30 February / March 2021
and the 100 ppb working solution was used to prepare the 50 ppb, 10 ppb, 5 ppb, and 1 ppb standards.
Results and discussion
Adding ammonia to the make-up flow provided the strongest m/z response
Figure 3. Gradient method provides a higher resolution separation compared to isocratic method. (A) IC-sCD chromatogram obtained for 2-butynoic acid analysed by gradient method (black) (B) extracted ion chromatogram of m/z 171 (pink) (C) extracted ion chromatogram of m/z 127 (blue) analysed using MeCN: H2
O + 25 mM ammonia as additives in the make-up flow [2].
of 51 mA was used unless otherwise stated. Dionex ADRS 600 suppressor applies a direct fixed voltage to give the most suitable current to match the concentration of the eluent to suppress the hydroxide to water. A voltage of 4 V was used. The flow rate was maintained at 0.38 mL/min and an injection volume of 25 µL was used for all injections. The specifications of the isocratic and gradient methods are as follows:
• Isocratic method: 30 mM KOH, 30 °C, lower current 29 mA
• Gradient method: Initial 1 mM KOH for 8.5 min, then linear gradient to 15 mM over 10 min, then to 30 mM over a further 10 min and then to 54 mM in 1.5 min.
MS conditions
All MS experiments were performed using the following conditions: vaporiser temperature of 450°C, ion transfer tube temperature of 200°C, ionisation voltage of -2500 V, sheath gas pressure of 50 psi and auxiliary gas pressure of 5 psi. The system was operated in negative ion ESI mode for all analyses. The MS system, when operated in full scan mode, recorded data for the mass range of m/z 40-250. Selected Ion Monitoring (SIM) mode was used for the analysis of deprotonated molecules. The MS ion source voltage (applied to aid desolvation) was set to 5 V for both the full scan and SIM analyses. Make-up flow of 1:1 acetonitrile: water (+ buffer additive, if used) was added at a rate of 0.1 mL/min.
Standards and sample preparation
Individual organic acid stock standard solutions were prepared at 10 ppm (v/v) or 10 mg/L, except for butynoic, crotonic,
and pentynoic acid solutions which were prepared at 50 ppm (v/v) or 50 mg/L. Mixed working stock solutions of 1 ppm and 100 ppb were prepared by diluting the stock solutions 10- and 100-fold respectively (50- and 500-fold for 50 ppm stock). These working stock solutions were then diluted to prepare a range of calibration standards between 1 ppb and 500 ppb. The 1 ppm working solution was used to prepare the 500 ppb, 200 ppb, and 100 ppb standards,
Table 1. R2 ppb using both IC-sCD and MS [2]. CD Component Acetic acid Linearity R2 0.9925
Propinoic acid 0.9926 Formic acid
0.9992
Butanoic acid 0.9958 Crotonic acid 0.9998 Pentanoic acid 0.9954 Propiolic acid n/a Butynoic acid n/a Pentynoic acid 0.9972
LOQ / ppb (vol/ vol)
1 1 1 1 5 1
n/a n/a 1
Linearity R2
0.9942 0.9863 0.9597 0.9903 0.9969 0.9927 0.9527 0.9880 0.9989
1 1 1 1 1 1 1 1 5
Table 2. Linearity and LOQ of organic acids within a 25 mg/L 2-butynoic acid sample matrix using IC-sCD and MS [2].
Component Acetic acid CD Linearity R2 0.9998
Propinoic acid 0.9998 Formic acid
0.9995
Butanoic acid 0.9970 Crotonic acid
Pentanoic acid 0.9997 Propiolic acid
n/a Pentynoic acid 0.9915 0.9997
0.005 0.005 0.005 0.005 0.02
0.005 n/a
0.005
LOQ / %w/w Linearity R2 0.9814 0.9966 0.9944 0.9991 0.9993 0.9982 0.9944 0.9853
MS LOQ / %w/w
0.005 0.005 0.005 0.005 0.005 0.005 0.005 0.05
MS LOQ / ppb
The main impurity of 2-butynoic acid, analysed using the isocratic method, produced an ion with m/z 127 with a partially co-eluting impurity at m/z 171 as shown by the extracted ion chromatograms (Figure 2A). Using different additives in the make-up flow resulted in different MS signal responses as shown in Figure 2. Based on its chemical composition, each additive (MeCN: H2
O, MeCN: H2 mM ammonium acetate and MeCN: H2
O + 12.5 O
+ 25 mM ammonia) helps to volatilise the analytes in solution and, in turn, contributes to the efficiency of the ionisation process. Of these additives, using 25mM ammonia in the make-up solvent provided a greater response for both m/z 127 and m/z 171 ions (Figure 2). Ammonia was therefore chosen for all subsequent experiments. As expected, the gradient method provided a
values demonstrating the linearity of a range of organic acid standards between 1 ppb and 500
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