Synthesis of tubuphenylalanine and epi-tubuphenylalanine via regioselective aziridine ring opening with carbon nucleophiles followed by hydroboration-oxidation of 1,1-substituted amino alkenes

Article abstract of DOI:10.1016/j.tet.2018.10.024

An efficient synthesis of N-Boc-tubuphenylalanine benzyl ester (N-Boc-Tup-OBn, 1a) and N-Boc-epi-tubuphenylalanine benzyl ester (N-Boc-epi-Tup-OBn, 1b) is reported herein. Regioselective aziridine 4 ring opening with carbon nucleophiles followed by hydrob

Full text of DOI:10.1016/j.tet.2018.10.024

Accepted Manuscript
Synthesis of tubuphenylalanine and epi-tubuphenylalanine via regioselective aziridine
ring opening with carbon nucleophiles followed by hydroboration-oxidation of 1,1-
substituted amino alkenes
Ramesh B. Reddy, Premansh Dudhe, Priyanka Chauhan, Sagnik Sengupta, Chelvam
Venkatesh
PII:
S0040-4020(18)31230-4
10.1016/j.tet.2018.10.024
TET 29857
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 21 August 2018
Revised Date: 10 October 2018
Accepted Date: 13 October 2018
Please cite this article as: Reddy RB, Dudhe P, Chauhan P, Sengupta S, Venkatesh C, Synthesis of
tubuphenylalanine and epi-tubuphenylalanine via regioselective aziridine ring opening with carbon
nucleophiles followed by hydroboration-oxidation of 1,1-substituted amino alkenes, Tetrahedron (2018),
doi: https://doi.org/10.1016/j.tet.2018.10.024.
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ACCEPTED MANUSCRIPT
Graphical Abstract
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Synthesis of tubuphenylalanine and epi-tubuphenylalanine via regioselective
aziridine ring opening with carbon nucleophiles followed by hydroboration-
oxidation of 1,1-substituted amino alkenes
Ramesh B. Reddy,a Premansh Dudhe,a Priyanka Chauhan,a Sagnik Sengupta,a and Chelvam
Venkatesh*,a,b
a
Discipline of Chemistry, Indian Institute of Technology Indore, Khandwa Road, Simrol, Indore-453
552, India.
b
Discipline of Biosciences and Biomedical Engineering, Indian Institute of Technology Indore,
Khandwa Road, Simrol, Indore-453 552, India.
*email: cvenkat@iiti.ac.in; fax: +91-731-2431482; Tel: +91-731-2438789
ABSTRACT
An efficient synthesis of N-Boc-tubuphenylalanine benzyl ester (N-Boc-Tup-OBn, 1a) and N-Boc-epi-
tubuphenylalanine benzyl ester (N-Boc-epi-Tup-OBn, 1b) is reported herein. Regioselective aziridine 4
ring opening with carbon nucleophiles followed by hydroboration of 1,1-substituted aminoalkene 3
using 9-BBN and subsequent oxidation in an alkaline medium are used as the key steps to provide N-
tosyl 1,4-aminoalcohols. The 1,4-aminoalcohols are successfully transformed into the desired products
with an overall yield of 23% for 1a and 11% for 1b over 8 consecutive steps separately.
Keywords: Regioselective aziridine ring opening, Grignard reaction, Hydroboration, 9-BBN,
Tubuphenylalanine, epi-Tubuphenylalanine
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1. Introduction
Mitotic spindle microtubules are important drug targets in designing new anticancer therapy.
Currently marketed anticancer drugs like paclitaxel and vinblastine work on suppression of microtubule
dynamics in fast dividing cells leading to their apoptosis.1 In the last decade, tubulysin compounds
have emerged as a new class of highly potent cytotoxic peptides which can inhibit tubulin
polymerisation more efficiently than vinca alkaloids.2 However, the scarce supply of these natural
anticancer agents has attracted the attention of medicinal chemists to develop novel routes towards the
synthesis of a tubulysin family of natural products.3
Höfle and co-workers first isolated tubulysins from myxobacteria culture broths as antimitotic
tetrapeptides.4 Later, Müller and co-workers identified a gene cluster in myxobacteria with mixed non-
ribosomal peptide synthetase-polyketide synthase system that is responsible for biosynthesis of
tubulysin.5 Interestingly, tubulysin was found to surpass cell growth inhibition activity of all the
conventional anticancer natural products such as epothilones, vinblastine and taxol by a factor of 20 to
1000.6 Tubulysin interacts with the eukaryotic cytoskeleton and prevents microtubule formation by
binding with tubulin protein leading to apoptosis of cells.7 The interesting biological activity of
tubulysin and its derivatives justifies the rapidly growing interest in the development of novel
chemosynthetic methods to access more potent tubulysin derivatives with cytotoxicity in the range of
low nanomolar to picomolar concentrations.
The tubulysin scaffold is composed of four fragments: N-methyl-D-pipecolic acid (D-Mep), -
L
isoleucine (L-Ile), tubuvaline (Tuv), and tubuphenylalanine (Tup) or tubutyrosine (Tut) (Fig. 1)
connected to each other through amide linkages. Mep is a carboxylic acid derivative of piperidine and
is biosynthesized from -lysine5a whereas, isoleucine is a natural amino acid which provides necessary
L
hydrophobic interactions in the tubulin protein binding pocket. Tubuvaline (Tuv) is the most complex
component of tubulysin compounds with a thiazole ring containing a side chain chiral acetoxy carbon.
The C-terminus fragments, Tup or Tut, are γ-amino acid homologous of phenylalanine and tyrosine,
respectively. SAR studies5b,8 have confirmed that few modifications in the N-terminus and Tuv units of
the molecule are well tolerated and will not adversely affect the overall potency of the molecule. In
contrast to this, the C-terminus end (Tup or Tut) has excellent scope for modifications. This part of the
tubulysin natural products has been exploited to develop several new potent derivatives of this type of
anticancer molecules.
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Fig. 1. Structure of tubulysin family of antimitotic agents.
Seiber and coworkers have designed pretubulysin photoaffinity probes by C-terminus
modification for imaging studies using fluorescent labelling.9 Leamon et al. have prepared targeted
chemotherapeutic warheads of tubulysin derivatives and delivered to cancer cells via folic acid
conjugation.10
The existing methods for the synthesis of Tup and epi-Tup mostly employ chiral auxiliary
controlled alkylation using SAMP-hydrazone or pseudoephedrine,11 oxazolidinones,12–13 and tert-
butanesufinamide,3c,14 along with substrate controlled diastereoselective methylation of chiral lactam,15
diastereoselective hydrogenation,3d,16 epoxide ring-opening17 and Ireland-Claisen rearrangement of
allyl ester of β-amino acid.18 Recently, Park and co-workers have reported a chiral auxiliary-free direct
synthetic pathway for the synthesis of tubuphenylalanine by Lewis acid-catalyzed Mukaiyama aldol
condensation reaction.19
Although a variety of methods have been developed to date, there is enough scope to introduce
refined synthetic protocols for a large-scale synthesis of these biologically active entities. Herein, we
report a novel and high yielding synthetic route for the synthesis of tubuphenylalanine (Tup) and epi-
tubuphenylalanine (epi-Tup) using hydroboration-oxidation of substituted alkene 3 as the crucial step
without a chiral auxiliary in the substrate. Moreover, our synthetic route is efficient in terms of a
number of the steps (8-steps) and overall yield (23%) compared to an earlier reported synthesis20 of
tubuphenylalanine fragment in 16-steps with an overall yield of 2.5%. This practical and efficient
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synthesis of key components, 1a and 1b, will allow building a diverse set of new tubulysin
architectures in the future.
2. Results and discussion
In order to develop a method for the synthesis of tubuphenylalanine (Tup) and its biologically
active stereoisomer epi-tubuphenylalanine (epi-Tup), a simple retrosynthetic analysis of their closest
precursors 2a and 2b are depicted in Scheme 1. According to this strategy, the crucial N-tosyl-1,4-
amino alcohols 2a and 2b can be obtained from a common precursor, 1,1-disubstituted aminoalkene 3
through a regioselective hydroboration-oxidation reaction. Further, N-tosyl-benzylaziridine 4 could be
a key starting material to obtain 1,1-disubstituted aminoalkene 3 via regioselective ring opening.
Scheme 1. Retrosynthetic analysis of N-Boc-Tup-OBn (1a) and N-Boc-epi-Tup-OBn (1b).
Since its inception in 1956, hydroboration reaction has made available many alkylated borane
derivatives that are essential precursors for regio- and stereoselective introduction of hydroxy
functionality in a substituted alkene.21 Moreover, in 1979, Brown and co-workers reported pioneering
studies on hydroboration of allene22 using 9-BBN to address the drawbacks of previous hydroboration
reactions that lacks regioselectivity. Further studies on the application of bicycloboranes have revealed
that hydroboration of substituted terminal alkenes using 9-BBN generally results in the high
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regioselective23 preparation of organoboranes which can be easily oxidized to their corresponding
alcohols in alkaline H2O2.24
We began our synthesis starting from -phenylalanine to obtain the required 1,1-substituted
L
aminoalkene 3 for hydroboration reaction. -phenylalanine was converted to phenylalaninol 5 by
L
performing reduction of acid functionality with NaBH4 in the presence of I2 under refluxing
tetrahydrofuran in excellent yield. Without further purification, the hydroxy and amino functionalities
in phenylalaninol
5
was tosylated using p-toluenesulfonyl chloride to give (S)-2-(4-
methylphenylsulfonamido)-3-phenylpropyl 4-methylbenzenesulfonate intermediate. The bis(tosyl)
intermediate shown within parentheses was in situ cyclized by intramolecular sulfonamide nucleophilic
attack on tosyl protected hydroxyl group in the presence of a mild base, Et3N, to furnish N-Ts-
benzylaziridine 4 in 85% isolated yield (Scheme 2).25
Scheme 2. Synthesis of (S)-2-benzyl-1-tosylaziridine (4). Reagents and conditions (i) NaBH4, I2, THF,
0 °C to reflux, 24 h, 96% (ii) Et3N, TsCl, CH3CN, 0 °C to rt, 1 h, 85%.
The key intermediate N-Ts-benzylaziridine 4 was thus prepared in gram scale to obtain the
desired 1,1-substituted aminoalkene 3 which is an important substrate for regio- and stereoselective
hydroboration reactions proposed to prepare 2a and 2b. Literature survey shows that there are few
procedures for regioselective ring opening of aziridine by nucleophiles including Grignard reagents.26
Cu (I) salts play an instrumental role in Grignard reactions for generating active organocuprate species.
We have explored catalytic amounts of CuI, CuBr, and CuCN under different reaction conditions to
prepare the desired alkene 3 in moderate to high yields (Table 1). The initial reaction was performed in
the presence of CuI (Table 1, entry 1, 2) and after 12 h of reaction at room temperature, the expected
aminoalkene 3 was isolated only in a moderate yield of 50-55%. However, no noticeable enhancement
in the yield of 3 was observed when CuBr was used as a catalyst (Table 1, entry 3, 4). Copper(I)halides
form homocuprates by the reaction of either copper(I) bromide or copper(I) iodide with a minimum of
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two equivalents of Grignard reagents in ether or THF. The initially formed polymeric (RCu)n species
were insoluble in Et2O and THF, but dissolve only on addition of a second equivalent of Grignard
reagent. The resultant organocuprates are thermally labile and generally prepared at low temperatures.27
Because of this reason the yield of 3 was moderate when copper(I)halides were used as catalysts and
require longer reaction time for completion (Table 1, entries 2, 4).
Interestingly, the yield of aminoalkene 3 has dramatically improved in the presence of CuCN (0.2 eq)
to 90% within 2 h of the reaction time (Table 1, entry 6). The increase in yield of 3 is because when
copper(I)halides were substituted with copper(I) cyanide as a catalyst, higher order cyanocuprates28,29
were formed smoothly at 0 °C to rt. The cyanocuprates have higher thermal stability than
homocuprates which are generally generated at a lower temperature. Moreover, cyanocuprates retain
the reactivity of homocuprates as well as the stability of heterocuprates when copper(I)cyanide was
used as the catalyst.
Table 1. Regioselective ring opening of (S)-2-Benzyl-1-tosylaziridine (4).
Cu (I) salt
(equiv.)
Rxn
time
Grignard Yield
reagent
(equiv.)
2.0
of 3a
1
2
3
4
5
6
CuI (0.1)
CuI (0.2)
12 h
24 h
50%
55%
60%
60%
73%
90%
2.5
2.0
CuBr (0.1) 12 h
CuBr (0.2) 24 h
CuCN (0.1) 2 h
CuCN (0.2) 2 h
2.5
1.5
2.0
a Isolated
Substrate-controlled hydroboration can play a vital role in deciding the diastereoselectivity of
formation of N-tosyl-1,4-amino alcohols, 2a–b. Therefore, we plan to verify our hypothesis by
preparing two differently protected amino alkenes such as 3 and 6. Initially, N-tosyl protected alkene 3
was treated with Boc anhydride (Boc2O) in the presence of dimethyl amino pyridine (DMAP) in
dichloromethane to afford N-Boc-N-tosyl derivative of the alkene 6 in excellent yield.
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The crucial hydroboration-oxidation of alkene functionality in 3 and 6 was realized later with 9-BBN.
The hydroboration of 6 and subsequent oxidation using H2O2 in the presence of 2 M NaOH afforded
1,4-amino alcohols 7 (Scheme 3) only as a mixture of diastereomers in the ratio 2:1 (from 1H NMR) in
70% yield. Moreover, the diastereomeric mixture of alcohols 7 remains non-separable by silica gel
column chromatography.
Scheme 3. Synthesis of N-tosyl-1,4-amino alcohols, 2a–b and 7. Reagents and conditions (i) a. 9-BBN,
THF, rt, 12 h b. 2 M NaOH, H2O2, 0 °C to rt, 12 h, 90%, dr = 2 :1 (ii) Boc2O, DMAP, CH2Cl2, rt, 2 h,
97% (iii) a. 9-BBN, THF, rt, 12 h b. 2 M NaOH, H2O2, 0 °C to rt, 12 h, 70%, dr = 2:1 (non-separable).
On the contrary, hydroboration-oxidation of alkene 3 in the presence of 9-BBN/2 M NaOH /
30% H2O2 smoothly resulted into a separable 2:1 diastereomeric mixture of 1,4-amino alcohols (2a,
2b) in a combined yield of 90%. After successful purification, 1,4-amino alcohol 2a (Scheme 4) was
subjected to oxidation reaction using pyridinium dichromate (PDC). During the oxidation of the
primary hydroxyl group in 2a, the primary alcohol forms chromate ester and subsequently oxidized to
an unstable aldehyde. The unstable aldehyde undergoes further oxidation in the presence of PDC to
form chromate carboxylic acid anhydride, [RC(O)OCrO3H] followed by intramolecular cyclization
with the NH-tosyl amino group to form N-Ts-γ-lactam 8a as the only product.30 Detosylation of 8a was
achieved with Mg/NH4Cl via a single electron transfer mechanism under reflux conditions in methanol
to provide 9a. 31
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Me
Me
(i)
O
OH
N
Ts
TsHN
8a
2a
(ii)
Me
Me
(iii)
O
Boc
10a
O
N
N
H
9a
(iv)
Me
OBn
BocHN
O
1a
Scheme 4. Synthesis of N-Boc-Tup-OBn (1a). Reagents and conditions (i) PDC, DMF, rt, 12 h, 85%
(ii) Mg, NH4Cl, MeOH, reflux, 2 h, 84% (iii) Boc2O, DMAP, DCM, rt, 2 h, 96% (iv) a. LiOH, H2O2,
THF:H2O (1:1), rt, 12 h b. BnBr, Et3N, DCM, rt, 12 h, 75%.
The free NH group in 9a was protected as NHBoc using Boc2O and DMAP to afford 10a in
95% yield. The Boc derivative of γ-lactam 10a was reacted with LiOH in the presence of H2O2 to
facilitate opening of lactam ring to provide carboxylic acid intermediate. Without isolation, the
carboxylic acid intermediate was treated with benzyl bromide in the presence of a mild base, Et3N, to
afford N-Boc-Tup-OBn 1a in 75% yield over two steps.
Zanda and coworkers32 have established that the modification of Tup stereochemistry has only
minor effects on the tubulysin cytotoxicity, this makes epi-tubuphenylalanine an important synthetic
target. Therefore, in our synthetic strategy, 1,4-amino alcohol derivative 2b was next subjected to
synthetic transformations similar to the conversion of 2a to 1a as shown in Scheme 5. The primary
alcohol 2b was oxidized to chromate carboxylic acid anhydride, which undergoes in situ cyclisation to
yield 8b. Subsequent tosyl deprotection and Boc protection of amide NH yielded N-Boc γ-lactam 10b.
Lactam ring opening and benzyl protection at the C-terminus provided the desired molecule N-Boc-epi-
Tup-OBn 1b in good yield.
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Scheme 5. Synthesis of N-Boc-epi-Tup-OBn (1b). Reagents and conditions (i) PDC, DMF, rt, 12 h,
83% (ii) Mg, NH4Cl, MeOH, reflux, 2 h, 81% (iii) Boc2O, DMAP, DCM, rt, 2 h, 99% (iv) a. LiOH,
H2O2, THF:H2O (1:1), rt, 12 h b. BnBr, Et3N, DCM, rt, 12 h, 72%.
It was imperative to determine the stereochemistry of major (2a) and minor (2b) 1,4-aminoalcohol
diastereoisomers. The stereochemistry was assigned by performing COSY and NOESY experiments on
the lactams 8a and 8b (N-tosyl-5-benzyl-3-methylpyrrolidin-2-one). The NOE (Fig. 2) was observed
between two protons at 3-Ha and 5-Hb in 8b, whereas 8a did not show any NOE between two protons
3-Ha and 5-Hb. The appearance of cross peaks off the diagonal peaks in 8b for protons 3-Ha and 5-Hb
indicated spatial coupling whereas its absence in 8a confirmed the proposed trans stereo chemical
arrangement of 3-Ha and 5-Hb protons in space. This study unequivocally established the
stereochemistry of major and minor 1,4-aminoalcohols 2a (2R, 4S) and 2b (2R, 4R) respectively (see
2D-NMR data in supporting information).
Fig. 2. Stereochemical assignments of 8a and 8b through 1H-1H COSY and NOESY studies.
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3. Conclusions
In summary, we have developed a high yielding synthetic approach for C-terminus fragments of
natural and synthetic tubulysins, i.e. tubuphenylalanine and epi-tubuphenylalanine via regioselective
aziridine ring opening with carbon nucleophiles followed by hydroboration-oxidation of 1,1-substituted
amino alkenes using 9-BBN. The chiral auxiliary-free synthetic protocol leads to overall good yields
for both the biologically active γ-amino acids. These practical, concise and scalable synthetic steps
have promising scope in the gram-scale synthesis of tubulysin and its derivatives.
4. Experimental Section
4.1. General information
All reactions were performed in oven-dried glassware under the inert atmosphere with magnetic
stirring. Air and moisture-sensitive liquids and solutions were transferred via glass syringes. TLC was
performed on 0.25 mm Merck TLC silica gel 60 F254 plates and visualized under UV light (254 nm) or
by staining with bromocresol green, ninhydrin, KMnO4. Flash chromatography was performed on 230–
400 mesh silica gel. All reagents were purchased from commercial suppliers and used without further
purification unless otherwise stated. Solvents were distilled using suitable drying agents (CaH2 or Na
1
wire, Mg turnings) under the nitrogen atmosphere. H, 13C-NMR, COSY, NOESY spectra were
recorded on Avance III 400 MHz Ascend Bruker. Chemical shifts are expressed in ppm relative to
TMS (1H, 0 ppm) or solvent signals: CDCl3 (1H, 7.26 ppm; 13C, 77.26 ppm) coupling constants are
expressed in Hz. High-resolution mass spectra electrospray ionization (HRMS-ESI) performed on
Bruker Daltonik LC/MS spectrometer. FT-IR spectra of samples dissolved in CH2Cl2 were recorded in
using Fourier Transform Infrared-Attenuated Total reflection (FTIR-ATR) Spectrometer, Bruker
(Tensor-27) over a range of 500–4000 cm-1.
4.2.
Experimental details and characterisation data
4.2.1. (S)-2-Amino-3-phenylpropan-1-ol (5)
L-phenylalanine (5 g, 30 mmol) was dissolved in dry THF (50 mL) in a 250 mL round-bottom
flask under an inert atmosphere. The reaction mixture was cooled to 0 °C, and solid NaBH4 (2.87 g,
75.70 mmol) was added to the reaction mixture in a single portion with stirring. A solution of Iodine
(6.6 g, 52.5 mmol) in dry THF (30 mL) was added to the reaction mixture dropwise using dropping
funnel over a period of 1 h. The reaction mixture was further heated to reflux on a preheated oil bath
overnight. The reaction mixture was cooled to room temperature, and methanol (30 mL) was slowly
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added until a clear solution is obtained. The solvent was evaporated under reduced pressure, and the
resulting white paste was dissolved in 2 M NaOH (50 mL). The mixture was stirred at room
temperature for 8 h. The aqueous phase was extracted with CH2Cl2 (100 × 3 mL) and the combined
organic layers were dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to
afford -phenylalaninol (4 g, 96%) as a white solid, which was used in the next step without further
L
purification. m.p. 90–94 C. TLC: Rf 0.19 (1:9, MeOH/CH2Cl2). [α]ꢁꢀꢁ.ꢂ = −7.9 (c 0.13, CH2Cl2). IR
(CH2Cl2): 3352 (O–H) 3279 (N–H), 3083, 3061, 3027, (=C–H), 2919, 2855(C–H), 1649 (N–H), 1551–
1
1502 (C=C) 1452 (C–H), 1054 (C–O), 701 (=C–H) cm-1. H NMR (400 MHz, CDCl3): δ = 7.25–7.11
(m, 5H), 3.56 (dd, J = 10.8, 2.8 Hz, 1H), 3.36–3.27 (dd, , J = 10.8, 8.0 Hz, 1H), 3.04 (brs, 1H), 2.72
13
(dd, J = 13.6, 4.8 Hz, 1H), 2.45 (dd, J = 13.6, 8.8 Hz, 1H), 1.97 (brs, 3H) ppm. C NMR (100 MHz,
CDCl3): δ = 138.7, 129.2, 128.6, 126.4, 66.2, 54.2, 40.8 ppm. HRMS (ESI) m/z [M+H]+ calcd. for
C9H13NO 152.1070, found 152.1101.
4.2.2. (S)-2-Benzyl-1-tosylaziridine (4)
4 Å molecular sieves (2 g) were flame dried in a 100 mL two neck round bottom flask
under reduced pressure for 15-min or more until there is no sign of the appearance of water
droplets or moisture in the flask. The heat dried molecular sieves were cooled to room
temperature under inert atmosphere. CH3CN (50 mL), magnetic bar and L-phenylalaninol
5 (2.8
g, 18.5 mmol) were charged to this flask. The reaction mixture was briefly cooled to 0 °C, and
Et3N (7.7 mL, 55.55 mmol) and tosyl chloride (3.52 g, 18.5 mmol) were added sequentially via
a syringe to the reaction mixture. The reaction mixture was warmed to room temperature and
further stirred for 1 h. After complete consumption of -phenylalaninol, as confirmed by TLC,
L
acetonitrile was evaporated under reduced pressure, and the residue was dissolved in EtOAc (50
mL). The resultant precipitate and molecular sieves were filtered using Buchner funnel and
washed with EtOAc (3 × 50 mL). The organic solvent was concentrated under reduced pressure
and the crude mixture was purified by silica gel column chromatography (1:24 EtOAc/hexane)
to afford 4 (4 g, 85%) as a white solid. m.p. 91–93 °C. TLC: Rf 0.21 (9:1, hexane/EtOAc).
[α]ꢁꢀꢁ.ꢃ = +16.4 (c 0.13, CH2Cl2). IR (CH2Cl2): 3029, (=C–H), 2922 (C–H), 1592, 1489, 1455
(C=C), 1317, 1157 (S=O), 712 (=C–H) cm-1. 1H NMR (400 MHz, CDCl3, 25 °C):
= 8.0 Hz, 2H), 7.21 (d, = 8.0 Hz, 2H), 7.16–7.14 (m, 3H), 7.05–7.03 (m, 2H), 2.97–2.91 (m,
1H), 2.81 (dd, = 14.4, 5.2 Hz, 1H), 2.70 (d, = 6.8 Hz, 1H), 2.68 (dd, = 14.4, 7.2 Hz, 1H),
2.42 (s, 3H), 2.16 (d,
δ = 7.68 (d, J
J
J
J
J
J
= 4.4 Hz, 1H) ppm. 13C NMR (100 MHz, CDCl3, 25 °C):
δ = 144.3,
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137.0, 134.8, 129.6, 128.7, 128.4, 127.8, 126.5, 41.1, 37.5, 32.8, 21.6 ppm. HRMS (ESI) m/z
[M+Na]+ calcd. for C16H17NO2S 310.0872, found 310.0883
.
4.2.3. (R)-4-Methyl-N-(4-methyl-1-phenylpent-4-en-2-yl) benzene sulfonamide (3)
A 250 mL two neck round-bottom flask was charged with catalytic amount of CuCN (0.2
equiv.) under an inert atmosphere. N-Tosyl-benzylaziridine 4 (2.5 g, 8.7 mmol) dissolved in dry THF
(50 mL) was added to the reaction mixture at 0 °C. Isopropenyl magnesium bromide (34.8 mL, 17.4
mmol, 0.5 M in THF) was added dropwise to the reaction mixture over a period of 20 minutes with
stirring. The reaction mixture was allowed to warm to room temperature and further stirred for 2 h.
After the consumption of N-Tosyl-benzylaziridine 4, as confirmed by TLC, the reaction mixture was
quenched with saturated NH4Cl (30 mL) solution and further diluted with EtOAc (100 mL). The
aqueous layer was extracted with EtOAc (3 × 50 mL) and the combined organic extracts were washed
with brine (100 mL) and dried over anhydrous Na2SO4. The organic layer was filtered, evaporated
under reduced pressure, and the crude residue was purified over silica gel column chromatography
(32:1 hexane/EtOAc) to afford the alkene 3 (2.1 g, 90%) as colorless liquid. TLC: Rf 0.17 (9:1,
hexane/EtOAc). [α]ꢁꢀꢁ.ꢁ = −1.5 (c 0.13, CH2Cl2). IR (CH2Cl2): 3284 (N–H), 3064, 3027 (=C–H), 2923,
1
(C–H), 1649, 1604 (C=C) 1499–1451 (C–H), 1322, 1153 (S=O), 701 (=C–H) cm-1. H NMR (400
MHz, CDCl3, 25 °C): δ = 7.65 (d, J = 8.0 Hz, 2H), 7.26–7.19 (m, 5H), 7.07–7.05 (m, 2H), 4.76–4.67
(m, 2H), 4.32 (d, J = 5.6 Hz, 1H), 3.49–3.41 (m, 1H), 2.85, (dd, J = 13.6, 5.6 Hz, 1H) ), 2.76, (dd, J =
13.6, 6.8 Hz, 1H) 2.41 (s, 3H), (dd, J = 14.0, 6.4 Hz, 1H), (dd, J = 14.0, 8.4 Hz, 1H), 1.38 (s, 3H) ppm.
13C NMR (100 MHz, CDCl3, 25 °C): δ = 143.2, 141.6, 137.2, 137.2, 129.5, 129.5, 128.4, 127.1, 126.5,
114.5, 52.6, 43.0, 41.2, 21.5, 21.5 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C19H23NO2S 352.1342,
found 352.1345.
4.2.4. (R)-tert-Butyl (4-methyl-1-phenylpent-4-en-2-yl)(tosyl)carbamate (6)
An oven dried 250 mL round-bottom flask was charged with a solution of alkene 3 (2.5 g, 7.59
mmol) in dry CH2Cl2 (50 mL). DMAP (0.185 g, 1.52 mmol) was added in single portion to the reaction
mixture and Boc2O (3.3 g, 15.2 mmol) was added using syringe. The reaction mixture was stirred at
room temperature for 2 h. After the completion of reaction, CH2Cl2 was evaporated under reduced
pressure and the residue was purified over silica gel column chromatography using 33:1 hexane/EtOAc
mixture as eluent to afford 6 (3.1 g, 97%) as a colorless liquid. TLC: Rf 0.54 (9:1, hexane/EtOAc).
[α]ꢁꢀꢁ.ꢄ = −99.6 (c 0.13, CH2Cl2). IR (CH2Cl2): 3064, 3027 (=C–H), 2972, 2922, 2852 (C–H), 1727
13

ACCEPTED MANUSCRIPT
1
(C=O), 1653 (C=C), 1493, 1455 (C–H), 1351, 1152 (S=O), 1088 (C–O), 673 (=C–H) cm-1. H NMR
(400 MHz, CDCl3, 25 °C): δ = 7.34–7.20 (m, 5H), 7.18–7.00 (m, 4H), 5.00–4.80 (m, 3H), 3.32 (dd, J
= 14.0, 9.6 Hz, 1H), 3.04 (dd, J = 14.0, 6.0 Hz, 1H), 2.79 (dd, J = 13.6, 8.0 Hz, 1H), 2.55 (dd, J = 13.6,
7.2 Hz, 1H), 2.35 (s, 3H), 1.84 (s, 3H), 1.34 (s, 9H) ppm. 13C NMR (100 MHz, CDCl3, 25 °C): δ =
150.7, 143.2, 142.3, 139.1, 137.8, 129.6, 128.8, 128.6, 127.7, 126.5, 114.2, 83.9, 59.3, 42.0, 38.7, 27.9,
22.2, 21.4 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C24H31NO4S 452.1866, found 452.2172.
4.2.5. General procedure for hydroboration reactions
9-BBN (3 equiv., 0.5 M THF solution) was added to a solution of alkene 3 or 6 (1.0 equiv.) in
dry THF at room temperature. The resulting solution was further stirred for 12 h at the same
temperature under nitrogen atmosphere. 2 M NaOH (3.5 equiv.) and 30% H2O2 (4 equiv.) were added
at 0 °C to the reaction mixture and stirring was continued for 12 h at room temperature. The reaction
mixture was quenched by adding saturated aq. NaCl (20 mL) at room temperature and the aqueous
layer was extracted with EtOAc (3 × 100 mL). The combined organic layers was dried over anhydrous
Na2SO4, filtered and concentrated under reduced pressure. The crude residue was purified using
hexane:EtOAc as eluent over silica gel column chromatography.
4.2.5.1. N-((2R,4S)-5-hydroxy-4-methyl-1-phenylpentan-2-yl)-4-methylbenzenesulfonamide (2a)
According to the general procedure described in section 4.2.5, 9-BBN (38.2 mL, 19.1 mmol, 0.5
M THF solution) was added to alkene 3 (2.1 g, 6.37 mmol) in dry THF (20 mL) and stirred at room
temperature for 12 h. 2 M NaOH (11.0 mL, 22.29 mmol) and 30% H2O2 (9.0 mL, 89.14 mmol) were
added at 0 °C to the reaction mixture and stirred for 12 h at room temperature and worked up as
mentioned in general procedure 4.2.5. The residue was purified over silica gel using column
chromatography (3:1 hexane/EtOAc), to afford 2a and 2b as colorless liquid and pale yellow liquid
respectively (9.1 g, combined yield 90 %, (2a:2b = 2:1). TLC: Rf 0.34 (1:1, hexane/EtOAc). [ꢅ]ꢁꢀꢂ.ꢁ = −43.0
(c 0.13, CH2Cl2). IR (CH2Cl2): 3504 (N–H), 3277 (O–H), 3061, 3028 (=C–H), 2925, 2872 (C–H), 1598
1
(C=C), 1494, 1453, (C–H), 1320, 1152 (S=O), 1090 (C–O), 700 (=C–H) cm-1. H NMR (400 MHz,
CDCl3, 25 °C): δ = 7.65 (d, J = 8.0 Hz, 2H), 7.26–7.19 (m, 5H), 6.69–6.97 (m, 2H), 4.66 (d, J = 7.2
Hz, 1H), 3.70–3.61 (m, 1H), 3.44 (dd, J = 10.4, 5.2 Hz, 1H), 3.33 (dd, J = 10.4, 6.8 Hz, 1H), 2.64–2.62
(m, 2H), 2.41 (s, 3H), 1.87–1.79 (m, 1H), 1.55–1.49 (m, 1H), 1.30–1.23 (m, 1H), 0.84 (d, J = 6.8 Hz,
3H) ppm. 13C NMR (100 MHz, CDCl3, 25 °C): δ = 143.2, 137.8, 136.9, 129.6, 129.5, 128.5, 127.0,
14

ACCEPTED MANUSCRIPT
126.6, 67.8, 52.9, 41.1, 39.1, 31.7, 21.5, 17.5 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C19H25NO3S
370.1447, found 370.1448.
4.2.5.2. N-((2R,4R)-5-hydroxy-4-methyl-1-phenylpentan-2-yl)-4-methylbenzenesulfonamide (2b)
TLC: Rf 0.27 (1:1, hexane/EtOAc). [α]ꢀꢁꢂ.ꢂ = +31.2 (c 0.13, CH2Cl2). IR (CH2Cl2): 3506 (N–H), 3283
(O–H), 3061, 3028 (=C–H), 2927, 2873 (C–H) 1598 (C=C), 1495, 1454, (C–H), 1322, 1154 (S=O),
1
1092 (C–O), 733 (=C–H) cm-1. H NMR (400 MHz, CDCl3, 25 °C): δ = 7.71 (d, J = 8.0 Hz, 2H),
7.27–7.18 (m, 5H), 7.02–7.01 (m, 2H), 4.85 (brs, 1H), 3.56–3.48 (m, 1H), 3.43 (dd, J = 10.4, 5.2 Hz,
1H), 3.28 (dd, J = 10.4, 7.2 Hz, 1H), 2.73 (dd, J = 13.6, 4.8 Hz 1H), 2.63 (dd, J = 13.6, 7.2 Hz 1H) 2.41
(s, 3H), 1.69 (brs, 1H), 1.64–1.57 (m, 1H), 1.55–1.48 (m, 1H), 1.22–1.16 (m, 1H), 0.68 (d, J = 6.4 Hz,
3H) ppm. 13C NMR (100 MHz, CDCl3, 25 °C): δ = 143.2, 137.9, 137.2, 129.6, 129.5, 128.4, 127.0,
126.5, 68.1, 53.2, 42.3, 38.5, 32.3, 21.5, 16.4 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C19H25NO3S
370.1447, found 370.1443.
4.2.5.3. tert-Butyl ((2R)-5-hydroxy-4-(S or R)methyl-1-phenylpentan-2-yl)(tosyl)carbamate (7) (dr
= 2:1)
According to the general procedure described in section 4.2.5, 9-BBN (28.14 mL, 13.97 mmol,
0.5 M THF solution) was added to alkene 6 (2 g, 4.65 mmol) in dry THF (20 mL) in a round bottom
flask (100 mL) at room temperature and stirred for 12 h. 2 M NaOH (8.14 mL, 16.28 mmol) was added
followed by addition of 30% H2O2 (6.6 mL, 65.0 mmol) at 0 °C. The reaction mixture was stirred for
12 h at room temperature and worked up as mentioned in general procedure 4.2.5. The crude residue
was purified over silica gel column chromatography using 3:1 hexane/EtOAc mixture as eluent to
afford colorless oil of 7 (1.65 g, 70%) as a diastereomeric mixture (2:1). TLC: Rf 0.23 (3:1,
hexane/EtOAc). [α]ꢁꢀꢁ.ꢁ = −49.8 (c 0.13, CH2Cl2). IR (CH2Cl2): 3564 (O–H), 3028 (=C–H) 2925, 2870
(C–H), 1722 (C=O), 1598 (C=C), 1494, 1453 (C–H), 1343, 1147 (S=O), 1085 (C–O), 669 (=C–H) cm-
1. 1H NMR (400 MHz, CDCl3, 25 °C): δ = 7.30–7.23 (m, 6H), 7.05–6.99 (m, 3H), 4.93–4.86 (m, 1H),
3.63–3.55 (m, 1H), 3.54–3.47 (m, 1H), 3.37–3.28 (m, 1H), 3.02–2.97 (m, 1H), 2.36 (s, 3H), 1.97–1.91
(m, 2H), 1.84–1.79 (m, 1H), 1.55–1.50 (m, 1H), 1.34 (s, 9H), 1.04 (d, J = 6.4 Hz, 2H), 1.01 (d, J = 6.8
Hz, 1H)* ppm. 13C NMR (100 MHz, CDCl3, 25 °C): δ = 150.7, 143.4*, 143.3, 139.0, 137.5, 129.6,
128.9, 128.7, 127.9*, 127.8, 126.5, 84.2*, 84.1, 68.2*, 67.8, 59.2, 59.0*, 39.3, 37.4, 37.3*, 33.2*, 33.1,
29.6*, 27.9, 22.7*, 21.5, 21.4*, 17.4, 14.1* ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C24H33NO5S
470.1970, found 470.2320. (*peaks from minor diastereoisomer)
15

ACCEPTED MANUSCRIPT
4.2.6. General procedure for the synthesis of N-Ts-γ-lactams 8a and 8b
Dry DMF (10 mL) was added to 1,4-amino alcohol (2a or 2b, 1.0 equiv.) taken in a 100 mL
round-bottom flask charged with a magnetic bead and the mixture was stirred at room temperature.
Pyridinium dichromate (5.0 equiv.) was added to above reaction mixture in one portion and stirred for
24 h at the same temperature. After the complete consumption of 1,4-amino alcohol, the reaction
mixture was quenched with cold water (10 mL). The aqueous layer was extracted with EtOAc (3 × 75
mL) and the combined organic layers were washed with cold brine (3 × 10 mL). The organic layer was
dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The residue was
purified over silica gel column chromatography.
4.2.6.1. (3S,5R)-5-Benzyl-3-methyl-1-tosylpyrrolidin-2-one (8a)
According to the general procedure described in section 4.2.6, to a solution of 1,4-amino
alcohol 2a (500 mg, 1.43 mmol) in dry DMF (10 mL) taken in a round bottom flask (50 mL), PDC (2.7
g, 7.19 mmol) was added to the reaction mixture and stirred for 24 h at room temperature. The work up
was performed as mentioned in the general procedure and the residue was purified over silica gel
column chromatography (6:1 hexane/EtOAc) to afford lactam 8a as colorless sticky liquid, (433 mg,
85%). TLC: Rf 0.53 (2:1, hexane/EtOAc). [α]ꢁꢀꢃ.ꢄ = +97.8 (c 0.13, CH2Cl2). IR (CH2Cl2): 3062, 3028,
(=C–H), 2931, 2874 (C–H) 1729 (C=O), 1597 (C=C), 1494, 1453 (C–H), 1352, 1159 (S=O), 711 (=C–
H) cm-1. 1H NMR (400 MHz, CDCl3, 25 °C): δ = 8.0 (d, J = 8.4 Hz, 2H), 7.35–7.21 (m, 7H), 4.54 (td,
J = 8.8, 3.2 Hz, 1H), 3.36 (dd, J = 13.2, 3.2 Hz, 1H), 2.81 (dd, J = 13.2, 9.6 Hz, 1H), 2.44 (s, 3H),
2.34–2.26 (m, 1H), 2.11–2.04 (m, 1H), 1.67–1.58 (m, 1H), 1.02 (d, J = 6.8 Hz, 3H) ppm. 13C NMR
(100 MHz, CDCl3, 25 °C): δ = 175.8, 145.0, 136.7, 136.0, 129.5, 129.5, 128.8, 128.3, 127.0, 58.8,
40.2, 35.7, 31.8, 21.6, 14.9 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C19H21NO3S 366.1134, found
366.1138.
4.2.6.2. (3R,5R)-5-Benzyl-3-methyl-1-tosylpyrrolidin-2-one (8b)
According to the general procedure describe in section 4.2.6, to a solution of 1,4-amino alcohol
2b (457 mg, 1.31 mmol) in dry DMF (10 mL) taken in a round bottom flask (50 mL), PDC (2.5 g, 6.57
mmol) was added to the reaction mixture and stirred for 24 h at room temperature. After workup, the
residue was purified over silica gel column chromatography (6:1 hexane/EtOAc) to afford γ-lactam 8b
as white solid, (366 mg, 83 %). m.p. 122–124 °C. TLC: Rf 0.52 (2:1, hexane/EtOAc). [α]ꢁꢀꢃ.ꢁ = +123.0
(c 0.13, CH2Cl2). IR (CH2Cl2): 3061, 3028, (=C–H), 2930, 2875 (C–H) 1732 (C=O), 1591 (C=C),
16

ACCEPTED MANUSCRIPT
1494, 1454 (C–H), 1355, 1164 (S=O), 702 (=C–H) cm-1. 1H NMR (400 MHz, CDCl3, 25 °C): δ = 8.00
(d, J = 8.4 Hz, 2H), 7.35–7.22 (m, 7H), 4.48–4.41 (m, 1H), 3.84 (dd, J = 12.8, 3.6 Hz, 1H), 2.63 (dd, J
= 12.8, 10.0 Hz, 1H), 2.44 (s, 3H), 2.41–2.35 (m, 1H), 2.20–2.12 (m, 1H), 1.46–1.39 (m, 1H), 1.09 (d,
J = 7.2 Hz, 3H) ppm. 13C NMR (100 MHz, CDCl3, 25 °C): δ = 176.8, 145.0, 136.4, 136.1, 129.5
(2C*), 128.7, 128.3, 126.9, 59.6, 42.6, 36.9, 31.6, 21.6, 16.1 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for
C19H21NO3S 366.1134, found 366.1140.
*higher intensity carbon
4.2.7. General procedure for tosyl deprotection
In a 100 mL round-bottom flask charged with magnetic bead, Mg turnings (10.0 equiv.) and
NH4Cl (5.0 equiv.), dry methanol was added under an inert atmosphere with stirring. A solution of Ts-
γ-lactam (8a or 8b, 1.0 equiv.) in dry methanol was added to the above suspension and stirred at room
temperature for 10 minutes. The reaction mixture was further refluxed at 70 °C for 2 h. After complete
consumption of Ts-γ-lactam (8a or 8b), methanol was concentrated under reduced pressure and the
crude residue was purified over silica gel column chromatography using hexane:EtOAc as eluent.
4.2.7.1. (3S,5R)-5-Benzyl-3-methylpyrrolidin-2-one (9a)
According to the general procedure described in section 4.2.7, N-tosyl- γ-lactam 8a (350 mg,
1.02 mmol) in dry methanol (7 ml) was added to a suspension of Mg turnings (244 mg, 10.2 mmol) and
NH4Cl (273 mg, 5.0 mmol) in methanol (5 mL) taken in a 100 mL round bottom flask. The suspension
was refluxed at 70 °C for 2 h under inert atmosphere and worked up as mentioned in the general
procedure. The crude residue was purified over silica gel column chromatography (1:1 hexane/EtOAc)
to afford 9a (161 mg, 84%) as colorless liquid. TLC: Rf 0.14 (1:3, hexane/EtOAc). [α]ꢁꢀꢃ.ꢆ = −66.1 (c
0.13, CH2Cl2). IR (CH2Cl2): 3230 (N–H), 3028 (=C–H), 2928, 2871 (C–H), 1693 (C=O), 1456 (C–H),
701 (=C–H) cm-1. 1H NMR (400 MHz, CDCl3, 25 °C): δ = 7.33–7.23 (m, 3H), 7.21–7.15 (m, 2H), 5.87
(brs, 1H), 3.85–3.78 (m, 1H), 2.81 (dd, J = 13.2, 5.6 Hz, 1H), 2.71 (dd, J = 13.2, 8.4 Hz, 1H), 2.50–
2.40 (m, 1H), 2.15–2.08 (m, 1H), 1.93–1.86 (m, 1H), 1.18 (d, J = 7.2 Hz, 3H) ppm. 13C NMR (100
MHz, CDCl3, 25 °C): δ = 180.3, 137.6, 129.1, 128.8, 126.8, 53.4, 42.8, 35.1, 34.9, 16.2 ppm. HRMS
(ESI) m/z [M+Na]+ calcd. for C12H15NO 212.1046, found 212.1059.
4.2.7.2. (3R, 5R)-5-Benzyl-3-methylpyrrolidin-2-one (9b)
17

ACCEPTED MANUSCRIPT
According to the general procedure described in section 4.2.7, a solution of N-tosyl-γ-lactam 8b
(200 mg, 0.582 mmol) ) in dry methanol (5 mL), was added to suspension of Mg turnings (140 mg,
5.82 mmol) and NH4Cl (156 mg, 3 mmol) in methanol (5 mL) in a round bottom flask. The suspension
was refluxed at 70 °C with stirring for 2 h under inert atmosphere and worked up as mentioned in the
general procedure. The crude residue was purified over silica gel column chromatography (1:1
EtOAc/hexane) to afford 9b (82 mg, 81%) as colorless liquid. TLC: Rf 0.14 (3:1, EtOAc/hexane).
[α]ꢁꢀꢃ.ꢇ = +83.2 (c 0.13, CH2Cl2). IR (CH2Cl2): 3230 (N–H), 3028 (=C–H), 2961, 2871 (C–H), 1693
1
(C=O), 1557–1456 (C–H), 701 (=C–H) cm-1. H NMR (400 MHz, CDCl3, 25 °C): δ = 7.33–7.22 (m,
3H), 7.20–7.12 (m, 2H), 5.96 (brs, 1H), 3.85–3.78 (m, 1H), 2.81 (dd, J = 13.6, 6.0 Hz, 1H), 2.72 (dd, J
= 13.6, 8.0 Hz, 1H), 2.49–2.40 (m, 1H), 2.14–2.08 (m, 1H), 1.92–1.85 (m, 1H), 1.18 (d, J = 7.2 Hz,
3H) ppm. 13C NMR (100 MHz, CDCl3, 25 °C): δ = 179.6, 137.6, 128.8, 128.8, 126.9, 53.7, 43.3, 37.0,
36.8, 15.9 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C12H15NO 212.1046, found 212.1055.
4.2.8. General procedure for synthesis of 10a and 10b
Dry CH2Cl2 (10 mL) was added to γ-lactam (9a or 9b, 1.0 equiv.) taken in a two-necked round
bottom flask (100 mL) under inert atmosphere. The reaction mixture was cooled to 0 °C and solid
DMAP (0.2 equiv.) was added dropwise to the reaction mixture in single portion with stirring followed
by dropwise addition of Boc2O (2.0 equiv.) using syringe. The reaction mixture was allowed to warm
to room temperature and stirred for 2.5 h. After the consumption of γ-lactam (9a or 9b), the solvent
was evaporated under reduced pressure using. The crude mixture was purified over silica gel column
chromatography using hexane:EtOAc as eluent.
4.2.8.1. (3S,5R)-tert-Butyl 5-benzyl-3-methyl-2-oxopyrrolidine-1-carboxylate (10a)
According to the general procedure described in section 4.2.8, to a solution of γ-lactam 9a (125
mg, 0.682 mmol) in dry CH2Cl2 (10 mL) in a round bottom flask (100 mL), DMAP (16.6 mg, 0.136
mmol) and Boc2O (0.3 mL, 1.364 mmol) were added to the reaction mixture at 0 °C and further stirred
at room temperature for 2.5 h. The solvent was evaporated under reduced pressure and the crude
mixture was purified over silica gel column chromatography (8:1 hexane/EtOAc) to afford 10a (190
mg, 96%) as colorless liquid. TLC: Rf 0.76 (1:1, hexane/EtOAc). [α]ꢁꢀꢂ.ꢈ = +41.0 (c 0.13, CH2Cl2). IR
(CH2Cl2): 3028 (=C–H), 2975, 2936 (C–H), 1785, 1711 (C=O), 1650 (C=C), 1455, 1305 (C–H), 1150
1
(C–O), 702 (=C–H) cm-1. H NMR (400 MHz, CDCl3, 25 °C): δ = 7.35–7.23 (m, 3H), 7.22–7.16 (m,
2H), 4.28 (td, J = 8.8, 3.2 Hz, 1H), 3.13 (dd, J = 13.2, 3.2 Hz, 1H), 2.71 (dd, J = 13.2, 9.2 Hz, 1H),
18

ACCEPTED MANUSCRIPT
2.44–2.35 (m, 1H), 2.04 (dd, J = 12.8, 8.4 Hz, 1H), 1.58 (s, 9H), 1.54–1.42 (m, 1H), 1.15 (d, J = 6.8
Hz, 3H) ppm. 13C NMR (100 MHz, CDCl3, 25 °C): δ = 176.6, 150.1, 137.4, 129.3, 128.7, 126.8, 82.9,
56.9, 39.2, 36.2, 30.7, 28.1, 15.3 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C17H23NO3 312.1570,
found 312.1657.
4.2.8.2. (3R,5R)-tert-Butyl 5-benzyl-3-methyl-2-oxopyrrolidine-1-carboxylate (10b)
According to the general procedure describe in section 4.2.8, to a solution of γ-lactam 9b (70
mg, 0.261 mmol) in dry CH2Cl2 (10 mL) in a round bottom flask (100 mL), DMAP (6.37 mg, 0.05
mmol) and Boc2O (0.12 mL, 0.522 mmol) were added to the reaction mixture at 0 °C and further
stirred at room temperature for 2.5 h. The solvent was evaporated under reduced pressure and the crude
reaction mixture was purified over silica gel column chromatography (8:1 hexane/EtOAc) to afford
10b (109 mg, 99%) as colorless liquid. TLC: Rf 0.76 (1:1, hexane EtOAc). [α]ꢁꢀꢃ.ꢉ = +51.8 (c 0.13,
CH2Cl2). IR (CH2Cl2): 3028 (=C–H), 2928, 2873 (C–H), 1746, 1711 (C=O), 1650 (C=C), 1494, 1455,
1
1305 (C–H), 1150 (C–O), 702 (=C–H) cm-1. H NMR (400 MHz, CDCl3, 25 °C): δ = 7.33–7.23 (m,
3H), 7.22–7.15 (m, 2H), 4.24–4.17 (m, 1H), 3.49 (dd, J = 13.2, 3.2 Hz, 1H), 2.60 (dd, J = 13.2, 10 Hz,
1H), 2.54–2.44 (m, 1H), 2.17–2.09 (m, 1H), 1.60 (s, 9H), 1.40–1.33 (m, 1H), 1.18 (d, J = 7.2 Hz, 3H)
ppm. 13C NMR (100 MHz, CDCl3, 25 °C): δ = 177.2, 150.4, 137.0, 129.5, 128.6, 126.7, 83.1, 57.3,
41.3, 37.3, 30.5, 28.1, 16.6 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for C17H23NO3 312.1570, found
312.1574.
4.2.9. General procedure for the synthesis of 1a and 1b
Dry THF (5mL) was added to N-Boc-γ-lactam (10a or 10b, 1.0 equiv.) taken in a round-bottom
flask (100 mL) with a magnetic bead. An aq.solution of LiOH (5.0 equiv.) and 30% aq.H2O2 (5.0
equiv.) were added to the reaction mixture at room temperature with constant stirring. The reaction
mixture was stirred for further 12 h at room temperature, acidified with an aq. solution of 5% KHSO4
(5 mL), and extracted with EtOAc (20 × 3 mL). The combined organic layers was washed with brine
(10 mL), dried over anhydrous Na2SO4, filtered, evaporated under reduced pressure to afford the
intermediate carboxylic acid which was used as such for benzylation reaction without further
purification. The intermediate crude carboxylic acid was dissolved in dry CH2Cl2 and Et3N (2.0 equiv.)
was added to the mixture with stirring at room temperature. BnBr (1.5 equiv.) was added to the reaction
mixture via a syringe and stirred for 24 h at room temperature. The reaction mixture was quenched with
cold water (5 mL) and extracted with EtOAc (3 × 20 mL). The combined organic layers was washed
19

ACCEPTED MANUSCRIPT
with brine (10 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure.
The crude residue was purified over silica gel column chromatography using hexane:EtOAc as eluent
to afford 1a or 1b.
4.2.9.1. (2S,4R)-Benzyl 4-((tert-butoxycarbonyl)amino)-2-methyl-5-phenylpentanoate (1a)
According to the general procedure described in section 4.2.9, to a solution of N-Boc-γ-lactam
10a (75 mg, 0.259 mmol) in THF (5 mL) in a round bottom flask, an aq.solution of LiOH (31 mg in 2
mL of H2O, 1.29 mmol), 30% aq.H2O2 (1 mL) were added and the reaction mixture was stirred at room
temperature for 12 h. After work up, as mentioned in general procedure, the solvent was evaporated
under reduced pressure to afford the intermediate carboxylic acid. The crude carboxylic acid (75 mg,
0.243 mmol) was dissolved in dry CH2Cl2 (5 mL) and Et3N (0.06 mL, 0.486 mmol) was added to the
mixture with stirring followed by addition of BnBr (0.04 mL, 0.364 mmol). The reaction mixture was
stirred at room temperature for 24 h, worked up as mentioned in the general procedure. The crude
residue was purified over silica gel column chromatography (11:1 hexane/EtOAc) to afford 1a (72 mg,
75%) as pale yellow liquid. TLC: Rf 0.48 (2:1, hexane/EtOAc). [α]ꢁꢀꢂ.ꢃ = +48.9 (c 0.13, CH2Cl2). IR
(CH2Cl2): 3366 (N–H), 3062, 3029 (=C–H), 2973, 2931 (C–H), 1731, 1700 (C=O), 1651 (C=C), 1498,
1
1455 (C–H), 1167 (C–O), 698 (=C–H) cm-1. H NMR (400 MHz, CDCl3, 25 °C): δ = 7.37–7.13 (m,
10H), 5.15–5.04 (m, 2H), 4.30 (d, J = 7.6 Hz, 1H), 3.94–3.83 (m, 1H), 2.77–2.76 (d, J = 5.6 Hz, 2H),
2.69–2.59 (m, 1H), 1.94–1.88 (m, 1H), 1.50–1.41 (m, 1H), 1.38 (s, 9H), 1.17 (d, J = 7.2 Hz, 3H) ppm.
13C NMR (100 MHz, CDCl3, 25 °C): δ = 176.0, 155.2, 137.8, 136.0, 129.4, 128.5, 128.3, 128.1 (2C*),
126.3, 79.1, 66.2, 49.7, 41.3, 37.9, 36.4, 28.3, 17.6 ppm. HRMS (ESI) m/z [M+Na]+ calcd. For
C24H31NO4 420.2145, found 420.2160.
*higher intensity carbon
4.2.9.2. (2R,4R)-Benzyl 4-((tert-butoxycarbonyl) amino)-2-methyl-5-phenylpentanoate (1b)
According to the general procedure described in section 4.2.9, to a solution of N-Boc-γ-lactam
10b (75 mg, 0.259 mmol) in THF (5 mL) in a round bottom flask, an aq.solution of LiOH (31 mg in 2
mL of H2O, 1.29 mmol), 30% aq.H2O2 (1 mL) were added and the reaction mixture was stirred at room
temperature for 12 h. After work up, as mentioned in general procedure, the solvent was evaporated
under reduced pressure to afford the intermediate carboxylic acid. The crude carboxylic acid (75 mg,
0.243 mmol) was dissolved in dry CH2Cl2 (5 mL) and Et3N (0.06 mL, 0.486 mmol) was added to the
mixture with stirring followed by addition of BnBr (0.04 mL, 0.364 mmol). The reaction mixture was
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stirred at room temperature for 24 h, worked up as mentioned in the general procedure. The crude
residue was purified over silica gel column chromatography (11:1 hexane/EtOAc) to afford 1b (70 mg,
72%) as colorless liquid. TLC: Rf 0.48 (2:1, hexane/EtOAc). [α]ꢁꢀꢂ.ꢄ = −43.7 (c 0.13, CH2Cl2). IR
(CH2Cl2): 3366 (N–H), 3063, 3029 (=C–H), 2973, 2931 (C–H), 1729, 1699 (C=O), 1651 (C=C), 1496,
1
1454 (C–H), 1164 (C–O), 697 (=C–H) cm-1. H NMR (400 MHz, CDCl3, 25 °C): δ = 7.37–7.13 (m,
10H), 5.15–5.04 (m, 2H), 4.30 (d, J = 7.6 Hz 1H), 3.96–3.81 (m, 1H), 2.77–2.76 (d, J = 5.6 Hz, 2H),
2.70–2.56 (m, 1H), 1.95–1.88 (m, 1H), 1.53–1.41 (m, 1H), 1.38 (s, 9H), 1.17 (d, J = 7.2 Hz, 3H) ppm.
13C NMR (100 MHz, CDCl3, 25 °C): δ = 176.0, 155.2, 137.8, 136.0, 129.4, 128.5, 128.3, 128.1 (2C*),
126.3, 79.1, 66.2, 49.7, 41.3, 37.9, 36.4, 28.3, 17.6 ppm. HRMS (ESI) m/z [M+Na]+ calcd. for
C24H31NO4 420.2150, found 420.2160.
*higher intensity carbon
Conflict of Interest
Declarations of interest: none
Supplementary data
Electronic Supplementary Information (ESI) available: Copy of 1H and 13C NMR spectra of 1a, 1b, 2a,
2b, 3, 4, 5, 6, 7, 8a, 8b, 9a, 9b, 10a, 10b.
Acknowledgments
This work was supported by the Science and Engineering Research Board, Department of
Science and Technology, Govt. of India under the grant number EMR/2015/001764. The authors are
thankful to Ministry of Human Resource Development (MHRD), Government of India, Indian Institute
of Technology (IIT) Indore and Council of Scientific and Industrial Research (CSIR), India for
research student’s fellowship. We also thank Sophisticated Instrumentation Centre (SIC), IIT Indore for
compound characterization facilities.
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Legends
Table 1. Regioselective ring opening of (S)-2-Benzyl-1-tosylaziridine (4).
Fig. 1. Structure of tubulysin family of antimitotic agents.
Fig. 2. Stereochemical assignments of 8a and 8b through 1H-1H COSY and NOESY studies.
Scheme 1. Retrosynthetic analysis of N-Boc-Tup-OBn (1a) and N-Boc-epi-Tup-OBn (1b).
Scheme 2. Synthesis of (S)-2-benzyl-1-tosylaziridine (4). Reagents and conditions (i) NaBH4, I2,
THF, 0 °C to reflux, 24 h, 96% (ii) Et3N, TsCl, CH3CN, 0 °C to rt, 1 h, 85%.
Scheme 3. Synthesis of N-tosyl-1,4-amino alcohols, 2a–b and 7. Reagents and conditions (i) a. 9-
BBN, THF, rt, 12 h b. 2 M NaOH, H2O2, 0 °C to rt, 12 h, 90%, dr=2 :1 (ii) Boc2O, DMAP, CH2Cl2,
rt, 2 h, 97% (iii) a. 9-BBN, THF, rt, 12 h b. 2 M NaOH, H2O2, 0 °C to rt, 12 h, 70%, dr=2:1 (non-
separable).
Scheme 4. Synthesis of N-Boc-Tup-OBn (1a). Reagents and conditions (i) PDC, DMF, rt, 12 h, 85%
(ii) Mg, NH4Cl, MeOH, reflux, 2 h, 84% (iii) Boc2O, DMAP, DCM, rt, 2 h, 96% (iv) a. LiOH, H2O2,
THF:H2O (1:1), rt, 12 h b. BnBr, Et3N, DCM, rt, 12 h, 75%.
Scheme 5. Synthesis of N-Boc-epi-Tup-OBn (1b). Reagents and conditions (i) PDC, DMF, rt, 12 h,
83% (ii) Mg, NH4Cl, MeOH, reflux, 2 h, 81% (iii) Boc2O, DMAP, DCM, rt, 2 h, 99% (iv) a. LiOH,
H2O2, THF:H2O (1:1), rt, 12 h b. BnBr, Et3N, DCM, rt, 12 h, 72%.
24