N-Aryl Amides as Chemical Exchange Saturation Transfer Magnetic Resonance Imaging Contrast Agents

Article abstract of DOI:10.1002/chem.202002415

Chemical exchange saturation transfer (CEST) MRI has recently emerged as a versatile molecular imaging approach in which diamagnetic compounds can be utilized to generate an MRI signal. To expand the scope of CEST MRI applications, herein, we systematical

Full text of DOI:10.1002/chem.202002415

Accepted Article
Accepted Article
Title: N-aryl amides as chemical exchange saturation transfer magnetic
resonance imaging contrast agents
Authors: Xuekang Cai, Jia Zhang, Jiaqi Lu, Long Yi, Zheng Han,
Shuixing Zhang, Xing Yang, and Guanshu Liu
This manuscript has been accepted after peer review and appears as an
Accepted Article online prior to editing, proofing, and formal publication
of the final Version of Record (VoR). This work is currently citable by
using the Digital Object Identifier (DOI) given below. The VoR will be
published online in Early View as soon as possible and may be different
to this Accepted Article as a result of editing. Readers should obtain
the VoR from the journal website shown below when it is published
to ensure accuracy of information. The authors are responsible for the
content of this Accepted Article.
To be cited as: Chem. Eur. J. 10.1002/chem.202002415
Link to VoR: https://doi.org/10.1002/chem.202002415
01/2020

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N-aryl amides as chemical exchange saturation transfer magnetic
resonance imaging contrast agents
Xuekang Cai† [a,b], Jia Zhang† [c,d], Jiaqi Lu[e], Long Yi[b], Zheng Han[c,d], Shuixing Zhang*[f], Xing
Yang*[a,g], Guanshu Liu*[c,d]
[a]
[b]
Department of Nuclear Medicine, Peking University First Hospital, Beijing, China. E-mail: yangxing2017@bjmu.edu.cn
State Key Laboratory of Organic-Inorganic Composites and Beijing Key Lab of Bioprocess, Beijing University of Chemical Technology, 15 Beisanhuan East
Road, Chaoyang District, Beijing, China.
[c]
Russell H. Morgan Department of Radiology, The Johns Hopkins University School of Medicine, Baltimore, Maryland, United States. Email:
guanshu@mri.jhu.edu
[d]
[e]
[f]
[b]
†
F.M. Kirby Research Center for Functional Brain Imaging, Kennedy Krieger Institute, Baltimore, Maryland, United State.
Department of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, Maryland, United States.
Department of Radiology, The First Affiliated Hospital of Jinan University, Guangzhou, Guandong, China. Email: shui7515@126.com
Institute of Medical Technology, Peking University, Beijing, China
These authors contributed equally to this work.
Supporting information for this article is given via a link at the end of the document
the natural substrates of the enzyme as the imaging probe. In this
context, a variety of enzymes, including cytosine deaminase
(CDase)[10], carboxypeptidase G2[11], proteases[12], protein kinase
Abstract: Chemical exchange saturation transfer (CEST) MRI has
recently emerged as a versatile molecular imaging approach in which
diamagnetic compounds can be utilized to generate MRI signal. To
expand the scope of CEST MRI applications, herein, we
systematically investigated the CEST properties of N-aryl amides with
different N-aromatic substitution, revealing their chemical shifts (4.6-
5.8 ppm) and exchange rates (up to thousands s-1) are favorable to
be used as CEST agents as compared to alkyl amide. As the first
proof-of-concept study, we used CEST MRI to detect the enzymatic
metabolism of the drug acebutolol directly by its intrinsic CEST signal
without any chemical labeling. Our study implies that N-aryl amides
may enable the label-free CEST MRI detection of the metabolism of
many N-aryl amide-containing drugs and a variety of enzymes that act
on N-aryl amide, greatly expanding the scope of CEST MR molecular
imaging.
, , , g-
A[13] deoxycytidine kinase[14] alkaline phosphatases[15]
glutamyl transferase (GGT)[16], sulfatase[17], and esterase[18], have
been studied. All these studies showed CEST MRI was capable
of detecting enzymes in small qualities simply because enzymes
can convert the substrates to products continuously without the
loss of the enzyme activity, providing a practical way to leverage
the sensitivity of CEST MRI.
To expand the scope of CEST MRI applications, it is essential
to screen, characterize, select and optimize new types of
exchangeable protons and functional groups with desirable CEST
properties. In such an investigation, two CEST characteristics are
needed to be optimized, i.e., chemical shift (also called CEST
offset) and exchange rate, both of which are sensitive to the
chemical and physical environment (i.e., pH) around the
exchangeable protons. Ideally, the chemical shift (∆ω) of the
exchangeable proton should be well-separated from the bulk
water resonance frequency (set to 0 ppm in MRI convention), and
the exchange rate (kex) should be fast as long as in the slow to
intermediate range, i.e., kex <= ∆ω. For instance, it has been
shown previously that natural phenol[19] and heterocyclic amine[20]
can be used for generating CEST MRI contrast with highly shifted
chemical shifts. In the present study, we systematically studied
the CEST properties of aryl amide, another important type of
exchangeable protons, and expanded our knowledge further in
the development of CEST-based MR molecular imaging.
Amide proton is one of the most abundant type of exchangeable
protons in biomolecules in the human body. With a frequency
offset of ~3.5-3.7 ppm apart from water resonance, endogenous
alkyl amide has been used for amide proton transfer (APT)
imaging, which has entered clinical testing for diagnosing tumors
and stroke[21]. The success of APT imaging is attributed to the
high concentration of endogenous amide protons in mobile
proteins and peptides, despite the exchange rate of alkyl-amide
protons typically is slow (~ 30-40 s-1)[22], which indeed is not
favorable for CEST detectability. Interestingly, recent studies
showed a few iodinated X-ray agents, such as Iopamidol and
Magnetic resonance imaging (MRI) is one of the most
commonly used medical imaging modalities. The capacity of MRI
for precision diagnosis depends on highly sensitive MRI contrast
agents that are designated specifically to the detection of
particular molecular targets. Among the currently available MRI
contrast mechanisms suitable for molecular imaging, chemical
exchange saturation transfer (CEST) is relatively new and
versatile, holding great promise for future MR molecular imaging.
Instead of paramagnetic metals, CEST MRI utilizes water
exchangeable protons to generate specific, sensitive MRI
contrast.[1],[2] In particular, the CEST contrast is produced by
applying radiofrequency (RF) pulses continuously at the chemical
shift of exchangeable protons to completely null the MR signal of
these protons (namely saturation). Subsequentially, via proton
exchange, these saturated protons will relocate to the
surrounding water and transfer the saturation to water, resulting
in a detectable level of (water) MRI signal attenuation, which is
named CEST contrast.[3] Increasing evidence shows that, with the
capability of directly using a broad spectrum of diamagnetic
compounds as contrast agents, CEST MRI paves a new avenue
to accomplish MR molecular imaging[4]. To date, CEST MRI has
been applied to detect anti-cancer drugs,[5] metabolites,[6] amino
acids,[7] sugars,[8] and many other important bioorganic agents[9]
directly without chemical labeling (namely label-free). Recently,
the label-free CEST MRI has also been used to detect enzymatic
activity[10]. As the active sites of many biologically relevant
enzymes are directly on the exchangeable functional group, one
can utilize the difference in the CEST signals between the
substrate and product to directly detect the activity of the enzyme
of interest. Such an approach, unlike the widely used fluorometric
and colorimetric methods, completely eliminates the requirement
for chemical labeling and hence favors in vivo detection by using
iopromide, can be used as CEST MRI agents, attributable to their
[23],[24]
inherently carried aryl amide protons
. Compared to alkyl
amide, these aryl amide protons have larger frequency offsets
(i.e., Δω ~ 5.2-5.6 ppm) and a much faster exchange rate (kex is
~2560 s-1 [23],[24]. Inspired by these studies, we hypothesized that
)
many other N-aryl-amide-containing compounds can be
developed as CEST contrast agents with more favorable CEST
properties than alkyl amide-containing agents. To this end, we
first systematically investigated the CEST properties of a variety
of N-aryl amides, characterized the effect of N-aromatic
substitution (Scheme 1); and then, as the first proof-of-concept
study, we used the inherent N-aryl amide CEST signal of
1
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acebutolol, a drug for treating hypertension and arrhythmias, to
assess the enzymatic metabolism of acebutolol.
320 s-1), likely attributed to electron-donating and steric effect. A
relatively large Δω of 5.6 ppm was seen for compound 15,
indicating the formation of intramolecular hydrogen-bonding
between the N-H and ester. To determine the effects of pKa
(acidity) of amide protons on the CEST properties, we also
calculated the pKa value of each compound (supplementary
information Figure S4.4) using Advanced Chemistry
Development (ACD/Labs) Software V11.02 (1994-2020
ACD/Labs). The results show significantly negative correlation
between pKa and kex (and maximal MTRasym values), indicating
that lower pKa (or higher acidity) of aryl amide protons would
result in faster exchange and thereby higher CEST signal,
whereas no correlation with Δω (supplementary information
Figure S4.5). Table 2 also includes three compounds (16-18)
whose benzene ring is replaced by heterocyclic rings. The
heterocyclic substitutions were effective to increase the chemical
shifts (up to 5.8 ppm for compound 18). Compound 18 also
exhibited a much faster exchange rate, likely due to dramatically
reduced pKa of the N-H.
Overall, chemical modification only has a limited effect on the
chemical shift of amide protons, i.e., Δω varies from 4.6 to 5.8
ppm for all the structures studied. In contrast, the exchange rate
and hence the maximum CEST contrast varied greatly. Among all
the compounds studied, we found that compound 14 has the
highest CEST signal (i.e., 9.7 % in MTRasym , 10 mM at pH 7.4 and
37 oC), which is 1.7-fold higher than that of the model compound
succinanilic acid (compound 1) and more than 3-fold higher than
that of compound 2.
O
O
R2
Alkyl
R1
N
H
R1
N
H
N-Alkyl amide
3.7 ppm
Slow exchange
N-Aryl amide
Larger chemical shift
Faster exchange rate
Scheme 1. The rational design of the present study.
We first examined the CEST signal of succinanilic acid, which
represents the basic structure of N-aryl amide (Figure 1A, with
reference to a structure with R1 = propionic acid and R2=H in
Scheme 1). The Z-spectrum and MTRasym plot of succinanilic acid
(10 mM in PBS, pH =7.5) were shown in Figure 1B, revealing a
detectable CEST signal peaked at 4.9 ppm. As shown in Figure
1C, N-aryl amide has a strong pH dependence, with the maximum
CEST signal =0.9, 2.1, 5.6, and 7.8%, at pH = 6.5, 7.0 and 7.5,
and 8.0 respectively. The increase of the CEST signal with pH
(Figure 1D) is attributed to the faster exchange rates at higher
pHs (Figure 1D insert, supplementary method), where higher
concentrations of OH catalyze the proton exchange.
Table 1. CEST properties of para- and meta-substituted N-aryl amides
Compound
Signal [ppm]
Contrast [%]
kex [s-1]
4.9
5.6
544
4.8
4.8
2.7
5.8
104
590
Figure 1. CEST characteristics of succinanilic acid. (a) Chemical structure; (b)
CEST signal as depicted by Z-spectrum and MTRasym plot; (c) MTRasym plots at
different pH; (d) pH dependence of the peak CEST MRI signal (quantified by
MTRasym at 4.9 ppm) and exchange rates (insert). The CEST experiments were
performed on a Bruker 9.4 T vertical bore MR scanner with a 25-mm birdcage
transmitter/receiver volume RF coil. Unless otherwise noted, all CEST
measurements were acquired using a 3-second long, continuous wave (CW)
RF pulse with saturation field strength B1= 3.6 µT (details provided in
supplementary information S2).
5
6.2
6.5
7.5
820
883
923
5.2
5.2
To elucidate the effect of chemical modifications, we
synthesized eighteen different N-aryl amides with different
substitutions and studied their CEST properties (supplementary
information S3 and S4). The measured CEST offsets, maximum
CEST contrast, and exchange rate are listed in Table 1 and Table
2. Of note, all compounds contain a carboxyl group so as to
ensure adequate water solubility >10 mM.
5.0
7.3
846
Table 1 summarizes the compounds with para- and meta-
substituted anilides. The largest Δω was obtained in compounds
5 and 6, with para-electron withdrawing substitutions, i.e., 5.2 ppm.
The para- substitutions only have a small effect on kex. An electron
withdrawing substitution accelerates the exchange rate kex (5-6),
while an electron-donating substitution slows kex (2). Compared
to succinanilic acid (1), meta-substituted compounds (7-9)
showed similar Δω (i.e., 5.0 ppm) and slightly faster kex (i.e., 648
-846 s-1).
5.0
5.0
7.5
7.5
684
677
The inherent CEST signal of N-aryl amide allows label-free MRI
detection of the pharmacokinetics and pharmacodynamics of
many N-aryl-amide-containing drugs. To demonstrate it, we
chose acebutolol, a β-adrenergic receptor-blocker that is
In Table 2, ortho- Me/MeO- substituted anilides (10 and 11)
showed slightly reduced Δω (down to 4.6 ppm) and kex (down to
2
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signal change of acebutolol at different concentrations after the
incubation with 10 mg/mL esterase for 5 min (supplementary
information S6), and fitted the data with the Michaelis-Menten
kinetic model. The estimated KM is 93 mM (Figure 2d), which is
on the same order as the values in the literature, e.g., 29.5 and
238 mM for human liver microsomes and carboxylesterase 1,
respectively[26a], demonstrating the ability of CEST MRI to detect
enzyme and drug metabolism.
Table 2. CEST properties of N-aryl amides with ortho- and other
heterocyclic substitutions
Compound
Signal [ppm]
Contrast [%]
kex[s-1]
4.6
5.5
320
H
N
H
N
H
N
(a)
4.6
4.6
7.1
8.6
733
Hydrolysis
OH
OH
Acetylation
OH
O
O
O
O
O
N
H
N
H
H2
N
O
O
O
1124
Acebutolol
Acetolol
Diacetolol
(b)
MTRasym
MTRasym
MTRasym
0.08
0.08
0.08
0.04
0.04
0.04
0.12
0.09
0.06
0.03
0.00
-0.03
0.12
0.09
0.06
0.03
0.00
0.12
0.09
0.06
0.03
0.00
-0.03
T2w CEST
T2w CEST
T2w CEST
0
0
0
4.8
8.3
1003
-0.03
8
6
4
2
0
8
6
4
2
0
8
6
4
2
0
Offset (ppm)
Offset (ppm)
Offset (ppm)
4.8
5.6
5.2
5.4
5.8
9.7
7.1
7.7
3.7
3.5
905
636
747
511
15
10
5
0.05
0.04
0.03
0.02
0.01
0.00
(c)
(d)
Vmax = 0.14 mM s-1
KM = 93 mM
0
0
10
20
30
40
50
0
2
4
6
8
10
Acebutolol concentration (mM)
[Esterase] mg/ml
Figure 2. CEST MRI detection of the esterase-catalyzed hydrolysis of
acebutolol. (a) In vivo metabolic pathway of acebutolol; (b) MTRasym plots of
acebutolol, acetolol, and diacetolol in PBS (pH 7.4); (c) CEST signal (quantified
by MTRasym at 5 ppm) of 20 mM acebutolol incubated with esterase at different
concentrations for 1 h; (d) Fitting CEST MRI data with the Michaelis-Menten
kinetic model, in which the initial reaction velocity was calculated using the
CEST signal change at each concentration of acebutolol after the first 5 min’s
incubation. For (b)-(d), all data are averaged on the measurement of three
samples, the error bars in (c) and (d) shows the standard deviations. Conditions
for CEST MRI acquisition: B1=3.6 µT/4 s, pH=7.4, 37 oC.
4838
commonly used to treat hypertension and arrhythmia[25]
.
Acebutolol is a N-aryl amide with two substituents on aromatic
ring. While the structure of acebutolol is not a direct derivative of
the library compounds listed in Tables 1 and 2, it also contains
substituents that tend to enhance the CEST effect as observed in
compounds 5 and 11. Indeed, the CEST signal of acebutolol is
similar to succinic acid derivatives, with a Δω of 5.0 ppm, kex of
959 s-1, and MTRasym=6.3±0.7% for 10 mM acebutolol in PBS, at
In summary, we systematically investigated the CEST
properties of a library of N-aryl amides with different chemical
modifications. Compared to alkyl amide, N-aryl amides have
further separated chemical shift from water in the range of 4.6-5.8
ppm and much faster exchange rates (up to thousands s-1). The
CEST properties of N-aryl amide can be fine-tuned using different
chemical modifications. We further demonstrated the application
of the CEST signal of N-aryl amide in the label-free MRI detection
o
pH 7.4 and 37 C. In the liver and intestines, acebutolol is first
metabolized to acetolol by carboxylesterases, and subsequently
to diacetolol by N-acetyltransferases (Figure 2a).[26] The first step
of metabolism results in the disappearance of CEST signal at 5
ppm due to the conversion of N-aryl amide to amine, while the
second step leads to the re-appearance of the CEST signal
(Figure 2b). The CEST signals of the substrate acebutolol and
final product diacetolol are nearly identical, whereas that of the
intermediate product acetolol (synthesized by acid-assisted
hydrolysis of acebutolol, supplementary information S5) is
markedly different, providing a practical way to detect the
conversion of acebutolol to acetolol and monitor acetolol-related
toxicity caused by N-acetyltransferase deficiency, e.g., lupus
erythematosus (DILE).[27]
of the esterase-catalyzed metabolism of
a N-aryl amide-
containing drug acebutolol. Our study demonstrated that many N-
aryl amides can be directly used as CEST agents with favorable
CEST properties, which may greatly expand the scope of the
biomedical applications of CEST MR molecular imaging.
Acknowledgements
X.Yang is supported by NSFC (21877004) and Clinical Medicine
Plus X-Young Scholars Project of Peking University
(PKU2020LCXQ029).
To demonstrate the capacity of CEST MRI to monitor the
enzymatic conversion of acebutolol to acetolol, we first incubated
20 mM acebutolol with carboxylesterase (from porcine liver,
Sigma, E3019) at different concentrations for one hour and
measured the CEST MRI signal of the reaction systems afterward.
As shown in Figure 2c, the decrease of the CEST signal at 5 ppm
was proportional to the enzyme concentration. At the highest
enzyme concentration (i.e., 10 mg/mL), the intensity of CEST
signal decreased from 10.4±0.7% to 5.4±1.0%, accounting for a
48% relative signal change. The minimal concentration of enzyme
allowing for a reliable CEST detection was determined to be c.a.
1 mg/mL (~ 6 µM, MW=168 kD). We then measured the CEST
Keywords: MRI contrast agent • N-aryl amide • CEST MRI •
enzyme detection• acebutolol metabolism
[1] K. M. Ward, A. H. Aletras and R. S. Balaban, J Magn Reson 2000, 143, 79-
87.
[2] J. Y. Zhou and P. C. M. van Zijl, Progress in Nuclear Magnetic Resonance
Spectroscopy 2006, 48, 109-136.
[3] P. C. M. Van Zijl, C. K. Jones, J. Ren, C. R. Malloy and A. D. Sherry, Proc.
Natl. Acad. Sci. USA 2007, 104, 4359-4364.
[4] a) Z. Han and G. Liu, Wiley Interdiscip Rev Nanomed Nanobiotechnol
2019, 11, e1551; b) G. Liu, S. Ray Banerjee, X. Yang, N. Yadav, A. Lisok, A.
3
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Jablonska, J. Xu, Y. Li, M. G. Pomper and P. van Zijl, Nat Biomed Eng 2017,
1, 977-982.
[5] a) Y. Li, H. Chen, J. Xu, N. N. Yadav, K. W. Chan, L. Luo, M. T. McMahon,
B. Vogelstein, P. C. van Zijl, S. Zhou and G. Liu, Oncotarget 2016, 7, 6369-
6378; b) H. Liu, A. Jablonska, Y. Li, S. Cao, D. Liu, H. Chen, P. C. Van Zijl, J.
W. Bulte, M. Janowski, P. Walczak and G. Liu, Theranostics 2016, 6, 1588-
1600; c) L. L. Lock, Y. Li, X. Mao, H. Chen, V. Staedtke, R. Bai, W. Ma, R. Lin,
Y. Li, G. Liu and H. Cui, ACS Nano 2017, 11, 797-805.
[6] K. Cai, M. Haris, A. Singh, F. Kogan, J. H. Greenberg, H. Hariharan, J. A.
Detre and R. Reddy, Nat Med 2012, 18, 302-306.
[7] G. Liu, M. Moake, Y. E. Har-el, C. M. Long, K. W. Chan, A. Cardona, M.
Jamil, P. Walczak, A. A. Gilad, G. Sgouros, P. C. van Zijl, J. W. Bulte and M.
T. McMahon, Magn Reson Med 2012, 67, 1106-1113.
[8] T. Jin, H. Mehrens, P. Wang and S. G. Kim, Neuroimage 2016, 143, 82-90.
[9] D. Ryoo, X. Xu, Y. Li, J. A. Tang, J. Zhang, P. C. M. van Zijl and G. Liu,
Anal Chem 2017, 89, 7758-7764.
[10] G. Liu, Y. Liang, A. Bar-Shir, K. W. Chan, C. S. Galpoththawela, S. M.
Bernard, T. Tse, N. N. Yadav, P. Walczak, M. T. McMahon, J. W. Bulte, P. C.
van Zijl and A. A. Gilad, J Am Chem Soc 2011, 133, 16326-16329.
[11] Y. Jamin, T. R. Eykyn, E. Poon, C. J. Springer and S. P. Robinson, Mol
Imaging Biol 2014, 16, 152-157.
[12] M. Haris, A. Singh, I. Mohammed, R. Ittyerah, K. Nath, R. P. Nanga, C.
Debrosse, F. Kogan, K. Cai, H. Poptani, D. Reddy, H. Hariharan and R.
Reddy, Sci Rep 2014, 4, 6081.
[13] R. D. Airan, A. Bar-Shir, G. Liu, G. Pelled, M. T. McMahon, P. C. van Zijl,
J. W. Bulte and A. A. Gilad, Magn Reson Med 2012, 68, 1919-1923.
[14] Z. Han, Y. Li, J. Zhang, J. Liu, C. Chen, P. C. van Zijl and G. Liu, Cancer
Res 2019, 79, 2775-2783.
[15] I. Daryaei, M. M. Ghaffari, K. M. Jones and M. D. Pagel, ACS Sens 2016,
1, 857-861.
[16] S. Sinharay, E. A. Randtke, K. M. Jones, C. M. Howison, S. K. Chambers,
H. Kobayashi and M. D. Pagel, Magn Reson Med 2017, 77, 2005-2014.
[17] S. Sinharay, G. Fernandez-Cuervo, J. P. Acfalle and M. D. Pagel,
Chemistry 2016, 22, 6491-6495.
[18] R. Thomas, Chembiochem 2016, 17, 2293.
[19] a) X. Yang, X. Song, Y. Li, G. Liu, S. R. Banerjee, M. G. Pomper and M.
T. McMahon, Angew Chem Int Ed Engl 2013, 52, 8116-8119; b) J. Zhang, Y.
Yuan, Z. Han, Y. Li, P. C. M. van Zijl, X. Yang, J. W. M. Bulte and G. Liu,
Biosens Bioelectron 2019, 141, 111442.
[20] X. Yang, X. Song, S. Ray Banerjee, Y. Li, Y. Byun, G. Liu, Z. M.
Bhujwalla, M. G. Pomper and M. T. McMahon, Contrast Media Mol Imaging
2016, 11, 304-312.
[21] H. Zhu, C. K. Jones, P. C. van Zijl, P. B. Barker and J. Zhou, Magn Reson
Med 2010, 64, 638-644.
[22] a) J. Zhou, D. A. Wilson, P. Z. Sun, J. A. Klaus and P. C. Van Zijl, Magn
Reson Med 2004, 51, 945-952; b) H. Y. Heo, Z. Han, S. Jiang, M. Schar, P. C.
M. van Zijl and J. Zhou, Neuroimage 2019, 189, 202-213.
[23] S. Aime, L. Calabi, L. Biondi, M. De Miranda, S. Ghelli, L. Paleari, C.
Rebaudengo and E. Terreno, Magn Reson Med 2005, 53, 830-834.
[24] S. Sinharay, E. A. Randtke, C. M. Howison, N. A. Ignatenko and M. D.
Pagel, Mol Imaging Biol 2018, 20, 240-248.
[25] W. H. Frishman, American Heart Journal 1987, 113, 1190-1198.
[26] a) K. Muta, T. Fukami and M. Nakajima, Biochem Pharmacol 2015, 98,
659-670; b) U. Rakibe, R. Tiwari, A. Mahajan, V. Rane and P. Wakte, J Pharm
Anal 2018, 8, 357-365.
[27] S. Fenniche, A. Dhaoui, F. B. Ammar, R. Benmously, H. Marrak and I.
Mokhtar, Skin Pharmacol Physiol 2005, 18, 230-233.
4
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Entry for the Table of Contents
CEST MRI
N-aryl amide
20
15
10
5
1.0
0.8
0.6
0.4
0.2
0.0
O
R2
N
R1
Chemical shift ↑
H
0
8
6
4
2
0
-2 -4 -6 -8
Exchange rate ↑↑
Offset (ppm)
Label free MRI detection
MTRasym
MTRasym
0.08
0.08
0.04
0
0.04
0
T2w CEST
T2w CEST
H
N
H
N
0.10
0.08
0.06
0.04
0.02
0.00
Acebutolol
Acetolol
Enzymatic
activity
OH
OH
O
O
O
Drug
metabolism
N
H
H2
N
O
O
6
4
2
0
Offset (ppm)
Acebutolol
Acetolol
The chemical exchange saturation transfer (CEST) MRI properties of N-aryl amides were systematically investigated. With relatively
large chemical shifts and fast exchange rate, this type of diamagnetic, biocompatible compounds can be directly used as CEST
agents for label-free MRI detection of enzymatic activity and drug metabolism.
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