Optimization of the Ugi reaction using
parallel synthesis and automated liquid
handling
Authors and Affiliations:
Jean-Claude Bradley* Khalid Baig Mirza and Kevin Owens (Drexel University); Tom
Osborne (Mettler-Toledo); Antony Williams (ChemSpider)
Introduction
The Ugi reaction has proved to be a convenient way to quickly create diverse libraries of
compounds (1-3). It involves the reaction of an amine, an aldehyde, a carboxylic acid and
an isonitrile typically in methanol at room temperature. The Ugi reaction has often been
used as a tool in the synthesis of pharmacologically active molecules and we have
exploited it to quickly access compounds in the search for new anti-malarial agents (4). It
has been observed that Ugi products sometimes precipitate in pure form from the reaction
mixture (1,5). This is a very fortunate outcome since the reaction can then be easily scaled
up without requiring costly purification procedures such as chromatography. It would be
most beneficial to optimize the Ugi product yield as obtained directly from filtering the
reaction mixture without further treatment. To this end we utilized a 48-slot Mettler-Toledo
MiniBlock (6) equipped with filtration tubes. A Mettler-Toledo MiniMapper (7) automated
liquid handler was used to deliver the reagents and solvent. The parameters of interest were
the concentration, the solvent composition and the excess of some of the reagents.
Experimental
The MiniMapper automated liquid handler was programmed to deliver liquids in the
following sequence to empty filter tubes in a 48-position MiniBlock: additional solvent,
furfurylamine (2M in methanol), benzaldehyde (2M in methanol), boc-glycine (2M in
methanol) and t-butylisocyanide (2M in methanol). The default addition volume was 100
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microliters. If a reagent is noted as being added in excess, 120 microliters were delivered.
The MiniBlock was then placed on a shaker for 16 hours before being filtered using house
vacuum. Two washes are performed by adding methanol (1 mL) to each tube followed by
15 min shaking before filtering off. The tubes were then dried under high vacuum in a
dessicator for at least 30 min. The yield was calculated from the increase in weight of the
filter tubes. The purity was assessed by H NMR for one sample from each solvent system
and concentration. The reactions were run in triplicate (8-10) and the average yields are
reported in Table 1.
Characterization of the Ugi product tert-butyl (2-{[2-(tert-butylamino)-2-oxo-1-
phenylethyl](furan-2-ylmethyl)amino}-2-oxoethyl)carbamate : white solid; m.p.(11)
202-204 C; 1H NMR (12, spectrum) (500MHz, ppm, CDCl3) 1.33 (s, 9H), 1.45 (s, 9H),
4.21 (m, 2H), 4.49 (d, J=18Hz, 1H), 4.50 (d, J=18Hz, 1H), 5.47 (s, 1H), 5.60 (s, 1H), 5.62
(s, 1H), 5.89 (s, 1H), 6.10 (s, 1H), 7.19 (s, 1H), phenyl 7.21-7.37 (m, 5H); 13C NMR (13,
spectrum) (500MHz, ppm, CDCl3) 28.3, 28.6, 42.3, 42.8, 51.7, 62.9, 79.5, 107.7, 110.4,
128.5, 128.7, 129.6, 134.7, 141.9, 149.8, 155.7, 168.4 170.2; IR (14, spectrum) (Ʋmax cm-
1 ATR): 1645,1673,1699, 3331; FAB-HRMS (15, spectrum) (calculated for C24H33N3O5
m/z 444.2498 [M+H], obtained 444.2517.)
Data Analysis
The precipitate yield data was analyzed using the single-factor or two-factor (with
replicates) analysis of variance (ANOVA) tools available in Microsoft Excel. (16) For
those variables found to be statistically significant in the ANOVA analysis, Fisher's Least
Significant Difference (LSD) test was used to determine the specific experimental settings
exhibiting significant differences in yield of the Ugi precipitate (17). All significance tests
were performed at the 95% confidence level.
Results and Discussion
Figure 1 summarizes the effect of solvent composition on the yield of the Ugi precipitate.
A single factor ANOVA indicates a significant difference in average yield for the four
solvents studied at a reagent concentration of 0.2M. Fisher's LSD test indicates that the
precipitate yields obtained in methanol and ethanol are not statistically different at the 95%
confidence level (signified by the bar over the names of solvents in the figure); the yield of
Ugi precipitate is significantly greater in ethanol and methanol than in acetonitrile, which
in turn is significantly greater than that in THF.
Figure 1: Summary of the effect of solvent composition on the yield of Ugi precipitate at a
reagent concentration of 0.2M. The listed solvents include 40% methanol at this
concentration.
Considering the effect of reagent concentration, the data in Table 1 show a precipitous drop
in yield was found when going from 0.2M to 0.07M using methanol, ethanol and
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acetonitrile as the solvent; the yields were comparable in methanol at the 0.2M and 0.4M
concentrations. Note that Ugi reactions are generally run in the 0.5-2M range (2). However,
when using an automated liquid handler, it may be difficult to start with reagent solutions
much more concentrated than 2M, thus placing a practical upper limit on the final
concentration at about 0.5M. The results of this optimization study suggest caution in
interpreting the success or failure of Ugi reactions below 0.2M, since non-linear effects are
apparently at play.
Figure 2 summarizes the effect of solvent composition and reagent excess on the yield of
Ugi precipitate at a reagent concentration of 0.2M. The two-factor ANOVA results indicate
a statistically significant solvent and reagent excess effect. In addition, there is a
statistically significant interaction between the two variables. Details of the interaction are
uncovered in figure 2 which shows that the yield of precipitate is affected by the solvent
used. Again note that the bars over the factors in the figure indicate the yields are not
statistically different at the 95% confidence level. For both ethanol and methanol the best
yields are obtained with the imine or isonitrile in excess, while for acetonitrile,
significantly improved results are obtained with the amine, aldehyde or isonitrile in excess.
As noted above in the discussion of the results in Table 1, the overall yield of precipitate is
low for THF as the solvent, and only an excess of imine leads to a statistically significant
improvement in precipitate yield. These different patterns of improved yield for the four
solvents is what leads to the statistically significant interaction effect, and may be an
indicator of different interaction of the Ugi product with the different solvents. Although
the numerical results in Table 1 show the best yield was obtained in 0.4M methanol with
1.2 equivalents of excess imine, 0.2M acetonitrile/methanol and 0.2M ethanol/methanol
mixtures also provided significant yields. Note particularly that all of the THF/methanol
mixtures at 0.2M gave very poor results. The Ugi reaction has been successfully carried out
in a variety of solvents, including THF (2). Note that the low yields here do not imply that
the Ugi reaction did not take place in THF mixtures, only that a precipitate was not formed.
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Figure 2: Summary of the effect of solvent composition and reagent excess on the yield of
Ugi precipitate at a reagent concentration of 0.2M. The listed solvents include 40%
methanol at this concentration.
Figure 3 summarizes the effect of reagent concentration and reagent excess on the yield of
Ugi precipitate using methanol as the solvent. The two-factor ANOVA results indicate a
statistically significant concentration and reagent excess effect. As described above, the
yield of precipitate at reagent concentrations of of 0.2M and 0.4M are significantly greater
than at 0.07M (but do not differ from each other). In addition, there is a statistically
significant interaction between the two variables under study. Note that the pattern of the
improvement in precipitate yield with identity of the excess reagent is generally the same at
the higher reagent concentrations.
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Figure 3: Summary of the effect of concentration and reagent excess on the yield of Ugi
precipitate in methanol solvent.
Overall, the results described in figures 2 and 3 demonstrate that there were interactions
between the solvent choice, reagent concentration and identity of the reagent in excess, but
overall these were more subtle than the straight solvent and concentration effects. Others
have found similar results when optimizing a Ugi reaction (18).
Conclusion
The optimization experiment found the highest yield of the 48 runs to be 66% from 0.4 M
methanol with 1.2 eq. of imine. This is significantly above the 49% yield obtained from the
initial reaction under equimolar concentration at 0.4 M in methanol. Good reproducibility
of the precipitate yields was obtained in these replicate experiments, allowing for subtle
interaction effects to be positively identified.
Table 1: Yield of Ugi precipitate as a function of solvent composition, reagent
concentration and reagent excess. All averages and standard deviations are for N=3
replicate reactions.
Run Solvent concentration (M) ratios average yields std dev
1 methanol 0.4 Eqimolar 49.41 1.18
2 methanol 0.4 Amine xs 57.31 2.58
3 methanol 0.4 Ald xs 58.62 2.66
4 methanol 0.4 Acid xs 55.39 5.69
5 methanol 0.4 tBUIC xs 58.66 1.86
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6 methanol 0.4 Imine xs 66.17 3.17
7 methanol 0.2 Eqimolar 50.84 1.37
8 methanol 0.2 Amine xs 47.20 2.29
9 methanol 0.2 Ald xs 50.47 0.97
10 methanol 0.2 Acid xs 54.15 1.21
11 methanol 0.2 tBUIC xs 57.83 4.06
12 methanol 0.2 Imine xs 57.49 2.59
13 methanol 0.07 Eqimolar 0.53 0.42
14 methanol 0.07 Amine xs 1.09 0.23
15 methanol 0.07 Ald xs 9.06 1.49
16 methanol 0.07 Acid xs 15.03 2.08
17 methanol 0.07 tBUIC xs 9.88 1.53
18 methanol 0.07 Imine xs 3.23 2.37
19 ethanol/methanol (60/40) 0.2 Eqimolar 47.99 3.67
20 ethanol/methanol (60/40) 0.2 Amine xs 48.29 1.73
21 ethanol/methanol (60/40) 0.2 Ald xs 50.39 1.46
22 ethanol/methanol (60/40) 0.2 Acid xs 54.04 0.90
23 ethanol/methanol (60/40) 0.2 tBUIC xs 55.12 2.51
24 ethanol/methanol (60/40) 0.2 Imine xs 57.91 2.50
25 ethanol/methanol (87/13) 0.07 Eqimolar 2.25 1.96
26 ethanol/methanol (87/13) 0.07 Amine xs 1.61 0.68
27 ethanol/methanol (87/13) 0.07 Ald xs 7.18 0.69
28 ethanol/methanol (87/13) 0.07 Acid xs 1.43 1.05
29 ethanol/methanol (87/13) 0.07 tBUIC xs 1.80 1.66
30 ethanol/methanol (87/13) 0.07 Imine xs 0.41 0.43
31 acetonitrile/methanol (60/40) 0.2 Eqimolar 37.09 1.86
32 acetonitrile/methanol (60/40) 0.2 Amine xs 41.00 3.01
33 acetonitrile/methanol (60/40) 0.2 Ald xs 42.88 1.50
34 acetonitrile/methanol (60/40) 0.2 Acid xs 32.09 3.22
35 acetonitrile/methanol (60/40) 0.2 tBUIC xs 41.00 0.85
36 acetonitrile/methanol (60/40) 0.2 Imine xs 37.95 1.36
37 acetonitrile/methanol (87/13) 0.07 Eqimolar 0.94 0.66
38 acetonitrile/methanol (87/13) 0.07 Amine xs 1.84 0.17
39 acetonitrile/methanol (87/13) 0.07 Ald xs 0.90 0.74
40 acetonitrile/methanol (87/13) 0.07 Acid xs 0.98 0.68
41 acetonitrile/methanol (87/13) 0.07 tBUIC xs 1.20 0.43
42 acetonitrile/methanol (87/13) 0.07 Imine xs 1.01 0.88
43 THF/methanol (87/13) 0.2 Eqimolar 7.89 1.35
44 THF/methanol (87/13) 0.2 Amine xs 8.15 0.98
45 THF/methanol (87/13) 0.2 Ald xs 8.45 1.58
46 THF/methanol (87/13) 0.2 Acid xs 10.75 0.72
47 THF/methanol (87/13) 0.2 tBUIC xs 7.29 0.91
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48 THF/methanol (87/13) 0.2 Imine xs 11.76 0.62
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