Volume 28 Issue 2 Article 10

A multi-analyte LC-MS/MS method for screening and quantification of

nitrosamines in sartans

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Recommended Citation
Chang, Shu-Han; Chang, Ching-Chia; Wang, Li-Jing; Chen, Wei-Ching; Fan, Shu-Yu; Zang, Chi-Zong; Hsu, Ya-Hui; Lin,
Mei-Chih; Tseng, Su-Hsiang; and Wang, Der-Yuan (2020) "A multi-analyte LC-MS/MS method for screening and
quantification of nitrosamines in sartans," Journal of Food and Drug Analysis: Vol. 28 : Iss. 2 , Article 10.
Available at: https://doi.org/10.38212/2224-6614.1063

This Original Article is brought to you for free and open access by Journal of Food and Drug Analysis. It has been accepted for
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ORIGINAL ARTICLE

A multi-analyte LC-MS/MS method for screening and

quantification of nitrosamines in sartans

Shu-Han Chang, Ching-Chia Chang, Li-Jing Wang, Wei-Ching Chen, Shu-Yu Fan,
Chi-Zong Zang*, Ya-Hui Hsu, Mei-Chih Lin, Su-Hsiang Tseng, Der-Yuan Wang

Food and Drug Administration, Ministry of Health and Welfare, Executive Yuan, 161-2 Kunyang St., Nangang Dist., Taipei 11561,
Taiwan

Abstract

An incident of sartan medicine contamination was notified by Europe in June 2018. The contaminant was identified
as a probable carcinogenic nitrosamine and the recalls of sartan medicines were soon made. Since then, more nitro-
samine contaminants in sartan medicines were reported. To broaden the applicability and variety in nitrosamine
determination, a multi-analyte method is required. In this study, a feasible and sensitive multi-analyte LC-MS/MS
method for determination of 12 nitrosamines in sartans was established, where the active pharmaceutical ingredients
and final products merchandised in Taiwan were also examined. Chromatographic separation was achieved on an
Xselect® HSS T3 column (15 cm £ 3 mm i.d., 3.5 mm) with gradient elution using mobile phase A consisting of 0.1%
formic acid in water and mobile phase B consisting of 0.1% formic acid in acetonitrile/methanol (2:8). Validation of the
proposed method was also carried out. The limit of detection and limit of quantification for 12 nitrosamines were
20 ng/g and 50 ng/g, respectively. The intra-day and inter-day recoveries of nitrosamines were among 80e120% with
precision of 20% for most nitrosamines within sartans matrices. The method was successfully established and applied
to authentic samples which a total of 98 positive samples containing 5 distinct nitrosamines, including N-nitro-
sodiethylamine, N-nitrosodimethylamine, N-nitroso-N-methyl-4-aminobutyric acid, N-nitrosomorpholine and N-
nitrosopiperidine, were detected from 557 authentic samples.

Keywords: Carcinogens, Liquid chromatography, Mass spectrometry, Nitrosamines, Sartans

1. Introduction These agents have been manufactured for years

where impurities and related substances are found

A ngiotensin II is a biologically active

component that plays an important role in
as analogs or degraded fragments of sartans [5e8].
Recently, unexpected carcinogenic impurities were
the regulation of blood pressure, secretion of detected in sartan medicines. Those compounds
aldosterone, and homeostasis of fluid in human were further identified as nitrosamine analogs N-
body. Besides, it is an etiological factor for car- nitrosodiethylamine (NDEA) and N-nitro-
diovascular diseases [1]. Angiotensin II receptor sodimethylamine (NDMA). Those nitrosamine
blockers (ARBs), also known as angiotensin II re- impurities are produced as by-products resulted
ceptor type 1 antagonists or “sartans”, have been from the alteration of the manufacturing processes
widely applied for treating cardiovascular diseases [9,10]. Generally, nitrosamines are found in natural
such as hypertension, heart failure, myocardial environment and industrial manufacturing, such
infarction, and diabetic nephropathy [1e3]. To as pesticides, rubbers, dyes, and pharmaceuticals
date, several sartans, such as losartan and valsar- [11,12]. In industry, nitrosamines are inferred to
tan, have been launched by many countries [4]. forming in manufacturing processes that employ

Received 9 September 2019; accepted 19 December 2019.

Available online 27 June 2020

* Corresponding author at: Fax: þ886 2 2653 1764.

E-mail address: [email protected] (C.-Z. Zang).

https://doi.org/10.38212/2224-6614.1063
2224-6614/Copyright © 2020 Food and Drug Administration, Taiwan. This is an open access article under the CC BY NC ND 4.0 license
(http://creativecommons.org/licenses/by-nc-nd/4.0/).
 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301 293

ORIGINAL ARTICLE
amines, nitrates, and nitrites under a wide range of method for determination of 8 nitrosamines in water
pH levels, via different mechanisms such as which demonstrated a good performance of deter-
nitrosation by nitrite [13,14]. Oxidation of unsym- mination at a low concentration level [22]. Only few
metrical dimethylhydrazine (UDMH) also affords works focused on screening nitrosamines in phar-
maceutical matrices. Parr and Joseph have reviewed
nitrosamines [15]. For sartans, the application of
the determination method of nitrosamines
alternative solvents in sartan production led to the including GC-based, LC-based and non-chromato-
production of nitrosamines, for example, NDMA graphic methods [24]. Among those methods, LC-
was generated from dimethylformamide (DMF) MS/MS has various advantages in analysis
and NDEA was generated from triethylamine including less amount of sample usage, thermal
(TEA) [16]. Nitrosamines are considered as po- applicability of analytes and lower detection limit
tential human carcinogens and the cancer risk has for determination of nitrosamines in pharmaceuti-
been estimated which the excess lifetime is 105 to cals [25,26]. Therefore, this study aimed at estab-
lishing a feasible and sensitive method for screening
106 in the concentration range 0.7e100 ng/L
of 12 nitrosamines in sartans using LC-MS/MS,
(drinking water) [17e19]. The risk of nitrosamines whereas method validation was carried out. The
for human health has been highly dressed that proposed method was further applied for the anal-
many governments impose strict regulations on ysis of sartan APIs and final products merchandised
nitrosamines. For example, NDEA and NDMA are in Taiwan.
categorized as Group 2A substances (probably
carcinogenic to humans) by International Agency 2. Materials and methods
for Research on Cancer (IARC) of WHO, category 2.1. Chemicals and reagents
1B by the European Union (Presumed to have
carcinogenic potential for humans), and category N-nitrosodiethylamine (NDEA), N-nitrosodimethyl-
B2 (probable human carcinogen) by U.S.; besides, amine (NDMA), N-nitrosodipropylamine (NDPA), N-
N-nitrosodipropylamine (NDPA), N-nitro- nitrosodimethylamine-d6 (NDMA-d6) and N-nitro-
sodipropylamine-d14 (NDPA-d14) were purchased
somethylethylamine (NMEA), N-nitrosomorpho-
from AccuStandard (CT, USA). N-nitro-
line (NMOR), N-nitrosopiperidine (NPIP) and N- soethylisopropylamine (NEIPA) was purchased from
nitrosopyrrolidine (NPYR) are categorized as B2 BOC Sciences (NY, USA). N-nitrosodiisopropylamine
level by IARC [19]. As a consequence, U.S. EPA (NDiPA) was purchased from Chem Service (PA,
(Environmental Protection Agency) incorporates 6 USA). N-nitrosodiethanolamine (NDELA), N-nitro-
nitrosamines in the regulation, including N-nitro- somorpholine (NMOR) and N-nitrosopyrrolidine
sodibutylamine (NDBA), NDEA, NDMA, NDPA, (NPYR) were purchased from SigmaeAldrich (MO,
NMEA and NPYR [20]. Due to the probable hazard USA). N-nitrosopiperidine (NPIP) was purchased
from Supelco (PA, USA). N-nitrosodiisopropanol-
to human body, limits of nitrosamines in sartans
amine (NDiPLA), N-nitroso-N-methyl-4-aminobutyric
were regulated that U.S. FDA drew up an interim acid (NMBA), N-nitrosomethylethylamine (NMEA),
limit which the acceptable intakes for NDEA, N-nitrosodiethylamine-d4 (NDEA-d4), N-nitro-
NDMA and NMBA in 8 sartans are 26.5, 96 and sodiethanolamine-d8 (NDELA-d8), and N-nitroso-N-
96 ng/day, respectively [21]. methyl-4-aminobutyric acid-d3 (NMBA-d3) were pur-
The recent incident of nitrosamine contamination chased from TRC (ON, Canada). The chemical struc-
in sartan medicines has raised concerns regarding tures of 12 nitrosamines are shown in Fig. 1. 557
the medication safety in many countries. Inevitably, samples from 5 sartans, including candesartan cilexetil
Taiwan was also affected that active pharmaceutical (4 APIs and 6 final products), irbesartan (35 APIs and
ingredients (APIs) and final products of 5 sartans 28 final products), losartan (68 APIs and 198 final
(candesartan cilexetil, irbesartan, losartan, olme- products), olmesartan cilexetil (8 APIs and 18 final
sartan cilexetil and valsartan), and final products of products), and valsartan (65 APIs and 127 final prod-
2 sartans (azilsartan and telmisartan) were im- ucts), were collected and submitted by local public
ported. It's required to develop an analytical method health bureaus of Taiwan. The chemical structures of
for screening nitrosamines in pharmaceuticals. In 5 sartans are shown in Fig. 2. Methanol and formic
many studies, it focused on the detection of nitro- acid of LC-MS grade were purchased from
samines in water matrices [12,18,22,23]. For SigmaeAldrich (MO, USA). Acetonitrile of LC-MS
example, Ripoll
es et al. developed a sensitive grade was purchased from J.T Baker (NJ, USA). Water
 294 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301
ORIGINAL ARTICLE

of LC-MS grade was purchased from Scharlau (Bar- standard solution was prepared by diluting the in-
celona, Spain). ternal standard stock solution with methanol to a
concentration of 400 ng/mL prior to use. A series of
2.2. Instrumentation and chromatographic working standard solutions for calibration curve
conditions were prepared at concentrations of 2.5, 5.0, 10.0,
20.0, 25.0 and 50.0 ng/mL with 20 ng/mL of the in-
The experiments were performed on a Waters ternal standard. All the stock and working solutions
Acquity UPLC® (Waters Assoc., Milford, MA, USA) were stored at 30  C and acclimated to room
coupled to a 6500 triple quadrupole linear ion temperature prior to use.
trap mass spectrometer (MS) equipped with an at-
mospheric-pressure chemical ionization (APCI) 2.5. Sample preparation
source (AB Sciex, USA), and operated in multiple re-
action monitoring (MRM) mode. The MRM parame- Transfer 250 mg of sartan API (or ground tablet
ters including MRM transitions, quantifier, qualifier, powder of final product) to a 15 mL centrifuge tube.
collision energy and declustering potential are pro- Add 250 mL of the internal standard solution
vided in Table 1. The chromatographic separation was (400 ng/mL) as well as 250 mL of methanol (for
carried out on a trifunctional alkyl C18 bonded phase validation, 250 ml of nitrosamine standard solution at
column Xselect® HSS T3 column (15 cm  3 mm i.d., concentration of interest was added) to the tube, and
3.5 mm) (Waters Assoc., Milford, MA, USA) at 40  C sonicated for 5 min to make an extraction solution.
with a constant flow rate of 0.6 mL/min using gradient Add 4.5 mL of DI water to the extraction solution
elution of mobile phase A consisting of 0.1% formic above and sonicate for 5 min. Centrifuge the sample
acid in water and mobile phase B consisting of 0.1% solution at 3000 g for 5 min, filter the solution
formic acid in acetonitrile/methanol (2:8). Each sample through a 0.22 mm PVDF filter, and then collect the
was analyzed with an injection volume of 10 mL. The filtrate, which was used as the sample solution.
total chromatographic run time was 15 min. The
elution was performed with varied gradient as follows: 2.6. Method validation
0e1.0 min 5% B, 1.0e5.0 min 5e50% B, 5.0e6.5 min
50% B, 6.5e7.5 min 50e65% B, 7.5e8.5 min 65% B, 2.6.1. Linearity and sensitivity
8.5e9.5 min 65e100% B, 9.5e12.0 min 100% B, and The linearity was assessed by analyzing standard
12.1e15.0 min 5% B for equilibrium. After the injec- solutions of 12 nitrosamines (n ¼ 3) at 6 concentra-
tion of each sample, the needle was rinsed alternately tions ranging from 2.5 to 50.0 ng/mL by plotting the
with methanol and water. A two-position diverter peak area ratio of standard/internal standard versus
valve was used and settings were as follows: detector the concentration of standard using the least-
over 0e8.5 min, waste over 8.5e15.0 min. The MS ion squares method estimated by logistic regression.
source was set as APCI in positive mode, under the The linearity is acceptable when the correlation co-
following conditions: temperature, 400  C; curtain gas efficient r was higher than 0.995. The sensitivity of
pressure, 25 psi; collision gas pressure of medium the method was evaluated using the limit of detec-
level, ion source gas, 50 psi; and nebulizer current, tion (LOD) and limit of quantification (LOQ) ac-
5 mA. The selectivity for precursor ion (Q1) and cording to ICH validation of analytical procedures
product ions (Q3) was set to 0.7 m/z (FWHM). [27]. The LOD is the lowest concentration of analyte
that can be detected with the estimated signal-to-
2.3. Robustness noise (S/N) ratio of 3. The LOQ is the lowest con-
centration of analyte that can be quantified with the
To examine the robustness of the proposed method, suitable precision and accuracy using an estimated
the effect of deliberation variations was investigated S/N ratio of 10.
by following parameters such as mobile phase addi-
tive (0.1e0.2% formic acid), organic components of 2.6.2. Accuracy and precision
mobile phase (20e40% acetonitrile in methanol) and Before assessing the accuracy (expressed as re-
flow rate of mobile phase (0.3e0.6 mL/min). covery, %), all sartan matrices were examined to be
absent of nitrosamines and so that the interference
2.4. Preparation of standard solutions to standards could be avoided. The accuracy was
evaluated by introducing quality control (QC) for
Stock solutions of 12 nitrosamines and 5 internal nitrosamines in sartan materices [27]. The proced-
standards (IS) were prepared in methanol at ure was in reference of ICH and followed the criteria
1000 mg/mL and 50 mg/mL, respectively. The internal for QC in chemical analysis set up by TFDA (for
 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301 295

ORIGINAL ARTICLE
Fig. 1. Chemical structures of 12 nitrosamines analyzed in this study.

Fig. 2. Chemical structures of 5 sartans analyzed in this study.

 296 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301
ORIGINAL ARTICLE

Table 1. MRM parameters of 12 nitrosamines and 5 internal standards. when the target is not vaporized by APCI [27]. In the
Analyte RT (min) MRM transitions DP (V) CE (eV) literature, the advantages of APCI such better per-
(m/z) formance in selecting ions and less susceptible to
NDELA 1.88 135 > 74a 29 17
matrices for determination of nitrosamines have
135 > 104b 9 been reported [22,28]. Preliminary tests were carried
NDMA 2.7 75 > 58a 28 15 out to assess the adaptability of ionization sources in
75 > 43b 21 detecting nitrosamines. Accordingly, APCI showed
NMOR 3.5 117 > 87a 38 17 higher signals for all nitrosamines and thus selected
117 > 86b 20
NDiPLA 3.75 163 > 88a 44 17
as the ionization source in this study. The MRM
163 > 70b 28 chromatograms of 12 nitrosamines with APCI are
NMBA 3.84 147 > 117a 10 8 presented in Fig. 3. The MRM parameters with
147 > 87b 16 APCI including are summarized in Table 1. Q1 and
NMEA 3.95 89 > 61a 7 16 Q3 for all analytes are within 0.7 m/z FWHM. De-
89 > 29b 27
NPYR 4.0 101 > 55a 58 22
fects for some chromatograms were observed dur-
101 > 41b 37 ing analysis. A doublet peak of NDiPLA
NDEA 5.3 103 > 75a 25 13 chromatogram with same MRM transitions
103 > 47b 29 appeared which may indicate the separation of
NPIP 5.62 115 > 69a 42 21 racemic mixture or zwitterions of NDiPLA analyzing
115 > 41b 33
NEIPA 6.27 117 > 75a 16 14
on Xselect® HSS T3 column. The peak shoulder
117 > 47b 22 observed in the NMEA and NMBA chromatograms
NDiPA 7.46 131 > 89a 50 13 was attributed to pH adaptation of mobile phase. All
131 > 43b 22 the doublet peak and peak shoulder mentioned
NDPA 8.15 131 > 89a 59 14 above were adopted to reduce the deviation in
131 > 43b 21
NDELA-d8 1.87 143 > 111a 23 8
quantification and validation.
NDMA-d6 2.65 81 > 46a 106 25
NMBA-d3 3.82 150 > 120a 36 9 3.2. Robustness
NDEA-d4 5.25 107 > 77a 48 16
NDPA-d14 7.99 145 > 50a 130 46 To evaluate the effect of mobile phase on sepa-
MRM, multiple reaction monitoring; RT: retention time. ration, quantification and peak width of analytes,
a
Quantifier.
b parameters such as additive (0.1e0.2% formic
Qualifier; CE: collision energy; DP: declustering potential.
acid), organic component (20e40% acetonitrile in
1.0e10.0 ng/mL, recovery 60e125%, RSD 30%; for methanol) and flow rate (0.3e0.6 mL/min) were
10.0e100.0 ng/mL, recovery 70e120%, RSD 20%). investigated. The results indicated that the condi-
The intra-day and inter-day accuracy (%) and pre- tions of mobile phase had only slight influence on
cision (RSD in %) of the assay were assessed at 3 peak width (less than 0.1 min for all analytes)
concentration levels from low to high within the which supports the robustness of the proposed
calibration curve (5.0, 25.0, and 50.0 ng/mL) in 3 method.
replicates. The accuracy was assessed by comparing
the estimated concentration of analyte in matrix (A) 3.3. Method validation
with that of standard solution (B), whereas the ratio
was calculated as following formula: A/B  100%. 3.3.1. Linearity and sensitivity
The assessment was performed as follows: for intra- The linearity of 12 nitrosamines was obtained up
day, 3 samples for each concentration, and 1 repli- to 50.0 ng/mL and the data are presented in Table 2.
cate for each sample, n ¼ 3; for inter-day, 3 samples The values of correlation coefficient r were higher
for each concentration, 1 replicate for each sample, than 0.995 for all analytes which indicated that the
for 3 consecutive days, n ¼ 9. most IS applied in qualification were highly
recommendable for the targets analyzed in this
3. Results and discussion study. The LOD and LOQ determined for all ana-
lytes in sartan APIs and final products were 20.0 ng/
3.1. Optimization of chromatographic conditions g and 50.0 ng/g, respectively. In the literatures,
methods have been developed for determination of
To optimize the chromatographic conditions, the nitrosamines in sartans. Schmidtsdorff and Schmidt
APCI was selected as the ionization source for developed a method for determination of 9 nitro-
determination of nitrosamines in this study; never- samines (NDBA, NDEA, NDMA, NDPA, NDPhA,
theless, ESI can also be applied as an alternative NMEA, NMOR, NPIP and NPYR) using
 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301 297

ORIGINAL ARTICLE
supercritical fluid chromatography which the LODs 3.3.2. Accuracy and precision
ranged from 50 to 140 ng/g in valsartan and The intra-day accuracy of nitrosamines for sartan
20e460 ng/g in losartan [29]; el-Atma and Gutsche APIs was 81.4e117.0%, while the precision (RSD)
reported a LC-MS/MS method for determination of located in the range of 0.7e9.8%; for sartan final
2 nitrosamines which the LODs for NDEA and products, the value was 82.6e118.4%, while the
NDMA were 20 and 70 ng/g, respectively, while the precision located in the range of 0.3e13.1%. The
LOQs for NDEA and NDMA were 40 and 100 ng/g, inter-day accuracy of nitrosamines for sartan APIs
respectively [30]. Compared to the published was 81.7e117.5%, while the precision located in the
methods, this study demonstrated a variety of tar- range of 1.2e10.5%; for sartan final products, the
gets and sensitivity of detection for determination of value was 82.9e118.0%, while the precision located
nitrosamines in sartans. in the range of 0.8e12.8%. The assessment of

Fig. 3. MRM chromatograms applying APCI ionization source for 12 nitrosamines and 5 internal standards.
 298 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301
ORIGINAL ARTICLE

Fig. 3. (continued).
 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301 299

ORIGINAL ARTICLE
Fig. 3. (continued).

accuracy and precision met the criteria (see section Table 2. Linearity data of 12 nitrosamines.
2.6.2) for most nitrosamines within 5 sartan Analyte Linear Slope Intercept Correlation
matrices, except for NDiPA and NEIPA within range (ng/mL) coefficient (r)
irbesartan matrix. The intra-day accuracy of NDiPA NDELA 2.5e50.0 0.23769 0.00298 0.9989
for irbesartan API was 139.1e160.3%, where it was NDMA 2.5e50.0 0.40608 0.00448 0.9988
97.3e124.1% for irbesartan final products; the inter- NMOR 2.5e50.0 0.34094 0.00193 0.9993
NDiPLA 2.5e50.0 0.75543 0.00435 0.9995
day accuracy of NDiPA for irbesartan API was NMBA 2.5e50.0 0.82797 0.00702 0.9977
140.3e176.9%, where it was 113.1e126.0% for irbe- NMEA 2.5e50.0 2.18029 0.01566 0.9994
sartan final products. The intra-day accuracy of NPYR 2.5e50.0 2.23857 0.02065 0.9992
NEIPA for irbesartan API was 64.5e70.3%, where it NDEA 2.5e50.0 0.58442 2.22599e-4 0.9994
was 47.5e49.3% for irbesartan final products; the NPIP 2.5e50.0 2.06747 0.04224 0.9983
NEIPA 2.5e50.0 2.11105 0.00859 0.9992
inter-day accuracy of NEIPA for irbesartan API was NDiPA 2.5e50.0 27.90931 0.51163 0.9994
65.9e71.2%, where it was 46.3e48.9% for irbesartan NDPA 2.5e50.0 31.94901 0.85289 0.9996
final products. The result of accuracy and precision
for NDiPA (NEIPA) within irbesartan matrix indi-
cated the chromatographic intensity was enhanced 3.4. Analysis of authentic samples
(suppressed) owing to the matrix interference of
irbesartan, i.e. the intensity of corresponding inter- 557 samples from 5 different sartans acquired
nal standards NDPA-d14 (NDEA-d4) were enhanced from local public health bureaus were analyzed and
(suppressed) and resulted in the mis-estimation in the result was shown in Fig. 4. A total of 98 positive
quantification. To improve the disadvantage, the samples were detected from 557 authentic samples
corresponding internal standards for NDiPA and which 3 sartan medicines, including irbesartan,
NEIPA could be replaced by alternative isotropic losartan, and valsartan were detected positive of
nitrosamines to receive a better performance in the nitrosamines. Among those samples, valsartan had
assessment of accuracy and precision. the highest detection rate of nitrosamine-positive
 300 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301
ORIGINAL ARTICLE

Fig. 4. Amount and detection rate of nitrosamine-positive samples from authentic samples.

Table 3. Amount and content of corresponding nitrosamine detected in positive samples.

Item NDMA NDEA NMBA NMOR NPIP
Amounta Content Amount Content Amount Content Amount Content Amount Content
(pcs) (mg/g) (pcs) (mg/g) (pcs) (mg/g) (pcs) (mg/g) (pcs) (mg/g)
Valsartan 53 0.10e137.60 11 0.07e6.90 1 0.14 2 0.16 2 0.12
Losartan 0 ND 4 0.08e0.23 32 0.22e27.40 0 ND 0 ND
Irbesartan 0 ND 5 0.11e0.12 0 ND 0 ND 0 ND
a
Amount of contaminated samples sorted according to corresponding nitrosamine.

samples (29.17%), i.e. 56 out of 192 samples were medicines, 8 samples possessed 2 contaminants
detected positive, followed by losartan (36/266, NDMA and NDEA; 2 samples possessed 3 con-
13.53%) and irbesartan (6/63, 9.52%). The result of taminants NDEA, NMOR and NPIP; 1 sample
detection was further interpreted to sort the types of possessed 3 contaminants NDEA, NDMA and
nitrosamines, and demonstrated in Table 3. 5 ni- NMBA. Echo to the description in introduction, as
trosamines, including NDEA, NDMA, NMBA, long the conditions were sufficient, various nitro-
NMOR and NPIP, were detected in the 3 sartan samines would be produced which followed that the
medicines mentioned above. For valsartan, most of modification of sartan APIs manufacturing process
the positive samples (53/56) were detected having intensively affected the quality of pharmaceuticals
NDMA, which the content attained to an enormous and led to the cross-border incident of medication
level of 137.6 mg/g and indicated NDMA as the main safety.
contaminant produced as by-product in the modi-
fication of manufacturing process of valsartan. As to 4. Conclusion
losartan, the majority of the positive samples (32/36)
were detected having NMBA at the content level of In this study, a multi-analyte LC-MS/MS method
27.40 mg/g which posed that NMBA was the main was successfully developed for screening and
contaminant produced as by-product in the modi- determination of 12 nitrosamines in sartan medi-
fication of manufacturing process of losartan. For a cines. The proposed method was validated and
better understanding, all positive samples were provided satisfactory result of validation for target
employed to examine if it is possible that more than nitrosamines in most APIs and final products which
2 contaminants were detected in sartan medicines. demonstrated a good performance and specificity
Not surprisingly, multiple nitrosamine contents for the method in screening and qualification of
were found. In the positive samples of valsartan nitrosamines.
 JOURNAL OF FOOD AND DRUG ANALYSIS 2020;28:292e301 301

ORIGINAL ARTICLE
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Health and Welfare, Taiwan. neering systems and atmospheric chemistry engineering
systems division and department of earth, atmospheric and
planetary Sciences, Massachusetts Institute of Technology.
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