Hydrotropic solubilization of paclitaxel: Analysis of chemical structures for hydrotropic property

Article abstract of DOI:10.1023/A:1024458206032

Purpose. To identify hydrotropic agents that can increase aqueous paclitaxel (PTX) solubility and to study the chemical structures necessary for hydrotropic properties so that polymeric hydrotropic agents can be synthesized. Methods. More than 60 candidate hydrotropic agents (or hydrotropes) were tested for their ability to increase the aqueous PTX solubility. A number of nicotinamide analogues were synthesized based on the observation that nicotinamide showed a favorable hydrotropic property. The identified hydrotropes for PTX were used to examine the structure-activity relationship. Results. N,N-Diethylnicotinamide (NNDENA) was found to be the most effective hydrotropic agent for PTX. The aqueous PTX solubility was 39 mg/ml and 512 mg/ml at NNDENA concentrations of 3.5 M and 5.95 M, respectively. These values are 5-6 orders of magnitude greater than the intrinsic solubility of 0.30 ± 0.02 μg/ml. N-Picolylnicotinamide, N-allylnicotinamide, and sodium salicylate were also excellent hydrotropes for PTX. Solubility data showed that an effective hydrotropic agent should be highly water soluble while maintaining a hydrophobic segment. Conclusions. The present study identified several hydrotropic agents effective for increasing aqueous solubility of PTX and analyzed the structural requirements for this hydrotropic property. This information can be used to find other hydrotropic compounds and to synthesize polymeric hydrotropes that are effective for PTX and other poorly water-soluble drugs.

Full text of DOI:10.1023/A:1024458206032

Pharmaceutical Research, Vol. 20, No. 7, July 2003 (© 2003)
Research Paper
increase their water solubility. Poorly water-soluble drugs
have been formulated into micron- or submicron-size particu-
late preparations (3), liposomes and micelles (8), and solid
dispersions (9,10). Cosolvent systems can increase the drug
solubility significantly, but the choices of clinically used sol-
vents are limited to ethylene glycol, dimethylsulfoxide, N,N-
dimethylformamide, Cremophore, and ethanol (11).
In an attempt to find an alternative or supplementary
method for increasing water solubility of poorly soluble
drugs, we have examined the possibility of using hydrotropes.
Hydrotropic agents (hydrotropes) have been used to increase
the water solubility of poorly soluble drugs, and in many in-
stances, the water solubility has increased by orders of mag-
nitude (12). Hydrotropy is a collective molecular phenom-
Hydrotropic Solubilization of
Paclitaxel: Analysis of Chemical
Structures for Hydrotropic Property
1
1
Jaehwi Lee, Sang Cheon Lee,
1
1
Ghanashyam Acharya, Ching-jer Chang, and
1
,2
Kinam Park
Received February 26, 2003; accepted March 27, 2003
Purpose. To identify hydrotropic agents that can increase aqueous enon describing an increase in the aqueous solubility of a
paclitaxel (PTX) solubility and to study the chemical structures nec-
poorly soluble compound by addition of a relatively large
essary for hydrotropic properties so that polymeric hydrotropic
agents can be synthesized.
amount of a second solute (i.e., a hydrotrope) (1). Each hy-
drotropic agent is effective in increasing the water solubility
Methods. More than 60 candidate hydrotropic agents (or hydro-
of selected hydrophobic drugs, and no universal hydrotropic
tropes) were tested for their ability to increase the aqueous PTX
agent has been found to be effective with all hydrophobic
solubility. A number of nicotinamide analogues were synthesized
drugs. Thus, finding the right hydrotropic agents for a par-
ticular hydrophobic drug requires the screening of a large
number of candidate hydrotropes. In this study, we examined
based on the observation that nicotinamide showed a favorable hy-
drotropic property. The identified hydrotropes for PTX were used to
examine the structure–activity relationship.
Results. N,N-Diethylnicotinamide (NNDENA) was found to be the various candidate agents for their abilities to solubilize PTX
most effective hydrotropic agent for PTX. The aqueous PTX solubil- so that the structures of effective agents can be used for iden-
ity was 39 mg/ml and 512 mg/ml at NNDENA concentrations of 3.5
tification of other hydrotropic agents and for synthesis of hy-
M and 5.95 M, respectively. These values are 5–6 orders of magnitude
drotropic polymers.
greater than the intrinsic solubility of 0.30 ± 0.02 ␮g/ml. N-
Picolylnicotinamide, N-allylnicotinamide, and sodium salicylate were
also excellent hydrotropes for PTX. Solubility data showed that an
MATERIALS AND METHODS
effective hydrotropic agent should be highly water soluble while
maintaining a hydrophobic segment.
Materials
Conclusions. The present study identified several hydrotropic agents
PTX was obtained from Samyang Genex Corp. (Taejeon,
effective for increasing aqueous solubility of PTX and analyzed the
South Korea). 6-Hydroxynicotinic acid, 1,1Ј-carbonyl-
structural requirements for this hydrotropic property. This informa-
diimidazole (CDI), diethylamine, 3-picolylamine, nicotinoyl
tion can be used to find other hydrotropic compounds and to synthe-
chloride hydrochloride, allylamine, acetic anhydride, pyri-
size polymeric hydrotropes that are effective for PTX and other
dine, and triethylamine were purchased from Aldrich Chemi-
cal Company (Milwaukee, WI) and used without further pu-
rification. Methylene chloride was dried and distilled over
calcium hydride. Tetrahydrofuran (THF) was distilled from
sodium benzophenone before use. n-Hexane, diethyl ether,
chloroform, and methanol were of reagent grade. All other
chemicals were purchased from Fisher Scientific (Pittsburgh,
PA). Freshly prepared distilled water was used throughout.
poorly water-soluble drugs.
KEY WORDS: hydrotropic agents; solubilization; poorly water-
soluble drug; paclitaxel; structure–activity relationship.
INTRODUCTION
Poor water solubility of many drugs and drug candidates
causes significant problems in producing formulations with
sufficiently high bioavailability (1–3). Paclitaxel (PTX)
presents a good example of the importance of water solubil-
ity. Its use in cancer therapy has been hindered by its low
water solubility (4), which has required special formulations
utilizing ethanol and Cremophore EL (polyoxyethylated cas-
tor oil), which has significant side effects such as hypersensi-
tivity reactions (5). Testing PTX in preclinical tumor model
systems is also difficult (6). In addition, the cosolvent mixture
is diluted in isotonic saline solution before intravenous ad-
ministration, and the diluted solution remains stable for only
several hours (7). For hydrophobic drugs with poor water
solubility, including PTX, several methods have been used to
Synthesis and Characterization of Nicotinamide Analogues
Instrumental Analysis
1
13
H and C NMR spectra were obtained using a Bruker
ARX300 spectrometer at 300 MHz and 75 MHz, respectively.
Elemental analysis was performed on a Perkin Elmer Series
II CHNS/O Analyzer 2400. UV-VIS spectra were obtained by
a Beckman DU® 640 spectrophotometer. Electrospray ion-
ization mass spectrometry (ESI-MS) assay was done using a
FinniganMAT LCQ (ThermoFinnigan Corp, San Jose, CA).
The electrospray needle voltage was set at 4.5 kV, the heated
capillary voltage was set to 10 V, and the capillary tempera-
ture to 225°C. Typical background source pressure was 1.2 ×
1
Departments of Pharmaceutics and Biomedical Engineering, Pur-
−
5
10
torr. The sample flow rate was approximately 10 ␮l/min.
due University, West Lafayette, Indiana 47907.
2
To whom correspondence should be addressed. (e-mail: kpark@ Nitrogen gas was used for drying. The LCQ was scanned to
purdue.edu)
2,000 amu for these experiments.
0
724-8741/03/0700-1022/0 © 2003 Plenum Publishing Corporation
1022

Hydrotropic Solubilization of Paclitaxel
1023
N-Picolylnicotinamide
ס 7.2 Hz, 6H), 3.32 (q, J ס 7.2 Hz, 4H), 6.34 (d, J ס 9.1 Hz,
1H), 7.45 (dd, J ס 2.4, 9.1 Hz, 1H), 7.50 (d, J ס 2.4 Hz, 1H);
To a solution of 3-picolylamine (0.1 mol) and pyridine 13
C NMR (DMSO-d ) ␦ 13.3, 41.0, 114.4, 119.3, 135.5, 139.7,
6
(
0.2 mol) in dry methylene chloride (600 ml) was added nico-
+
161.9, 166.8; ESI-MS, m/z 195 ([M+H] ); analysis calculated
tinoyl chloride hydrochloride (0.1 mol) at 0°C. The reaction
mixture was stirred at room temperature for 24 h under ni-
trogen. After 24 h, the solvent was removed under reduced
pressure, and the crude product was dissolved in water, neu-
tralized with sodium bicarbonate, and extracted with chloro-
form (3 × 200 ml). The solution was dried over anhydrous
magnesium sulfate. The solvent was removed at reduced pres-
sure, and the product was isolated by column chromatogra-
for C H N O : C, 61.84; H, 7.27; N, 14.42. Found: C, 61.73;
H, 7.16; N, 14.47.
1
0
14
2
2
2-Hydroxy-N,N-Diethylnicotinamide
To a stirred suspension of 2-hydroxynicotinic acid (0.216
mol) in THF (700 ml) was added CDI (0.216 mol) in one
portion. The reaction mixture was stirred at reflux under ni-
trogen. After 24 h, diethylamine (0.323 mol) was added drop-
wise to the stirred suspension of N-(2-hydroxynicotinyl)-
imidazole in THF at reflux. The reaction was maintained for
phy on a silica gel using THF/n-hexane. Yield was 80%; m.p.
1
1
05–107°C; ␭max(THF) 256 nm; H NMR (DMSO-d ) ␦ 4.52
6
(
d, J ס 5.8 Hz, 2H), 7.34 (dd, J ס 4.8, 7.7 Hz, 1H), 7.49 (dd,
J ס 4.8, 8.1 Hz, 1H), 7.72–7.75 (m, 1H), 8.20–8.23 (m, 1H),
.46 (dd, J ס 1.0, 4.8 Hz, 1H), 8.58 (d, J ס 2.4 Hz, 1H), 8.70
dd, J ס 1.0, 4.8 Hz, 1H), 9.06 (d, J ס 2.0 Hz, 1H), 9.29 (t, J
24 h under nitrogen. After cooling of the reaction mixture to
8
room temperature, the solution was concentrated under re-
duced pressure. The pale yellow precipitate was filtered,
(
1
3
ס 5.8 Hz, 1H); C NMR (DMSO-d ) ␦ 40.4, 123.4, 129.6,
6
washed with diethyl ether, and dried in vacuo. Yield was 70%;
1
34.7, 135.0, 135.1, 148.2, 148.3, 148.5, 148.9, 151.9, 164.9; ESI-
1
m.p. 90–92°C; ␭max(THF) 313 nm; H NMR (DMSO-d ) ␦
+
6
MS, m/z 214 ([M+H] ); analysis calculated for C H N O: C,
12
11
3
1
7
.00 (t, J ס 7.2 Hz, 3H), 1.07 (t, J ס 7.2 Hz, 3H), 3.12 (q, J ס
.2 Hz, 2H), 3.35 (q, J ס 7.2 Hz, 2H), 6.20 (m, 1H), 7.38 (dd,
6
7.59; H, 5.20; N, 19.71. Found: C, 67.73; H, 5.10; N, 19.51.
1
3
J ס 2.4, 6.9 Hz, 1H), 7.41 (dd, J ס 2.4, 6.9 Hz, 1H); C NMR
DMSO-d ) ␦ 12.8, 14.1, 38.4, 42.2, 104.6, 129.3, 136.1, 138.4,
N-Allylnicotinamide
(
1
6
+
59.2, 166.2; ESI-MS, m/z 195 ([M+H] ); analysis calculated
To a stirred solution of allylamine (0.168 mol) in dry
methylene chloride (500 ml), nicotinoyl chloride hydrochlo-
ride (0.112 mol) and triethylamine (0.225 mol) were added at
for C H N O : C, 61.84; H, 7.27; N, 14.42. Found: C, 62.14;
10 14
2
2
H, 7.18; N, 14.42.
0
°C. The reaction mixture was stirred at room temperature
for 24 h under nitrogen. After 24 h, the solvent was removed N-Picolylacetamide
under reduced pressure. The brown liquid was dissolved in
To a stirred solution of acetic anhydride (0.069 mol) in
THF (50 ml), a solution of 3-picolylamine (0.046 mol) in THF
20 ml) was added dropwise at room temperature. The reac-
tion mixture was stirred for 5 h under nitrogen. After 5 h, an
excess of water was added, and the aqueous solution was
neutralized with 1 N sodium hydroxide. The solvent was re-
moved at reduced pressure, and the product was isolated by
distilled water and neutralized with sodium bicarbonate, fol-
lowed by extraction with chloroform (3 × 200 ml). The solvent
was removed at reduced pressure, and the crude product was
column chromatographed with THF/n-hexane on a silica gel
(
to produce a light yellow liquid. Yield was 85%; ␭max(THF)
1
2
60 nm; H NMR (DMSO-d ) ␦ 3.89–3.94 (m, 2H), 5.05–5.20
6
(m, 2H), 5.82–5.94 (m, 1H), 7.47 (dd, J ס 5.0, 8.3 Hz, 1H),
column chromatography on a silica gel using THF/n-hexane.
8
.20 (m, 1H), 8.68 (dd, J ס 1.7, 5.0 Hz, 1H), 8.87 (t, J ס 5.6
1
13
Yield was 85%; ␭max(THF) 257 nm; H NMR (CDCl ) ␦ 1.90
Hz, 1H), 9.04 (d, J ס 1.7 Hz, 1H); C NMR (DMSO-d ) ␦
3
6
(s, 3H), 4.29 (d, J ס 5.7 Hz, 2H), 7.16 (dd, J ס 4.8, 8.2 Hz,
4
1.5, 115.3, 123.3, 129.9, 134.9, 135.0, 148.5, 151.7, 164.6; ESI-
+
1H), 7.39 (s, 1H), 7.54 (m, 1H), 8.34 (dd, J ס 1.4, 4.8 Hz, 1H),
MS, m/z 163 ([M+H] ); analysis calculated for C H N O: C,
9
10
2
1
3
8
1
.35 (d, J ס 1.4, 1H); C NMR (CDCl ) ␦ 22.7, 40.7, 123.4,
6
6.65; H, 6.21; N, 17.27. Found: C, 66.53; H, 6.07; N, 16.97.
3
+
34.2, 135.5, 148.1, 148.6, 170.4; ESI-MS, m/z 151 ([M+H] );
analysis calculated for C H N O: C, 63.98; H, 6.71; N, 18.65.
Found: C, 63.85; H, 6.61; N, 18.54.
8
10
2
6
-Hydroxy-N,N-Diethylnicotinamide
To a stirred suspension of 6-hydroxynicotinic acid (0.108
mol) in THF (600 ml) was added CDI (0.108 mol) in one
portion. The reaction mixture was stirred at reflux under ni-
trogen. After 24 h, diethylamine (0.216 mol) was added drop-
NMR Measurement
1
The H NMR spectra of NNDENA in D O in the con-
2
wise to the stirred suspension of N-(6-hydroxynicotinyl)- centration range of 0.0025 to 1.36 M were obtained. The ratios
imidazole in THF at reflux. The reaction was further main- of chemical shifts of nicotinamide protons to the chemical
tained for 24 h under nitrogen. After cooling of the reaction shift of HDO protons (4.63 ppm) were monitored with in-
mixture to room temperature, 1N sodium hydroxide solution creasing NNDENA concentrations.
(
120 ml) was added. THF was evaporated, and the aqueous
solution of the crude product was washed with diethyl ether
5 × 200 ml). The aqueous solution was then neutralized with
N hydrochloric acid to pH 7 and extracted with chloroform
3 × 200 ml). The solution was dried over anhydrous magne- a fixed volume of the hydrotrope solution. This mixture was
Solubility Study
Excess PTX was added to screw-capped vials containing
(
1
(
sium sulfate. The solvent was removed at reduced pressure, stirred using a magnetic stirring bar for 24 h at 37°C. An
and the product was isolated by column chromatography on a aliquot of the sample was collected, and within 5 s, it was
silica gel using THF/n-hexane. Yield was 65%; m.p. 113– filtered through a 0.2-␮m nylon membrane. This immediate
1
1
15°C; ␭max(THF) 253 nm; H NMR (DMSO-d ) ␦ 1.09 (t, J filtering process prevented any possible formation of PTX
6

1
024
Lee et al.
a
Table I. Paclitaxel Solubilities in the Presence of Various Hydrotropic Agents at 37°C
b
Hydrotropic agent
Concentration used (M)
PTX solubility (mg/ml)
Standard deviation
c
None (PTX solubility in pure water)
N,N-Diethylnicotinamide
N-Picolylnicotinamide
N-Allylnicotinamide
Sodium salicylate
—
0.0003
0.0000
0.600
1.205
0.385
0.514
0.037
0.012
0.026
0.006
0.071
0.008
0.003
0.005
0.031
0.080
0.063
0.070
0.080
0.028
0.004
0.002
0.008
0.002
0.006
0.002
0.003
0.002
0.001
0.002
3.5
3.5
3.5
3.5
2.9
3.5
3.5
3.5
3.5
3.5
3.5
2.5
3.5
3.5
3.5
1.5
2.5
3.5
2.0
2.5
2.5
3.5
2.0
3.5
3.5
3.5
3.5
1.0
39.071
29.435
14.184
5.542
3.199
2.009
1.771
1.344
1.341
1.282
1.084
0.720
0.694
0.658
0.552
0.500
0.481
0.300
0.241
0.220
0.216
0.071
0.050
0.038
0.030
0.015
0.012
0.009
2-Methacryloyloxyethyl phosphorylcholine
Resorcinol
N,N-Dimethylnicotinamide
N-Methylnicotinamide
Butylurea
Pyrogallol
N-Picolylacetamide
Procaine HCl
Nicotinamide
Pyridine
3-Picolylamine
Sodium ibuprofen
Sodium xylenesulfonate
Ethyl carbamate
6-Hydroxy-N,N-diethylnicotinamide
Sodium p-toluenesulfonate
Pyridoxal hydrochloride
1
-Methyl-2-pyrrolidone
Sodium benzoate
-Pyrrolidone
2
Ethylurea
N,N-Dimethylacetamide
N-Methylacetamide
Isoniazid
a
Mean ± SD, n ס 3, except for PTX solubility in pure water, where n ס 10.
The concentrations less than 3.5 M represent the maximum solubilities of the hydrotropic agent.
The aqueous PTX solubility is 0.30 ± 0.02 ␮g/ml.
b
c
particles as a result of the temperature decrease to ambient. Milford, MA) at 25°C. The mobile phase consisted of aceto-
The filtrate was diluted with acetonitrile (1:1), and the con- nitrile–water (45:55 v/v) with a flow rate of 1.0 ml/min. A
centration of PTX was determined by an isocratic reverse- diode array detector was set at 227 nm and linked to Chem-
phase HPLC (Agilent 1100 series, Agilent Technologies, Wil- Station software for data analysis. The PTX concentrations in
mington, DE) using a Symmetry column (Waters Corp., the samples were obtained from a calibration curve.
Fig. 1. Paclitaxel solubility as a function of the molar concentration Fig. 2. The ratio of chemical shifts of nicotinamide protons to the
of N,N-diethylnicotinamide. The solubility of paclitaxel at 5.95 M of chemical shift of HDO protons in D O as a function of the concen-
2
N,N-diethylnicotinamide is 512.6 mg/ml. The inserted plot shows the tration of N,N-diethylnicotinamide. H-2, H-4, H-5, and H-6 indicate
paclitaxel solubility as a function of the log concentration of N,N- the proton position of the nicotinamide ring of N,N-diethylnico-
diethylnicotinamide.
tinamide.

Hydrotropic Solubilization of Paclitaxel
1025
orders of magnitude. Other hydrotropes that resulted in an
aqueous PTX solubility more than 1 mg/ml were resorcinol,
N,N-dimethylnicotinamide, N-methylnicotinamide, butyl-
urea, pyrogallol, and N-picolylacetamide. The PTX solubility
of 0.3 mg/ml by ethyl carbamate appears to be much smaller
than that by NNDENA but still represents a 1,000-fold in-
crease.
Another 35 agents not listed in Table I showed paclitaxel
solubilities of 0.005 mg/ml (or 5 ␮g/ml) or less. They are,
in the descending order of solubilizing effect, nipecotamide
(3.5 M), citric acid (2.0 M), sodium gentisate (1.0 M), N-
isopropylacrylamide (1.5 M), methylurea (3.5 M), 1,3-
diamino-2-hydroxypropane-N,N,NЈ,NЈ-tetramethylacetate
Fig. 3. Chemical structure of paclitaxel.
RESULTS AND DISCUSSION
(3.0 M), thiourea (2.5 M), 1-methylnicotinamide iodide (1.0
M), ␣-cyclodextrin (0.15 M), sodium thiocyanate (8.6 M),
urea (6.0 M), caffeine (0.1 M), glyceryl triacetate (0.2 M),
Paclitaxel Solubility by Hydrotropes
Because of its noted therapeutic potential and very low glycerin (3.5 M), adenosine (0.005 M), ␥-cyclodextrin (0.17
water solubility, PTX was chosen to examine hydrotropic M), ␤-cyclodextrin (0.02 M), diisopropylnicotinamide (0.05
properties of various agents in this study. The exact mecha- M), pyridine-3-sulfonic acid (1.0 M), o-benzoic acid sulfimide
nisms involved in solubilization of PTX and other poorly (0.01 M), 2,6-pyridinedicarboxamide (0.0025 M), 3,4-
soluble drugs by hydrotropic agents are not clearly under- pyridinedicarboxamide (0.025 M), 4-aminosalicylic acid
stood, so it is difficult to predict the structural requirements of (0.005 M), L-tryptophan (0.05 M), salicylaldoxime (0.1 M),
hydrotropes suitable for solubilizing PTX. For this reason, a sucrose (2.0 M), L-lysine (2.0 M), 4-aminobenzoic acid sodium
large number of candidate agents were screened. Table I lists salt (2.5 M), D-sorbitol (3.0 M), sodium L-ascorbate (3.0 M),
the agents tested and the corresponding water solubilities of sodium propionate (3.5 M), sodium acetate (4.0 M), 2-hy-
PTX measured in the presence of those agents. Our prelimi- droxy-N,N-diethylnicotinamide (0.2 M), 2-hydroxy-N-
nary study suggested that even good hydrotropes required a picolylnicotinamide (0.0035 M), and 6-hydroxy-N-
hydrotropic concentration of approximately 3 M. For this rea- picolylnicotinamide (0.08 M).
son, a concentration of 3.5 M was used for all agents to com-
pare their hydrotropic properties under the same condition.
Hydrotropic Property of NNDENA
The concentrations of some agents listed in Table I are
The hydrotropic property of NNDENA was examined in
smaller than 3.5 M because of their limited solubility. Table I more detail. Figure 1 shows the paclitaxel solubility as a func-
clearly identifies a number of hydrotropic agents effective for tion of the NNDENA concentration. NNDENA at 5.95 M
increasing the water solubility of PTX.
increased the PTX concentration to 512 mg/ml (equivalent to
The aqueous solubility of PTX at 37°C was determined 0.6 M because the molecular weight of PTX is 854 g/mol), and
to be 0.30 ± 0.02 ␮g/ml. Thus, a PTX concentration of 39 this corresponds to 10 NNDENA molecules per paclitaxel
mg/ml by N,N-diethylnicotinamide (NNDENA) in Table I molecule dissolved. At the concentration of hydrotropes used
indicates more than 100,000-fold increase in aqueous solu- in Table I, however, more than 100 hydrotropic agents are
bility. Equally effective was N-picolylnicotinamide. N- necessary for effective solubilization of PTX. The inserted
Allylnicotinamide, sodium salicylate, and 2-methacryloyloxy- plot in Fig. 1 shows the solubility increase of PTX as a func-
ethyl phosphorylcholine increased the PTX solubility by four tion of the log concentration of NNDENA in the range of
Table II.
Conc. used
(M)
PTX solubility
(mg/ml)
Hydrotropic agent
Structure
3.5
2.0
0.2
39.07
0.98
N,N-Diethylnicotinamide
0.001
a
a
6
-Hydroxy-N,N-diethylnicotinamide
2.0
0.2
0.24
0.00
2
-Hydroxy-N,N-diethylnicotinamide
a
The maximum solubility of the hydrotropic agent.

1
026
Lee et al.
0.0028 to 0.5 M. The water solubility of PTX begins to in- Structural Analysis of Hydrotropic Property for PTX
crease at 0.11 M of NNDENA, although the increase is small
compared to higher concentrations of NNDENA.
To gain insights into the structural requirements neces-
sary for hydrotropy, chemical structures of various agents
listed in Table I were analyzed for their ability to increase
aqueous PTX solubility. The structure of PTX is shown in Fig.
Because the dissolution of PTX in NNDENA is expected
to occur through association of NNDENA molecules, self-
association of NNDENA molecules was examined using
NMR. Figure 2 shows the NNDENA concentration depen-
dence of the ratio of chemical shifts of nicotinamide protons
3. A few common features of good hydrotropes for PTX were
identified.
High Water Solubility of Hydrotropic Agents
to the chemical shift of HDO protons in D O. As the con-
2
centration of NNDENA increased to about 0.1 M, the ratios
The main criterion for effective hydrotropy is high water
of chemical shifts of all protons of the nicotinamide ring solubility of the hydrotropic agent. If the water solubility is
started to decrease. The data indicate that NNDENA self- low (e.g., less than 2 M), the hydrotropic property is not ob-
associates via the vertical plane-to-plane interaction of the served to be significant. At 2.0 M, PTX solubility was higher
aromatic rings. The crossover point in Fig. 2 can be described in NNDENA than in 6-hydroxy-N,N-diethylnicotinamide.
as the minimum hydrotropic concentration (MHC), which is The PTX solubility in NNDENA was greatly increased with
the threshold concentration of self-aggregate formation. The increasing the NNDENA concentration. 2-Hydroxy-N,N-
MHC value of NNDENA in the aqueous media was esti- diethylnicotinamide with the maximum water solubility of
mated to be 0.12 M. Interestingly, this MHC value is almost only 0.2 M did not have any PTX-solubilizing effect. The
the same as the concentration of 0.11 M where NNDENA following example shows the importance of water solubility of
begins to exhibit the solubilizing ability for PTX in aqueous hydrotropic agents on increasing aqueous PTX solubility
solutions.
(Table II).
Table III.
Conc.
used (M)
PTX solubility
(mg/ml)
Hydrotropic agent
Structure
Sodium salicylate
3.5
5.54
1.28
Pyrogallol (1,2,3-
trihydroxybenzene)
3
.5
Nicotinamide
3.5
3.5
0.69
0.55
3-Picolylamine
Nipecotamide
3.5
0.005
N,N-Dimethylacetamide
N-Isopropylacrylamide
3.5
1.5
0.015
0.004
a
a
1,3-Diamino-2-hydoxypropane-
3
.0
0.004
N,N,NЈ,NЈ-tetramethylacetate
a
The maximum solubility of the hydrotropic agent.

Hydrotropic Solubilization of Paclitaxel
1027
Table IV.
Conc. used
Hydrotropic agent
PTX solubility
(mg/ml)
(
Conc. used)
(M)
Structure
N,N-Diethylnicotinamide
N,N-Dimethylnicotinamide
3.5
39.07
1.77
3.5
N-Methylnicotinamide
3.5
3.5
1.34
0.69
Nicotinamide
a
1
-Methylnicotinamide iodide
1.0
0.003
0.001
a
N,N-Diisopropylnicotinamide
0.05
a
The maximum solubility of the hydrotropic agent.
Table V.
PTX solubility
Hydrotropic agent
Conc. used)
(
(mg/ml)
Structure
a
Sodium xylenesulfonate (2.5 M)
Sodium p-toluenesulfonate (2.5 M)
0.48
0.22
a
1
2
-Methyl-2-pyrrolidone (3.5 M)
-Pyrrolidone (3.5 M)
0.07
0.04
N-Methylnicotinamide (3.5 M)
Nicotinamide (3.5 M)
1.34
0.69
a
The maximum solubility of the hydrotropic agent.

1
028
Lee et al.
Table VI.
agents lacking a significant hydrophobic component are not
effective at all. The following examples show the importance
of hydrophobic groups in promoting hydrotropic properties.
Conc.
used
(M)
PTX
solubility
(mg/ml)
Hydrotropic agent
Structure
Importance of Pyridine and Benzene Rings
Almost all highly effective hydrotropic agents listed in
Table I have either a pyridine or a benzene ring in their
structures. Molecules without such rings in their structures
were not as effective as those containing the ring structures.
As shown in Table III, nicotinamide and 3-picolylamine dis-
played about the same hydrotropic property, while nipecot-
amide, which has a saturated ring structure, is less than 1%
effective as nicotinamide. Other agents without pyridine or
benzene ring that had very small hydrotropic effect are urea
and its alky derivatives (methyl-, ethyl-, and butylurea), glyc-
erin, thiourea, N-isopropylacrylamide, N-methylacetamide,
Resorcinol
(1,3-dihydroxybenzene)
3.5
3.5
2.009
1.282
Pyrogallol
(1,2,3-trihydroxybenzene)
The agents that did not show any appreciable hydrotro- N,N-dimethylacetamide, sodium thiocyanate, and 1,3-
pic properties have low water solubilities, as expected. Ex- diamino-2-hydroxypropane-N,N,N’,N’-tetramethylacetate.
amples are glyceryl triacetate (0.2 M), caffeine (0.1 M), sali-
cylaldoxime (0.1 M), 3,4-pyridinedicarboxamide (0.025 M), Maximum Hydrophobicity without Losing Water Solubility
o-benzoic acid sulfimide (0.01 M), 4-aminosalicylic acid (0.005
The hydrotropic properties of nicotinamide derivatives
M), adenosine (0.005 M), and 2,6-pyridinedicarboxamide
show a positive correlation with the hydrophobicity of mol-
(
0.0025 M). Those agents have low water solubility and thus
ecules as long as water solubility is not lost. As shown in Table
IV, NNDENA showed more than a 20 times higher hydro-
tropic property than N,N-dimethylnicotinamide at the same
concentration. N,N-Dimethylnicotinamide, in turn, was more
effective than N-methylnicotinamide, and N-methylnic-
otinamide was twice as effective as nicotinamide. 1-Methyl-
nicotinamide iodide was too hydrophilic to be hydrotropic.
The poor hydrotropic property of N,N-diisopropylnicotin-
amide results from its poor water solubility, which is only
showed almost no hydrotropic effect.
High Hydrophobicity of Hydrotropic Agents
For those agents having high water solubilities, the hy-
drotropic property increases as the hydrophobicity of the
molecule increases. Poorly soluble organic drugs are all hy-
drophobic, i.e., nonpolar, and do not interact with water mol-
ecules through hydrogen bonding. Thus, the presence (or in-
sertion) of hydrophobic drug molecules in water (known as
hydrophobic hydration) causes a direct perturbation of water,
i.e., an alteration in the hydrogen-bonding state of water mol-
ecules. Water structure formers, such as sucrose and sorbitol,
inhibit dissolution of poorly soluble drugs, whereas water
structure disruptors such as nicotinamide increase the solu-
0.05 M.
Increase in the Hydrotropic Property by a Factor of Two
with a Methyl Group on the Ring
At the same concentration, sodium xylenesulfonate was
bility by destroying clusters of associated water molecules and more hydrotropic than sodium p-toluenesulfonate (Table V).
releasing water of solvation (13). Thus, effective hydrotropic A similar trend was seen with 1-methyl-2-pyrrolidone and
agents are expected to destabilize water structure and at the 2-pyrrolidone. In both examples, the presence of one methyl
same time interact with poorly soluble drugs. Hydrotropic group increased the hydrotropic property of the molecule by
Table VII.
Conc. used
(M)
PTX solubility
(mg/ml)
Hydrotropic agent
Structure
N-Picolylnicotinamide
3.5
3.5
3.5
29.44
14.18
1.77
N-Allylnicotinamide
N,N-Dimethylnicotinamide

Hydrotropic Solubilization of Paclitaxel
1029
a factor of 2. The same result was observed for N-methylni- More Effective Hydrotropic Property by One Long
cotinamide and nicotinamide.
Hydrophobic Chain Than by Two Shorter
Hydrophobic Chains
Decrease in Hydrotropic Property with a Hydroxyl Group
The hydrophilicity of a molecule can be increased by
As shown in Table VII, the high hydrotropic properties
attaching hydroxyl groups to the molecule. Increase in hydro- of N-picolylnicotinamide and N-allylnicotinamide suggest
philicity comes with a reduction in the hydrotropic properties that one longer carbon chain is better than two shorter carbon
of the molecule. For example, pyrogallol, which is more hy- chains, e.g., one allyl group vs. two methyl groups. A signifi-
drophilic than resorcinol, has lower hydrotropic property cant increase in PTX solubility by N-picolylnicotinamide may
(Table VI).
be partly related to the presence of another pyridine ring that
Table VIII.
Conc. used PTX solubility
Hydrotropic agent
Sodium salicylate
(M)
(mg/ml)
Structure
3.5
5.54
2
-Methacryloyloxyethyl
phosphorylcholine
a
2.9
3.20
0.72
a
Procaine и HCl
2.5
a
Sodium ibuprofen
1.5
0.50
0.48
a
Sodium xylenesulfonate
2.5
a
Sodium p-toluenesulfonate
2.5
0.22
a
Pyridoxal hydrochloride
Sodium benzoate
2.5
0.22
0.05
a
2.0
a
Isoniazid
1.0
0.01
a
The maximum solubility of the hydrotropic agent.

1
030
Lee et al.
is shown to be essential for hydrotropic property, as discussed picolylnicotinamide maintains their hydrotropic properties
above.
(14). Table I suggests several candidate hydrotropic agents
that can be synthesized into polymers. NNDENA, N-
picolylnicotinamide, N-allylnicotinamide, and sodium salicy-
late appear to be good candidates for making polymeric hy-
drotropic agents for PTX.
Hydrophobic Interaction between Hydrotropic Agent
and Solute
Aliphatic derivatives of urea were studied for their ef-
fects on increasing the water solubility of PTX. Butylurea ACKNOWLEDGMENTS
shows the highest solubilizing effect among the analogues
studied, which suggests that as the hydrophobicity decreases,
the hydrotropic property also decreases. Urea is known to
break up the hydrogen-bonded water molecule clusters sur-
rounding nonpolar solute molecules. The poor hydrotropic
property of urea suggests that disruption of water structure
alone, without substantial interaction with solute, is not
enough for effective hydrotropy.
This study was supported in part by National Institute of
Health through grant GM 65284, Samyang Corporation, and
NSF Industry/University Center for Pharmaceutical Process-
ing Research.
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use of low-molecular-weight hydrotropic agents at high con-
centrations may result in undesirable side effects, including
toxicity and cell damages. This concern, however, may be
alleviated by preparing polymeric forms of hydrotropes. For
this reason, we synthesized polymers of hydrotropic agents
effective for PTX, and hydrotropic polymers were as effective
as the low-molecular-weight counterpart in increasing the wa-
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1
1