Ca2? and Zn2? complexes of pyrroglutamate analogs detected by ESI–MS
681
3
m
Cl-Ph
H
173.0 (CO ester), 178.0 (d, J(C,P) = 7.4 Hz, CO lactam)
ppm. 31P NMR (CDCl3, 121 MHz) d 26.2 ppm.
H2N PO3R2
N
3 or 4
RO2C
CO2R
PO3R2
n
2,2-Bis-(diethoxyphosphoryl)-pyrrolidin-5-one
(5) 1H
R=Et
n=1,2
NMR(CDCl3,300 MHz)d1.36(t,3J(H,H) = 7.1 Hz,12H,49
CH3), 2.55 (m, 4H, 29 CH2), 4.24 (m, 8H, 49 OCH2), 5.84
(br s, 1H, NH) ppm. 13C NMR (CDCl3, 125 MHz) d 16.5
(CH3),24.8(CH2),29.2(CH2),58.7(t,1J(C,P) =152.5 Hz,C),
177.7(COlactam)ppm. 31PNMR(CDCl3, 121 MHz)d19.8,
19.9 ppm.
Scheme 2 Synthesis of molecules 3 and 4
H2N
PO3R2
PO3R2
m
Cl-Ph
H
N
5
PO3R2
R2O3P
RO2C
R=Et
Mass spectrometry
Scheme 3 Synthesis of molecule 5
All measurements were carried out in acetonitrile (HPLC
grade, Fisher Scientific). The salts used for the complexa-
tion studies were calcium perchlorate and zinc perchlorate.
They were purchased from Alpha (highest quality avail-
able) and were vacuum-dried over P2O5.
3
OCH2), 4.22 (q, J(H,H) = 7.2 Hz, 2H, OCH2), 6.41 (br s,
1H, NH) ppm. 13C NMR (CDCl3, 125 MHz) d 14.3 (CH3),
29.3 (CH2), 29.8 (CH2), 30.6 (CH2), 33.8 (CH2), 61.1
(CH2), 62.2 (CH2), 64.9 (C), 172.6 (CO ester), 173.2 (CO
ester), 177.1 (CO lactam).
Mass spectra were obtained on a ESI–MS triple quad-
rupole, TSQ Quantum (Thermo FINNIGAN, San Jose, CA,
USA). The spectra were recorded in the positive ion mode.
The ESI source was operated at 3.3 kV, and the heated
capillary set at 260°C. Different temperatures and tube lens
voltages and CID sources have been tested to set the
optimal conditions for detecting complexes and not adducts
statistically formed in the ESI source. For instance, capil-
lary temperature was varied between 150 and 320°C.
The perchlorate salts were added to 10 mL of acetoni-
trile solution containing one of the ligands 1–5 at a con-
centration of 10-4 M. Then four solutions were prepared
by mixing the solution of salt previously prepared and
solutions of ligand at a concentration of 10-4 M in order to
study the formation of complexes according to the number
of salt equivalents (0.5, 1, 2, 5 equiv.). The solution
obtained was then directly infused into the ESI source, the
flow rate was 3–7 lL min-1. Nitrogen was used as the
nebulizing and desolvating gas at flow rates of 100 and
300 L h-1, respectively. Every scan was recorded from
200 to 1500 m/z in no less than 5 s and at least 20 scans
were averaged to yield the final spectrum. Every spectrum
was reproduced three times. The spectra were recorded in
profile mode, for three different SCID (Source Collision
Induced Dissociation) values: 0, -20, -40 V but only
results at -20 V are presented here except for compound 5
(-40 V). These SCIDs have been chosen because they
permit to maximize the desolvation and to minimize the
fragmentation. To check the attribution of the observed
peaks, their isotopic patterns were compared with the
simulated pattern of their suggested molecular formula.
In order to represent the evolution of each species in
function with the metal concentration, the surface of the
peaks has been integrated and reported to the integral sum
of all peaks in each spectrum. The isotopic clusters with a
Trimethyl 2-(2-phenylacetamido)aminopropane-1,2,3-tri-
carboxylate (2) 1H NMR (CDCl3, 300 MHz) d 2.85 (d,
2J(H,H) = 15.5 Hz, 29 1H, CH2), 3.53 (s, 2H, CH2), 3.56
2
(s, 6H, 29 OCH3), 3.63 (d, J(H,H) = 15.5 Hz, 29 1H,
CH2), 3.79 (s, 3H, OCH3), 6.87 (br s, 1H, NH), 7.23–7.40
(m, 5H, CHaryl) ppm. 13C NMR (CDCl3, 75 MHz) d 39.7
(CH2), 44.5 (CH2), 52.0 (CH3), 53.6 (CH3), 59.2 (C), 127.5
(CHAr), 129.0 (CHAr), 129.4 (CHAr), 134.7 (CHAr), 170.0
(CO ester), 171.0 (CO ester), 171.8 (CO amide) ppm.
5-Diethoxyphosphoryl-5-(ethoxycarbonyl)methyl-pyrrolidin-
2-one ( 3) 1H NMR (CDCl3, 300 MHz) d 1.28 (t, 3J(H,H)
7.1 Hz, 3H, CH3), 1.35 (td, J(H,H) = 7.1 Hz, J(H,P)
=
=
3
4
2.7 Hz, 6H, 29 CH3), 2.22 (m, 1H, CH2), 2.36 (m, 1H, CH2),
2
2.57 (m, 3H, CH2), 2.90 (dd, J(H,H) = 15.1 Hz, J(H,P)
3
=
7.3 Hz, 1H, CH2), 4.18 (m, 6H, 39 OCH2), 6.21 (br s, 1H,
NH) ppm. 13C NMR (CDCl3, 75 MHz) d 14.2 (CH3), 16.7 (d,
3J(C,P) = 5.6 Hz, CH3), 28.5 (CH2), 29.7 (CH2), 40.7 (d,
3J(C,P) = 9.4 Hz, CH2), 57.8 (d, 1J(C,P) = 164.7 Hz, C), 61.3
2
2
(CH2), 63.3 (d, J(C,P) = 7.6 Hz, CH2), 63.8 (d, J(C,P)
=
7.4 Hz, CH2), 169.6 (d, J(C,P) = 11 Hz, CO ester), 177.3 (d,
3J(C,P) = 3.7 Hz, CO lactam) ppm. 31P NMR (CDCl3,
121 MHz) d 24.6 ppm.
5-(Diethoxyphosphoryl)-5-[2-(ethoxycarbonyl)ethyl]-pyrr-
olidin-2-one ( 4) 1H NMR (CDCl3, 300 MHz) d 1.25
3
3
(t, J(H,H) = 7.1 Hz, 3H, CH3), 1.34 (td, J(H,H) = 7.1 Hz,
4J(H,P) = 2.3 Hz, 6H, 29 CH3), 1.97 (m, 2H, CH2), 2.36
(m, 6H, 39 CH2), 4.16 (m, 6H, 39 OCH2), 6.24 (br s, 1H,
NH) ppm. 13C NMR (CDCl3, 125 MHz) d 14.3 (CH3), 16.7
3
3
(d, J(C,P) = 5.6 Hz, CH3), 27.3 (CH2), 28.5 (d, J(C,P)
=
3
7.5 Hz, CH2), 30.0 (CH2), 30.7 (d, J(C,P) = 9.2 Hz, CH2),
1
59.3 (d, J(C,P) = 162.2 Hz, C), 61.0 (CH2), 63.1 (d,
2J(C,P) =7.8 Hz, CH2), 63.7 (d, J(C,P) = 7.4 Hz, CH2),
2
123