports
of plants being used as systemic and topical antimicrobial agents in Ayurvedic [5] and
Traditional Chinese Medicine [6] as well as in western herbal medicine [7] because of their
self-protection strategy to counter bacteria and fungi in their own environment [8].
Due to the burgeoning global problem of anti-microbial resistance (AMR), there is an
increasing need for new chemistries to supplement the dwindling antibiotic arsenal [9]. St
John’s Wort (Hypericum perforatum), a medicinal plant used widely as an anti-depressant in
herbal medicine, has been reported to produce hyperforin, an antibacterial metabolite with a
minimum inhibitory concentration (MIC) value of 0.1 mg/L against methicillin-resistant
Staphylococcus aureus (MRSA) and penicillin-resistant variants [10]. Our bioassay-directed
drug discovery efforts to identify potent anti-staphylococcal agents from various species of
the genus Hypericum led to the discovery of a number of acylphloroglucinols including (S,E)-
1-(2-((3,7-dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)-2-methylbutan-1-one (6)
(trivial name olympicin A) from H. olympicum [11] with MIC values of 0.5 to 1 mg/L against
a series of MRSA strains. The potential activity and the complex but interesting chemistry of
such acylphloroglucinol compounds prompted us to carry out the total synthesis of olympicin
A and its related analogues in order to assess their structure-activity profile as a new group of

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antibacterial agents. Here we report the total synthesis of olympicin A (6) and a series of
acylphloroglucinols (7-22) that are structurally related to 6, as well as their antibacterial
activities against a panel of multi-drug-resistant (MDR) and methicillin-resistant
Staphylococcus aureus strains.
2. Results and discussion
2.1. Synthesis of (S)-(2-methylbutanoyl)phloroglucinols with ortho alkyloxy variants
(S,E)-1-(2-((3,7-Dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)-2-
methylbutan-1-one (6) (trivial name olympicin A),
a
promising antibacterial
acylphloroglucinol isolated from H. olympicum, and its analogues with variable ortho
alkyloxy substituents were synthesised in four steps starting from (S)-2-methylbutanoic acid
(1) (Scheme 1) with typical overall yields of 10-12%. This general approach involved
Friedel-Crafts acylation of phloroglucinol (3) with (S)-2-methylbutanoyl chloride (2) to give
the intermediate ketone (S)-2-methyl-1-(2,4,6-trihydroxyphenyl)butan-1-one (4), which was
regioselectively protected as the bis-silyl derivative 5 prior to alkylation of the remaining free
phenolic hydroxyl group (and simultaneous deprotection). The structures of all intermediate
compounds were confirmed unambiguously by NMR spectral data.
In the first step, commercially available (S)-2-methylbutanoic acid (1) was treated
with thionyl chloride to form the corresponding acid chloride (S)-2-methylbutanoyl chloride
(2; 93% yield), which was isolated by distillation. The boiling point of 2 was 119-120 °C,
which was in good agreement with the literature [12]. The structural identity of 2 was
confirmed by mass spectrometry, and 1H and 13C NMR spectroscopy (section 4.2.1).
Friedel-Crafts acylation of phloroglucinol (3) with (S)-2-methylbutanoyl chloride (2)
in the presence of AlCl3, CS2 and nitrobenzene led to the formation of (S)-2-methyl-1-(2,4,6-
trihydroxyphenyl)butan-1-one (4; 54% yield), which was purified by VLC. Unreacted
phloroglucinol and nitrobenzene were easily separated from 4 during purification and

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successful acylation was verified by mass spectrometry and NMR spectral data (section
4.2.2). In addition to the presence of the signals that were typical for a 2-methylbutanoyl
substituent, the 1H NMR spectrum showed a singlet at δH 5.81, integrating for two equivalent
meta aromatic hydrogen atoms, consistent with the plane of symmetry in 4.
Initial efforts to protect two (para and one ortho) of the three phenolic hydroxyl
groups in 4 utilised methoxymethyl (MOM) ether protecting groups. Isolated yields were
generally poor (below 20%) and this approach was ultimately abandoned as the acidic
conditions required for MOM ether deprotection also resulted in cleavage of the required
geranyl ether in the target compound 6. The use of silyl ethers for the desired bis-protection
proved more successful. Compound 4 was treated with TBDMS-Cl (2.1 molar equivalents)
under basic reaction conditions, forming (S)-1-(2,4-bis((tert-butyldimethylsilyl)oxy)-6-
hydroxyphenyl)-2-methylbutan-1-one (5) in an improved 81% yield, after purification by
VLC over silica gel. Small quantities of mono-protected derivatives were also identified in
the reaction mixture. The presence of a hydrogen-bonded hydroxyl (δH 13.43) and two meta-
coupled (J = 2.0 Hz) hydrogen atoms with two different chemical shifts (δH 5.85 and 6.04)
confirmed the protection of the hydroxyl groups with TBDMS at one ortho and the para
position. Intramolecular hydrogen bonding between the hydrogen of the one remaining free
phenolic hydroxyl and the carbonyl oxygen of the acyl substituent, as well as its lower steric
accessibility, are likely to account for the excellent regioselectivity of this bis-silyl ether
protection.
The final step in the synthesis of the (S)-2-methylbutanoylphloroglucinol series
involved the alkylation of the free ortho phenolic hydroxyl of 5 using the appropriate alkyl
bromide. Treatment of a solution of (S)-1-(2,4-bis((tert-butyldimethylsilyl)oxy)-6-
hydroxyphenyl)-2-methylbutan-1-one (5) with geranyl bromide in the presence of K2CO3
yielded
(S,E)-1-(2-((3,7-dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)-2-

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methylbutan-1-one (6; typically 9-16% yield), which was purified by VLC over silica gel.
The original intention had been to alkylate the free ortho phenolic hydroxyl followed by silyl
ether deprotection in a subsequent step, but the conditions used in the alkylation unexpectedly
led to simultaneous TBDMS group removal. While this suggested the possibility of direct
alkylation (geranylation) of unprotected 4, attempts to achieve this generally resulted in very
poor conversion (less than 5% yields) of starting acylphloroglucinols to typically ca. 1:1
mixtures of ortho and para mono-alkylated products, which were difficult to separate from
unreacted starting materials.
The NMR data of 6 were identical to those of the new acylphloroglucinol we reported
from H. olympicum [11]. The alkyl (geranyl) substituent included an oxymethylene group (δH
4.56 d, J = 6.5 Hz, geranyl C-1), the hydrogen atoms of which showed long-range (3J) 1H-13C
correlations with a deshielded aromatic carbon atom (
acylphloroglucinol nucleus and two carbons associated with an olefinic group (
geranyl C-2; 142.4, geranyl C-3). Together with the non-equivalence (asymmetry) of the
δ
162.6, aromatic C-2) of the
δ
118.2,
δ
aromatic C-H signals, this confirmed successful ortho O-alkylation. In a similar manner, a
series of other ortho alkyloxy acylphloroglucinols (7-15, Figure 1) was synthesised, purified
and characterised. The final ether substituents were varied to include both saturated and
unsaturated linear and branched alkyl/alkenyl groups such as prenyl (7), farnesyl (8), 3,7-
dimethyloctyl (reduced geranyl; 9), 3-methylbutyl (reduced prenyl; 10), benzyl (11), octyl
(12), decyl (13), dodecyl (14) and octadecyl (15) groups. In each analogue, the C-1
oxymethylene hydrogen atoms of the alkyl substituent demonstrated key HMBC interactions
with the aromatic C-2 (ortho) carbon atom of the acylphloroglucinol core, allowing
confirmation of the successful desired ortho alkylation in compounds 7-15. The yield of the
final alkylation/deprotection step was typically quite low at around 30% (though much lower

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for 8 and 10). Isolated by-products often included recovered 4, along with small quantities of
bis-alkylated (ortho plus para) and occasionally para alkylated derivatives.
2.2.
Synthesis of ortho geranylated acylphloroglucinols with acyl variants
The acyl chloride used in the initial Friedel-Crafts acylation of phloroglucinol was
varied to generate, after elaboration as described above, a series of geranylated
acylphloroglucinol analogues (Figure 2), incorporating simple linear acyl substituents such as
propanoyl (19), butanoyl (20), pentanoyl (21), decanoyl (22) and octadecanoyl (23) as well as
aromatic acyl substituents such as benzoyl (24). Commercially available acetylphloroglucinol
was also used to synthesise ortho geranylated acetylphloroglucinol (18). Regioselective di-
protection of two (para and one ortho) of the three hydroxyl groups of the acylphloroglucinol
(16) moiety again using TBDMS-Cl, followed by alkylation with geranyl bromide and
simultaneous deprotection of the silyl ethers (17) yielded the corresponding ortho geranylated
acylphloroglucinols 18-24 (Scheme 2). Final products were purified by SPE over silica gel
and confirmed by NMR spectroscopy. Again, a key aspect in the confirmation of the
structures of each of the compounds was the use of 2D-HMBC, in which the C-1
3
oxymethylene hydrogen atoms of the geranyl substituent demonstrated J correlations with
the C-2 (ortho) carbon of the acylphloroglucinol.
2.3. Antibacterial activity of acylphloroglucinols
The antibacterial activities (Table 1) of the acylphloroglucinol series were assessed
against a panel of clinical isolates of multidrug-resistant (MDR) and methicillin-resistant
Staphylococcus aureus (MRSA), notably SA1199B, XU212, RN4220, EMRSA15 and
EMSA16. These organisms represent a group of effluxing strains that are resistant to
common antibiotics including certain fluoroquinolones (SA1199B), tetracycline (XU212) and
macrolides (erythromycin; RN4220). These strains were chosen as antibacterials that are not
substrates for these MDR pumps are clinically desirable. It was evident from Table 1 that the

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(S)-2-methylbutanoylphloroglucinol derivatives with geranyl (6; the natural product
olympicin A) and 3,7-dimethyloctyl (reduced geranyl; 9) ether substituents at the ortho
position were found to be highly active with MIC values of 0.5-1 mg/L. The effect on activity
of replacing the geranyl substituent with shorter and longer alkyl/alkenyl groups (prenyl,
farnesyl, 3-methylbutyl, 3,7-dimethyloctyl) can also be observed. We found that increasing
the chain length from geranyl (6) to farnesyl (8) resulted in a dramatic loss of activity,
evidenced by an increase in MIC values from 0.5-1 mg/L to in excess of 512 mg/L (no
inhibition of the growth of any of the strains of bacteria was seen at the highest concentration
of 8 tested), while decreasing the chain length from geranyl to prenyl (7) showed at least a
two-fold reduction in activity. These results suggested an optimal length for the alkyloxy
group in terms of antibacterial activity, which may imply a specific molecular target or a
requirement for a particular lipophilicity for the mechanism of action. In addition, the 3-
methylbutyl (10) and 3,7-dimethyloctyl (9) derivatives were as active as the prenyl or geranyl
derivatives respectively, suggesting that the presence of the double bonds in the alkyl side
chain (and the consequent reduction in conformational freedom) might not have any
significant role in antibacterial activity. The most potent compound in terms of its activity
against the six S. aureus strains was 12, an (S)-(2-methylbutanoyl)phloroglucinol with a
straight chain octyl ether substituent – the same continuous linear chain length as the geranyl
and 3,7-dimethyloctyl derivatives – at the ortho position phenolic hydroxyl.
Among the ortho geranylated acylphloroglucinol series, the pentanoyl derivative
(21) demonstrated the highest activity with MICs of 0.5-1 mg/L, making it comparable in
activity to the parent natural product 6. The octadecanoyl analogue (23) did not show any
inhibition of bacterial growth at any of the concentrations tested, indicating that there was
clearly an optimal chain length (number of carbon atoms) for the acyl substituent.

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Geranylated analogues containing three to seven carbon atoms in their acyl substituents
(aliphatic or aromatic) seem be most active.
3. Conclusion
The acylphloroglucinol natural product olympicin A was synthesised from
commercial phloroglucinol over three straightforward steps (Friedel-Crafts acylation,
regioselective silyl protection, and simultaneous deprotection/O-alkylation). The same
approach provided access to a series of ortho alkyloxy and acyl analogues. Evaluation of the
activities of the analogues against several strains of MRSA seemed to indicate that the
presence of an alkyloxy group consisting of 8-10 carbon atoms ortho to a five-carbon-atom
acyl substituent is an important structural feature for promising anti-staphylococcal activity
within this class of acylphloroglucinol antibiotics. The most potent derivative 12 – the octyl
ether analogue of olympicin A – demonstrated improved activity over the natural product
against all MRSA strains tested. The most promising acylphloroglucinols identified in this
study are being evaluated for their clinical potential as systemic and topical antibacterial
agents. A more diverse range of structural analogues, including those incorporating side-
chain functionality, is also being synthesised in an effort to better understand the structure-
activity relationships of this class of agents. At present we do not fully understand how these
compounds function. However, given that the most active compounds all have a hydrophobic
portion, comprised of an ether and an acyl functionality, capable of membrane interaction,
and a hydrophilic diphenolic moiety, it is probable that their target is cell-wall located.
4. Experimental section
4.1. General methods
All chemicals and reagents used for the syntheses were purchased from Sigma-
Aldrich, Gillingham, UK. UV spectra were recorded on a Thermo Electron Corporation
Helios spectrophotometer and IR spectra were recorded on a Nicolet 360 FT-IR

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spectrophotometer. NMR spectra (both 1D and 2D) were recorded on a Bruker AVANCE
500 MHz spectrometer (500 MHz for 1H and 125 MHz for 13C). Chemical shift values (
δ) are
reported in parts per million (ppm) and are calibrated relative to residual solvent peaks as
internal standards, and coupling constants (J values) are expressed in Hz. Detailed
assignments of NMR spectra can be found in the Supporting Information. Mass spectra were
recorded on a Finnigan MAT 95 high resolution, double focusing, magnetic sector mass
spectrometer. Accurate mass measurement was achieved using voltage scanning of the
accelerating voltage. This was nominally 5 kV and an internal reference of heptacosa was
used. Resolution was set between 5000 and 10000. Both TLC and preparative TLC were
performed using silica gel 60 PF254 plates (Merck). Vacuum liquid chromatography (VLC)
columns were packed with silica gel 60 PF254 (Merck), while solid phase extraction (SPE)
was performed using 10 g pre-packed SPE columns (SiGel60) using mobile phases of hexane
and ethyl acetate of increasing polarity.
4.2. Synthesis of intermediate and final compounds
4.2.1. Synthesis of (S)-2-methylbutanoyl chloride (2) [12]
(S)-2-Methylbutanoic acid (1; 10 g, 97.91 mmol) and thionyl chloride (10.71 mL,
146.9 mmol, 1.5 equiv) were heated together at 80-90 °C under reflux for 2 h. Distillation of
the reaction mixture afforded (S)-2-methylbutanoyl chloride as a colourless liquid (10.89 g,
22
D
1
90.72 mmol, 93%).
[
α
]
+10.1 (c 0.54, CHCl3); b.p. 119-120 °C; H NMR (500 MHz,
CDCl3): δH 0.95 (3H, t, J = 7.5 Hz), 1.26 (3H, d, J = 7.5 Hz), 1.59 (1H, m), 1.80 (1H, m),
2.80 (1H, q, J = 7.0 Hz); 13C NMR (125 MHz, CDCl3): δC 11.3, 16.7, 26.7, 53.1, 177.8;
HRMS (ESI) m/z: [M-H]- Calcd for C5H8OCl 119.0264; Found 119.0263.
4.2.2. Synthesis of (S)-2-methyl-1-(2,4,6-trihydroxyphenyl)butan-1-one (4) [13]
Phloroglucinol (3; 10.81 g, 85.8 mmol) in carbon disulphide (50 mL) was transferred
into a two-necked round-bottomed flask and allowed to stir while aluminium trichloride

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(46.43 g, 351.8 mmol, 4.1 equiv) was added. Nitrobenzene (40 mL) was then added to the
solution over 30 min. The solution was then heated under reflux at 55 °C for 30 min. A
solution of 2-methylbutanoyl chloride (2) (10.89 g, 85.8 mmol) dissolved in 5 mL
nitrobenzene was added to the reaction mixture over 30 min, followed by heating for another
30 min. The reaction mixture was allowed to cool with stirring and then poured into an ice-
water bath (400 mL). 100 mL of 3
M hydrochloric acid was then added and the mixture
extracted with diethyl ether (3 × 500 mL). The organic solvents were removed under reduced
pressure. The oily residue containing the acylphloroglucinol was subjected to VLC over
SiGel PF254 using hexane and EtOAc of increasing polarity. The VLC fraction eluted with
30-45% EtOAc in hexane gave (S)-2-methyl-1-(2,4,6-trihydroxyphenyl)butan-1-one as a pale
22
D
yellow oil (9.71 g, 46.19 mmol, 54%).
[
α
]
+8.5 (c 0.35, CHCl3); UV (CHCl3) λmax (log ε):
1
240 (4.17), 290 (3.97) nm; IR νmax (thin film) cm-1: 3297, 1628, 1602, 1222; H NMR (500
MHz, CD3OD): δH 0.88 (3H, t, J = 7.5 Hz), 1.11 (3H, d, J = 6.5 Hz), 1.33 (1H, m), 1.78 (1H,
m), 3.84 (1H, m), 5.81 (2H, s); 13C NMR (125 MHz, CD3OD): δC 12.4, 17.2, 28.2, 46.7, 96.0,
96.0, 105.3, 165.8, 165.8, 211.4; HRMS (ESI) m/z: [M-H]- Calcd. for C11H13O4 209.0814;
Found 209.0813.
4.2.3. Synthesis
of
(S)-1-(2,4-bis((tert-butyldimethylsilyl)oxy)-6-hydroxyphenyl)-2-
methylbutan-1-one (5) [14]
Acylphloroglucinol 4 (9.71 g, 46.19 mmol) was dissolved in part in dry acetone (150
mL) and transferred into a 250 mL round bottom flask. Imidazole (3.43 g, 138.6 mmol, 3
equiv) was added to the solution and the reaction mixture stirred for 5 min followed by the
addition of TBDMS-Cl (14.61 g, 97.0 mmol, 2.1 equiv). The reaction mixture was stirred for
2 h at room temperature. Acetone was removed from the reaction mixture under reduced
pressure and the residue taken up in chloroform and washed with 1
M HCl (150 mL). The
organic layer was dried using anhydrous magnesium sulphate, filtered and the solvent was

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removed under reduced pressure. The crude product purified by VLC over silica gel using
hexane and EtOAc of increasing polarity. VLC fractions eluted with 2.5-7.5% EtOAc in
hexane
afforded
(S)-1-(2,4-bis((tert-butyldimethylsilyl)oxy)-6-hydroxyphenyl)-2-
22
D
methylbutan-1-one as a pale yellow oil (16.4 g, 37.26 mmol, 81%).
[
α
]
+4.9 (c 0.39,
CHCl3); UV (CHCl3) λmax (log
ε
): 239 (4.26), 290 (4.23) nm; IR νmax (thin film) cm-1: 3276,
1
2973, 1688, 1572, 1531,1256, 1131, 1072, 850; H NMR (500 MHz, CDCl3): δH 0.23 (2 ×
3H, s), 0.32 (2 × 3H, s), 0.88 (3H, t, J = 7.5 Hz), 0.97 (3 × 3H, s), 0.99 (3 × 3H, s), 1.12 (3H,
d, J = 6.5Hz), 1.43 (1H, m), 1.78 (1H, m), 3.82 (1H, m), 5.85 (1H, d, J = 2.0 Hz), 6.04 (1H,
13
d, J = 2.0 Hz), 13.43 (1H, s); C NMR (125 MHz, CDCl3): δC -4.2, -3.7, 11.0, 16.9, 18.1,
18.9, 25.5, 26.5, 26.1, 45.0, 102.0, 103.1, 108.4, 158.8, 161.7, 166.5, 210.5; HRM (ESI) m/z:
[M-H]- Calcd. for C23H41O4Si2 437.2549; Found 437.2554.
4.2.4. Synthesis of (S,E)-1-(2-((3,7-dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxy-phenyl)-
2-methylbutan-1-one (olympicin A) (6)
TBDMS-protected acylphloroglucinol 5 (6.6 g, 15.0 mmol) was dissolved in dry
DMF (100 mL) and anhydrous potassium carbonate (3.1 g, 22.5 mmol, 1.5 equiv) was added.
The mixture was stirred for approximately 5 min followed by the addition of geranyl bromide
(3.43 mL, 18 mmol, 1.2 equiv). The mixture was heated at 80 °C for 3 h with stirring. The
reaction mixture was poured over water and extracted with chloroform. The solvent in the
organic layer was removed under reduced pressure. The crude product was purified by
chromatography over silica gel by VLC. Compound 6 was eluted with 9:1 hexane-ethyl
acetate and removal of the solvents under reduced pressure yielded the title compound as a
pale yellow oil (450 mg, 1.3 mmol, 8.7%). All spectral data were identical to those of the
22
D
natural product [11].
[
α
]
+6.0 (c 0.30, CHCl3); UV (CHCl3) λmax (log
ε
): 239 (4.06), 240
(4.23) nm; IR νmax (thin film) cm-1: 3357, 2965, 2931, 1623, 1589, 1458, 1212, 1165, 1087,
825; 1H NMR (500 MHz, CDCl3): δH 0.88 (3H, t, J = 7.5 Hz), 1.12 (3H, d, J = 6.5 Hz), 1.36

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(1H, m), 1.61 (3H, s), 1.69 (3H, s), 1.74 (3H, s), 1.79 (1H, m), 2.10 (2H, m), 2.13 (2H, m),
3.68 (1H, m), 4.56 (2H, d, J = 6.5 Hz), 5.10 (1H, m), 5.44 (1H, br s), 5.50 (1H, m), 5.91 (1H,
d, J = 2.0 Hz), 5.98 (1H, d, J = 2.0 Hz), 14.10 (1H, s); 13C NMR (125 MHz, CDCl3): δC 11.9,
16.5, 16.6, 17.7, 25.7, 26.2, 26.9, 39.5, 46.2, 65.6, 91.5, 96.5, 105.9, 118.2, 122.6, 132.0,
142.4, 161.9, 162.6, 167.5, 210.4; HRMS (ESI) m/z: [M-H]- Calcd. for C21H29O4 345.2071;
Found 345.2067.
4.2.5. Synthesis of compounds 7-15 from TBDMS-protected acylphloroglucinol 5
Compounds 7-15 were synthesised from TBDMS-protected acylphloroglucinol 5
using the procedure as described for 6 above but replacing the geranyl bromide with the
appropriate alkyl bromide.
4.2.5.1. (S)-1-(2,4-Dihydroxy-6-((3-methylbut-2-en-1-yl)oxy)phenyl)-2-methylbutan-1-one (7)
Synthesised using 0.259 mmol of 5 and 0.311 mmol of prenyl bromide. Isolated after
22
D
SPE (SiGel; hexane/EtOAc 9:1) as a pale yellow oil (15 mg, 0.054 mmol, 21%).
(c 0.30, MeOH); UV (MeOH) λmax (log
[
α
]
+5.2
ε
): 239 (4.36), 290 (3.97) nm; IR νmax (thin film) cm-
1: 3423, 2968, 2914, 1630, 1595, 1560, 1420, 1350, 1200; 1H NMR (500 MHz, CDCl3): δH
0.88 (3H, t, J = 7.5 Hz), 1.13 (3H, d, J = 6.5 Hz), 1.33 (1H, m), 1.76 (3H, s), 1.78 (1H, m),
1.82 (3H, s), 3.66 (1H, m), 4.55 (2H, d, J = 6.5 Hz), 5.51 (1H, t, J = 6.5 Hz), 5.70 (1H, br s),
13
5.93 (1H, d, J = 2.0 Hz), 6.00 (1H, d, J = 2.0 Hz), 14.07 (1H, br s); C NMR (125 MHz,
CDCl3): δC 12.0, 16.7, 18.4, 25.9, 27.1, 46.3, 65.8, 91.7, 96.7, 106.1, 118.6, 139.3, 162.3,
162.8, 167.7, 210.6; HRMS (ESI) m/z: [M-H]- Calcd. for C16H21O4 277.1445; Found
277.1453.
4.2.5.2. (S)-1-(2,4-dihydroxy-6-(((2E,6E)-3,7,11-trimethyldodeca-2,6,10-trien-1-
yl)oxy)phenyl)-2-methylbutan-1-one (8)
Synthesised using 0.925 mmol of 5 and 1.11 mmol of farnesyl bromide. Isolated after
22
D
SPE (SiGel; hexane/EtOAc 92:8) as a yellow oil (25 mg, 0.060 mmol, 6.5%).
[
α
]
+6.7 (c

ACCEPTED MANUSCRIPT
0.51, MeOH); UV (MeOH) λmax (log
ε
): 239 (4.09), 285 (4.22) nm; IR νmax (thin film) cm-1:
314, 1529, 1442, 1292, 1166, 1074, 821; 1H NMR (500 MHz, CDCl3): δH 0.90 (3H, t, J = 7.5
Hz), 1.13 (3H, d, J = 6.5 Hz), 1.37 (1H, m), 1.61 (3H, s), 1.62 (3H, s), 1.69 (3H, s), 1.75 (3H,
s), 1.80 (1H, m), 1.99 (2H, s), 2.07 (2H, s), 2.11 (2H, m), 2.13 (2H, m), 3.67 (1H, m), 4.57
(2H, d, J = 6.5 Hz), 5.08 (1H, m), 5.12 (1H, m), 5.52 (1H, m), 5.93 (1H, d, J = 2.5 Hz), 5.99
13
(1H, d, J = 2.5 Hz), 14.05 (1H, s); C NMR (125 MHz, CDCl3): δC 12.1, 16.2, 16.8, 16.9,
17.9, 25.9, 26.5, 26.9, 27.0, 39.7, 39.9, 46.3, 65.9, 91.8, 96.7, 106.1, 118.4, 123.7, 124.5,
131.6, 135.9, 142.7, 162.3, 162.8, 167.7, 210.6; HRMS (ESI) m/z: [M-H]- Calcd. for
C26H37O4 413.2698; Found 413.2697.
4.2.5.3.(2S)-1-(2-((3,7-Dimethyloctyl)oxy)-4,6-dihydroxyphenyl)-2-methylbutan-1-one (9)
Synthesised using 1.073 mmol of 5 and 1.289 mmol of 1-bromo-3,7-dimethyloctane.
Isolated after SPE (SiGel; hexane/EtOAc 9:1) as a colourless oil (110 mg, 0.313 mmol, 29%).
[
α
]
22 +4.4 (c 0.40, CHCl3); UV (CHCl3) λmax (log
ε
): 239 (3.99), 290 (4.21) nm; IR νmax (thin
D
1
film) cm-1: 3233, 2964, 2921, 1661, 1583, 1432, 1241, 1103, 1057; H NMR (CDCl3): δH
0.88 (3H, t, J = 7.5 Hz), 0.89 (6H, d, J = 6.5 Hz), 0.97 (3H, d, J = 6.5 Hz), 1.15 (3H, d, J =
6.5 Hz), 1.18 (2H, m), 1.19 (1H, m), 1.35 (3H, m), 1.43 (1H, m), 1.54 (1H, m), 1.66 (3H, m),
1.82 (1H, m), 1.89 (1H, m), 3.72 (1H, m), 4.04 (2H, d, J = 6.5 Hz), 5.36 (1H, br s), 5.93 (1H,
d, J = 2.0 Hz), 5.99 (1H, d, J = 2.0 Hz), 14.05 (1H, s); 13C NMR (CDCl3): δC 11.7, 16.8, 19.6,
22.6, 24.6, 26.6, 28.0, 29.9, 36.0, 37.5, 39.2, 46.0, 66.7, 91.5, 96.5, 105.8, 162.4, 162.8,
167.4, 210.4; HRMS (ESI) m/z: [M+H]+ Calcd. for C21H35O4 351.2531; Found 351.2529.
4.2.5.4.(S)-1-(2,4-Dihydroxy-6-(isopentyloxy)phenyl)-2-methylbutan-1-one (10)
Synthesised using 0.579 mmol of 5 and 0.695 mmol of 1-bromo-3-dimethylbutane.
Isolated after SPE (SiGel; hexane/EtOAc 9:1) as a yellow oil (5 mg, 0.018 mmol, 3.1%).
[
α
]
22 +3.5 (c 0.34, MeOH); UV (MeOH) λmax (log
ε
): 242 (4.39), 288 (4.09) nm; IR νmax (thin
D
film) cm-1: 333, 1631, 1593, 1258, 1110; 1H NMR (500 MHz, CDCl3): δH 0.89 (3H, t, J = 7.5

ACCEPTED MANUSCRIPT
Hz), 0.98 (6H, d, J = 6.5 Hz), 1.14 (3H, d, J = 6.5 Hz), 1.41 (1H, m), 1.73 (2H, m), 1.80 (1H,
m), 1.81 (1H, m), 3.69 (1H, m), 4.02 (2H, d, J = 6.5 Hz), 5.40 (1H, br s), 5.92 (1H, d, J = 2.0
Hz), 5.99 (1H, d, J = 2.0 Hz), 14.17 (1H, s); 13C NMR (125 MHz, CDCl3): δC 11.7, 16.8, 22.5
(2C), 25.2, 26.6, 37.7, 46.0, 67.5, 91.5, 96.5, 105.8, 162.4, 162.8, 167.4, 210.4; HRMS (ESI)
m/z: [M-H]- Calcd. for C16H23O4 279.1602; Found 279.1598.
4.2.5.5.(S)-1-(2-(Benzyloxy)-4,6-dihydroxyphenyl)-2-methylbutan-1-one (11)
Synthesised using 0.934 mmol of 5 and 1.121 mmol of benzyl bromide. Isolated after
SPE (SiGel; hexane/EtOAc 8:2) as a pale yellow oil (85 mg, 0.283 mmol, 30%).
[
α
]
22 +4.2 (c
D
0.32, MeOH); UV (MeOH) λmax (log
ε
): 241 (4.30), 289 (3.94) nm; IR νmax (thin film) cm-1:
1
3239, 2958, 1624, 1568, 1500, 1456, 1383, 1255, 1161, 1105, 751; H NMR (500 MHz,
CDCl3): δH 0.68 (3H, t, J = 6.5 Hz), 1.00 (3H, d, J = 6.5 Hz), 1.29 (1H, m), 1.71 (1H, m),
3.57 (1H, m), 5.08 (2H, s), 6.00 (1H, d, J = 2.0 Hz), 6.01 (1H, d, J = 2.0 Hz), 7.38-7.42 (5H,
13
m), 14.00 (1H, s); C NMR (125 MHz, CDCl3): δC 11.7, 16.8, 26.9, 46.2, 71.6, 92.0, 97.1,
106.1, 128.4 (2C), 128.8, 129.0 (2C), 135.6, 162.2, 162.5, 167.8, 210.6; HRMS (ESI) m/z:
[M+H]+ Calcd. for C18H21O4 301.1440; Found 301.1430.
4.2.5.6.(S)-1-(2,4-Dihydroxy-6-(octyloxy)phenyl)-2-methylbutan-1-one (12)
Synthesised using 0.893 mmol of 5 and 1.931 mmol of 1-bromooctane. Isolated after
SPE (SiGel; hexane/EtOAc 92:8) as a pale yellow oil (82 mg, 0.255 mmol, 29%);
[
α
22 +6.1
D
]
(c 0.45, MeOH); UV (MeOH) λmax (log
ε
): 239 (4.39), 288 (3.98) nm; IR νmax (thin film) cm-
1
1: 3328, 2928, 2857, 1621, 1591, 1444, 1377, 1212, 1158, 1102, 826, 754; H NMR (500
MHz, CDCl3): δH 0.89 (3H, t, J = 7.5 Hz), 0.90 (3H, t, J = 7.5 Hz), 1.16 (3H, d, J = 7.0 Hz),
1.26-1.40 (8H, m), 1.38 (1H, m), 1.46 (2H, m), 1.81 m (1H, m), 1.85 (2H, m), 3.72 (1H, q, J
= 6.5 Hz), 4.00 (2H, t, J = 6.5 Hz), 5.42 (1H, br s), 5.91 (1H, d, J = 2.0 Hz), 5.99 (1H, d, J =
13
2.0 Hz), 14.06 (1H, s); C NMR (125 MHz, CDCl3): δC 12.0, 14.3, 17.1, 22.8, 26.5, 26.9,

ACCEPTED MANUSCRIPT
29.3, 29.4, 29.5, 32.0, 46.3, 69.4, 91.5, 96.8, 106.2, 162.2, 163.0, 167.8, 210.5; HRMS (ESI)
m/z: [M+H]+ Calcd. for C19H31O4 323.2222; Found 323.2211.
4.2.5.7.(S)-1-(2-(Decyloxy)-4,6-dihydroxyphenyl)-2-methylbutan-1-one (13)
Synthesised using 0.936 mmol of 5 and 1.123 mmol of 1-bromodecane. Isolated after
SPE (SiGel; hexane/EtOAc 9:1) as a yellow oil (96 mg, 0.264 mmol, 28%).
[
α
]
22 +4.5 (c
D
0.25, MeOH); UV (MeOH) λmax (log
ε
): 243 (4.33), 292 (3.99) nm; IR νmax (thin film) cm-1:
3327, 2926, 2855, 1739, 1621, 1592, 1443, 1368, 1216, 1156, 1103, 827, 752; 1H NMR (500
MHz, CDCl3): δH 0.88 (3H, t, J = 7.5 Hz), 0.89 (3H, t, J = 7.5 Hz), 1.17 (3H, d, J = 6.5 Hz),
1.28-1.31 (12H, m), 1.34 (1H, m), 1.46 (2H, m), 1.81 (1H, m), 1.84 (2H, m), 3.72 (1H, q, J =
6.5 Hz), 4.00 (2H, t, J = 6.5 Hz), 5.92 (1H, d, J = 2.0 Hz), 6.00 (1H, d, J = 2.0 Hz), 14.16
(1H, s); 13C NMR (125 MHz, CDCl3): δC 12.0, 14.3, 17.3, 26.5, 26.6, 29.1, 29.4, 29.5, 29.6,
29.8, 29.9, 32.0 (8C), 46.3, 69.3, 91.6, 96.8, 106.2, 162.3, 163.0, 167.7, 210.6; HRMS (ESI)
m/z: [M+H]+ Calcd for C22H37O4 365.2692; Found 365.2699.
4.2.5.8.(S)-1-(2-(Dodecyloxy)-4,6-dihydroxyphenyl)-2-methylbutan-1-one (14)
Synthesised using 0.913 mmol of 5 and 1.096 mmol of 1-bromododecane. Isolated
22
D
after SPE (SiGel; hexane/EtOAc 92:8) as a pale yellow oil (98 mg, 0.259 mmol, 28%).
+4.7 (c 0.30, MeOH); UV (MeOH) λmax (log
α
[ ]
ε
): 238 (4.32), 290 (3.99) nm; IR νmax (thin film)
cm-1: 3327, 2924, 2854, 1739, 1622, 1593, 1456, 1366, 1216, 1160, 1109, 827; 1H NMR (500
MHz, CDCl3): δH 0.88 (3H, t, J = 7.5 Hz), 0.89 (3H, t, J = 7.5 Hz), 1.16 (3H, d, J = 6.5 Hz),
1.28-1.30 (16H, m), 1.35 (1H, m), 1.46 (2H, m), 1.83 m (1H, m), 1.86 (2H, m), 3.73 (1H, q,
J = 6.5 Hz), 4.00 (2H, t, J = 6.5 Hz), 5.92 (1H, d, J = 2.0 Hz), 6.00 (1H, d, J = 2.0 Hz), 6.02
(1H, br s), 14.19 (1H, s); 13C NMR (125 MHz, CDCl3): δC 12.0, 14.3, 17.1, 22.9, 26.9, 26.5,
29.3, 29.5, 29.6, 29.77 (2C), 29.8, 29.9, 32.1, 46.3, 69.4, 91.7, 96.8, 106.1, 162.5, 163.0,
167.7, 210.7; HRMS (ESI) m/z: [M+H]+ Calcd. for C23H39O4 379.2848; Found 379.2864.
4.2.5.9.(S)-1-(2,4-Dihydroxy-6-(octadecyloxy)phenyl)-2-methylbutan-1-one (15)

ACCEPTED MANUSCRIPT
Synthesised using 0.9 mmol of 5 and 1.079 mmol of 1-bromooctadecane. Isolated
22
D
after SPE (SiGel; hexane/EtOAc 92:8) as a colourless oil (118 mg, 0.255 mmol, 28%).
+4.2 (c 0.35, MeOH); UV (MeOH) λmax (log
α
[ ]
ε
): 244 (4.32), 289 (3.98) nm; IR νmax (thin film)
cm-1: 3328, 2923, 2853, 1738, 1623, 1593, 1455, 1373, 1214, 1159, 1104, 774; 1H NMR (500
MHz, CDCl3): δH 0.89 (3H, t, J = 7.5 Hz), 0.91 (3H, t, J = 7.5 Hz), 1.16 (3H, d, J = 6.5 Hz),
1.27-1.33 (28H, m), 1.42 (1H, m), 1.46 (2H, m), 1.81 (1H, m), 1.85 (2H, m), 3.73 (1H, q, J =
6.5 Hz), 4.00 (2H, t, J = 6.5 Hz), 5.92 (1H, d, J = 2.0 Hz), 6.00 (1H, d, J = 2.0 Hz), 6.07 (1H,
br s), 14.12 (1H, s); 13C NMR (125 MHz, CDCl3): δC 12.0, 14.3, 17.1, 22.9, 26.5, 26.9, 29.3,
29.6, 29.8-29.9 (11C), 32.2, 46.3, 69.4, 91.6, 96.8, 106.1, 162.3, 163.0, 167.8, 210.6; HRMS
(ESI) m/z: [M+H]+ Calcd for C29H51O4 463.3787; Found 463.3796.
4.2.6. Synthesis of compounds 18-24 from phloroglucinol
The first step in the synthesis of compounds 19-24 was the Friedel-Crafts acylation of
phloroglucinol using the appropriate acyl (propionyl, butyryl, pentanoyl, decanoyl,
octadecanoyl and benzoyl) chloride using the procedure described for 4 above. Compound 18
was synthesised starting from commercially available acetylphloroglucinol. TBDMS
protection of each acylphloroglucinol 16 was achieved according to the procedure described
for 5 above. Finally, the TBDMS-protected acylphloroglucinols 17 were simultaneously
deprotected and geranylated at the ortho phenolic hydroxyl as described for compound 6 to
produce compounds 18-24. Further details of the experimental procedures used for the
synthesis of compounds 18-24, including full compound characterisation data for the
corresponding precursor acylphloroglucinols and TBDMS-protected acylphloroglucinols, are
included in the Supporting Information.
4.2.6.1. (E)-1-(2-((3,7-Dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)ethanone (18)
Isolated after SPE (SiGel; hexane/EtOAc 8:2) as a pale yellow oil (45 mg, 0.148
mmol, 4.3% over two steps starting from commercial acetylphloroglucinol). UV (MeOH)

ACCEPTED MANUSCRIPT
λmax (log
ε
): 244 (4.36), 288 (3.97) nm; IR νmax (thin film) cm-1: 3136, 2909, 1737, 1621,
1
1561, 1464, 1373, 1284, 1259, 1222, 1165, 1104, 1071, 822, 756; H NMR (500 MHz,
(CD3)2CO): δH 1.59 (3H, s), 1.64 (3H, s), 1.77 (3H, s), 2.11-2.16 (4H, m), 2.55 (3H, s), 4.64
(2H, d, J = 6.5 Hz), 5.11 (1H, m), 5.56 (1H, m), 5.93 (1H, d, J = 2.0 Hz), 56.03 (1H, d, J =
2.0 Hz), 13.91 (1H, s); 13C NMR (125 MHz, (CD3)2CO): δC 16.7, 17.8, 25.9, 27.0, 33.2, 40.1,
66.4, 92.8, 96.6, 106.1, 119.9, 124.7, 132.2, 142.6, 164.0, 166.0, 168.3, 203.5. HRMS (ESI)
m/z: [M+H]+ Calcd. for C18H25O4 305.1753; Found 305.1766.
4.2.6.2. (E)-1-(2-((3,7-Dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)propan-1-one
(19)
Isolated after SPE (SiGel; hexane/EtOAc 9:1) as a yellow oil (50 mg, 0.157 mmol,
2.6% over three steps starting from 3). UV (MeOH) λmax (log ε): 243 (4.32), 289 (3.99) nm;
IR νmax (thin film) cm-1: 3310, 2973, 2937, 1622, 1594, 1448, 1376, 1220, 1166, 1100, 828,
756; 1H NMR (500 MHz, CDCl3): δH 1.15 (3H, t, J = 7.0 Hz), 1.61 (3H, s), 1.68 (3H, s), 1.74
(3H, s), 2.10 (2H, m), 2.13 (2H, m), 3.03 (2H, q, J = 7.0 Hz), 4.57 (2H, d, J = 6.5 Hz), 5.10
(1H, m), 5.51 (1H, m), 5.53 (1H, br s), 5.91 (1H, d, J = 2.0 Hz), 5.98 (1H, d, J = 2.0 Hz),
14.05 (1H, br s); 13C NMR (125 MHz, CDCl3): δC 9.0, 16.9, 18.0, 25.9, 26.5, 37.8, 39.7, 66.0,
91.7, 96.6, 106.3, 118.6, 123.8, 132.3, 142.4, 162.4, 163.2, 167.4, 207.0; HRMS (ESI) m/z:
[M+H]+ Calcd. for C19H27O4 319.1909; Found 319.1917.
4.2.6.3. (E)-1-(2-((3,7-Dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)butan-1-one
(20)
Isolated after SPE (SiGel; hexane/EtOAc 94:6) as a pale yellow oil (110 mg, 0.331
mmol, 5.7% over three steps starting from 3). UV (MeOH) λmax (log ε): 243 (4.39), 290
(3.99) nm; IR νmax (thin film) cm-1: 3317, 2970, 1739, 1606, 1435, 1366, 1215, 745; 1H NMR
(500 MHz, CDCl3): δH 0.97 (3H, t, J = 6.5 Hz), 1.61 (3H, s),1.69 (3H, s), 1.71 (2H, s), 1.74
(3H, s), 2.10 (2H, m), 2.13 (2H, m), 2.97 (2H, t, J = 7.0 Hz), 4.54 (2H, d, J = 6.5 Hz), 5.10

ACCEPTED MANUSCRIPT
(1H, m), 5.50 (1H, t, J = 6.5 Hz), 5.91 (1H, s, HO4), 5.93 (1H, d, J = 2.0 Hz), 5.99 (1H, d, J =
13
2.0 Hz), 14.10 (1H, s); C NMR (125 MHz, CDCl3): δC 14.2, 16.9, 17.9, 18.9, 25.9, 26.5,
39.7, 46.5, 65.9, 92.1, 96.6, 106.0, 118.4, 123.8, 132.2, 142.7, 162.43, 163.4, 167.3, 206.9.
HRMS (ESI) m/z: [M+H]+ Calcd. for C20H29O4,333.2066; Found 333.2057.
4.2.6.4. (E)-1-(2-((3,7-Dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)pentan-1-one
(21)
Isolated after SPE (SiGel; hexane/EtOAc 8:2) as a colourless oil (95 mg, 0.274 mmol,
2.2% over three steps starting from 3). UV (MeOH) λmax (log ε): 239 (4.36), 290 (3.97) nm;
IR νmax (thin film) cm-1: 3263, 2957, 2929, 1739, 1621, 1569, 1416, 1346, 1198, 1164, 1102,
827, 791; 1H NMR (500 MHz, CDCl3): δH 0.92 (3H, t, J = 6.5 Hz), 1.37 (2H, q, J = 6.5 Hz),
1.61 (3H, s), 1.64 (2H, m), 1.69 (3H, s), 1.74 (3H, s), 2.10 (2H, m), 2.13 (2H, m), 3.00 (2H,
m), 4.55 (2H, d, J = 6.5 Hz), 5.10 (1H, t, J = 6.5 Hz), 5.50 (1H, t, J = 6.5 Hz), 5.92 (1H, d, J
13
= 2.0 Hz), 5.99 (1H, d, J = 2.0 Hz), 6.38 (1H, s), 14.04 (1H, s); C NMR (125 MHz): δC
14.2, 16.9, 17.9, 22.8, 25.9, 26.5, 27.4, 39.6, 44.4, 65.9, 91.9, 96.6, 106.1, 118.4, 123.8,
132.2, 142.6, 162.9, 163.1, 167.4, 206.8; HRMS (ESI) m/z: [M+H]+ Calcd. for C21H31O4
347.2222; Found 347.2223.
4.2.6.5. (E)-1-(2-((3,7-Dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)decan-1-one
(22)
Isolated after SPE (SiGel; hexane/EtOAc 96:4) as a colourless oil (85 mg, 0.204
mmol, 4.2% over three steps starting from 3). UV (MeOH) λmax (log ε): 244 (4.39), 293
(3.98) nm; IR νmax (thin film) cm-1: 3309, 2924, 2854, 1738, 1620, 1594, 1445, 1376, 1206,
1
1167, 1097, 829, 754; H NMR (500 MHz, CDCl3): δH 0.88 (3H, t, J = 6.5 Hz), 1.26 (12H,
br s), 1.35 (2H, m), 1.61 (3H, s), 1.68 (3H, s), 1.74 (3H, s), 2.09 (2H, m), 2.13 (2H, m), 2.99
(2H, m), 4.56 (2H, d, J = 6.5 Hz), 5.11 (1H, t, J = 6.5 Hz), 5.51 (1H, t, J = 6.5 Hz), 5.92 (1H,
d, J = 2.0 Hz), 5.99 (1H, d, J = 2.0 Hz), 14.12 (1H, s); 13C NMR (125 MHz, CDCl3): δC 14.4,

ACCEPTED MANUSCRIPT
16.9, 17.9, 22.9, 25., 25.9, 26.6, 29.6, 29.7-29.8 (3C), 32.2, 39.8, 44.7, 65.9, 91.8, 96.6,
106.1, 118.5, 123.8, 132.2, 142.6, 163.0, 163.1, 167.5, 206.5; HRMS (ESI) m/z: [M+H]+
Calcd. for C26H41O4 417.3005; Found 417.3001.
4.2.6.6. (E)-1-(2-((3,7-Dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)octadecan-1-
one (23)
Isolated after SPE (SiGel; hexane/EtOAc 96:4) as a pale yellow oil (650 mg, 1.230
mmol, 19% over three steps starting from 3). UV (MeOH) λmax (log
ε
): 237 (4.18), 286 (3.87)
1
nm; IR νmax (thin film) cm-1: 3020, 2924, 2854, 1715, 1662, 1447, 1214, 1097, 829, 750; H
NMR (400 MHz, CDCl3): δH 0.89 (3H, t, J = 6.8 Hz), 1.26-1.35 (34H, br s), 1.62 (3H, s),
1.69 (3H, s), 1.75 (3H, s), 2.10 (2H, m), 2.13 (2H, m), 2.99 (2H, m), 4.55 (2H, d, J = 6.5 Hz),
5.10 (1H, t, J = 6.5 Hz), 5.50 (1H, t, J = 6.5 Hz), 5.91 (1H, d, J = 2.0 Hz), 5.98 (1H, d, J = 2.0
13
Hz), 14.12 (1H, s); C NMR (100 MHz, CDCl3): δC 14.3, 16.9, 17.9, 22.9, 25.3, 25.9, 26.5,
29.6, 29.8-29.9 (15C'), 32.1, 39.7, 44.7, 65.9, 91.7, 96.6, 106.3, 118.5, 123.8, 132.2, 142.5,
162.6, 163.1, 167.5, 206.7; HRMS (ESI) m/z: [M+H]+ Calcd. for C34H57O4 529.4257; Found
529.3710.
4.2.6.7. (E)-(2-((3,7-Dimethylocta-2,6-dien-1-yl)oxy)-4,6-dihydroxyphenyl)(phenyl)
methanone (24)
Isolated after SPE (SiGel; hexane/EtOAc 9:1) as a pale yellow oil (96 mg, 0.262
mmol, 10% over three steps starting from 3); UV (MeOH) λmax (log ε): 237 (4.45), 288 (3.99)
nm; IR νmax (thin film) cm-1: 3308, 2970, 2923, 1738, 1621, 1590, 1446, 1376, 1271, 1154,
1095, 825, 752; 1H NMR (500 MHz, CDCl3): δH 1.52 (3H, s), 1.61 (3H, s), 1.71 (3H, s), 1.85
(2H, m), 1.94 (2H, m), 4.21 (2H, d, J = 6.5 Hz), 4.61 (1H, t, J = 6.5 Hz), 5.05 (1H, m), 5.89
(1H, d, J = 2.0 Hz), 6.07 (1H, d, J = 2.0 Hz), 7.36 (2H, dt, J = 8.0, 1.5 Hz), 7.42 (1H, d, J =
8.0 Hz), 7.49 (2H, dt, J = 8.0, 1.5 Hz), 12.27 (1H, s); 13C NMR (125 MHz, CDCl3): δC 16.8,
17.9, 25.9, 26.4, 39.4, 65.5, 92.4, 96.5, 106.2, 118.1, 124.0, 127.6 (2C), 127.8, 130.7, 132.0,

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140.7, 142.5, 162.4, 163.5, 165.9, 200.1; HRMS (ESI) m/z: [M+H]+ Calcd. for C23H27O4
367.1909; Found 367.1893.
4.3.
Antibacterial assay against Staphylococcus aureus
Unless otherwise stated, all chemicals were obtained from Sigma-Aldrich,
Gillingham, UK. Cation-adjusted Mueller-Hinton broth (MHB) was obtained from Oxoid and
was adjusted to contain 20 mg/L and 10 mg/L of Ca2+ and Mg2+, respectively. The S. aureus
strains used in this study included ATCC 25923, SA-1199B, RN4220, XU212, EMRSA-15
and EMRSA-16. A standard laboratory strain, ATCC 25923, which is sensitive to antibiotics
like tetracycline [15], was also used in this study. SA-1199B over-expresses the NorA MDR
efflux pump [16], RN4220 possesses the MsrA macrolide efflux protein [17], XU212 is a
Kuwaiti hospital isolate which is an MRSA strain possessing the TetK tetracycline efflux
pump [15], whilst the EMRSA-15 strain [18] and EMRSA-16 strain [19] are epidemic in the
UK. These strains were the generous gift of Dr Paul Stapleton (UCL).
All S. aureus strains were cultured on nutrient agar (Oxoid) and incubated for 24 h at
37 °C prior to MIC determination. A stock solution of norfloxacin was prepared by
dissolving the antibiotic (2 mg) in DMSO (244 µL; Sigma) and diluting 16-fold with MHB to
obtain the desired starting concentration (512 mg/L) of antibiotic. Similarly, stock solutions
of the test compounds were prepared by initial dissolution in DMSO followed by dilution
with MHB to produce target concentrations of 512 mg/L. No significant precipitation was
noted for any of the test compounds when stock solutions were prepared at this concentration.
An inoculum density of 5 × 105 colony forming units (cfu/mL) of each bacterial strain was
prepared in normal saline (9 g/L) by comparison with a 0.5 MacFarland turbidity standard.
During the experiment, MHB (125 µL) was added to each well of a 96-well plate,
save for the final column which was left empty. 125 µL of the stock solution of the
compound to be tested (or the control antibiotic norfloxacin) was then added to the MHB in

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the first well. Using a multi-channel pipette, the contents of the first well were mixed
thoroughly, followed by the transfer of 125 µL of the well contents to the second well. This
two-fold serial dilution process was continued up until the tenth well, and the final 125 µL
solution was added to the final (empty) well. The inoculum (125 µL) of each bacterium at a
density of 5 × 105 cfu/mL was added to all wells except those in the final column. The final
concentrations of each test compound thus ranged from 128 mg/L to 0.25 mg/L in the initial
evaluation (similarly, the final concentrations of DMSO in the assays ranged from 1.56% v/v
at the highest concentration of test compound to 0.0031% v/v at the lowest). The contents of
the wells in columns 11 and 12 represented growth control (bacteria but no compound) and
sterility control (compound but no bacteria) respectively. Every assay was performed in
duplicate. The microtitre plates were then incubated at 37 °C for the appropriate incubation
time. For the MIC determination, 20 µL of a 5 mg/mL methanolic solution of 3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was added to each of the
wells, followed by incubation for 20 min. Bacterial growth was indicated by a colour change
from yellow to dark blue. The MIC was recorded as the lowest concentration at which no
growth was observed [11]. If no growth was observed at any of the concentrations tested, the
assay was repeated starting with a stock solution of lower concentration. If growth was
observed at all of the concentrations tested, the assay was repeated starting with a stock
solution of higher concentration.
Acknowledgements
This work was supported by the Heptagon Fund.
Appendix A. Supplementary data
Supplementary data related to this article can be found at

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Fig. 1. (S)-(2-Methylbutanoyl)phloroglucinols with ortho alkyloxy variants

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Fig. 2. ortho geranylated acylphloroglucinols with acyl variants

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Scheme 1
Synthesis of compounds 6-15
SOCl2
HO
Cl
O
O
1
2 (93%)
HO
OH
HO
OH
O
2, AlCl3
CS2, PhNO2
OH
OH
3
4 (54%)
TBDMSCl,
imidazole
TBDMSO
OH
acetone
TBDMSO
O
5 (81%)
HO
OR
R-Br, K2CO3
DMF
OH
O
6-15 (3-30%)