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Paper spray ionization mass spectrometry for rapid
quantification of illegal beverage dyes+
Tianyang Guo,ab Zezhen Zhang,a Karen E. Yannell,a Yiyang Dongb
and R. Graham Cooks *a
A rapid method is described for quantification of six illegal colorants in beverages, i.e., soft beverages,
energy beverages, alcoholic beverages, teas, and fruit juices.Conventional mass spectrometry-based methods for detection
of dyes require complex pretreatment process, including
drying, extraction, and puri?cation.12,13 These treatments avoid
matrix interferences from various mono-, di- and polysaccharides
and modi?ed starch, and from compounds with
the same molecular weight as the analyte as well as minimizing
ionization suppression.14,15 These advantages come with the
need for time-consuming chromatographic separation
processes.12-14,16 Ambient ionization mass spectrometry17-19 is an
alternative way to rapidly ionize samples without or with only
minimal pretreatment.Four dyes in aqueous solution were detected by
DESI-MS at the ng mL1 level.27 Dyes with different structures
were identi?ed at pg mL1 level in wool samples by surface
acoustic wave nebulization (SAWN) mass spectrometry,
although degradation of certain analytes was observed.28
Paper spray (PS) ionization is a representative ESI-related
ambient ionization method which allows rapid and direct
analysis of an untreated sample from paper or other porous
(occasionally non-porous29) media which include a ?ne tip.In the case of foods, food dyes can compensate for color
loss, enhance natural colors, or add color to colorless food.1
Synthetic food dyes, synthesized from coal tar or petroleum byproducts,
2 are widely used, owing to their ease of production,
low cost, high stability, and good coloring properties.3 During
the 1900s, some 80 different food dyes were used in the United
States, but over the years, most of them have been taken off the
market because of safety concerns.2 Currently, only nine
synthetic colorings are allowed for food use by the U.S. Food
and Drug Administration (FDA), and these are subject to
certi?cation.4
Although maximum use levels for these dyes have been listed
by the FDA, their use still requires careful consideration.For
example, Fast Green, Orange B and Citrus Red 2 are legal in the
US, but they are not permitted in other countries, e.g., China5
and the European Union.6 Some studies showed that synthetic
food dyes may have a negative effect on the behavior of children,
such as hyperactivity.1,2,7 Furthermore, use of multiple synthetic
food colors is also a challenge for food safety, because of
potential synergistic effects.8
The addition to food of some industrial dyes, e.g., Sudan
dyes, Disperse dyes, Rhodamine B and Malachite Green, is
forbidden due to their genotoxic and carcinogenic activity.9
However, some producers still add these illegal non-food colors
to the product.10,11 Therefore, a sensitive method to screen both
legal and illegal dyes in food is required for food safety.The ESI-related method of desorption
electrospray ionization (DESI)20 and APCI-related direct analysis
in real time (DART),21 as well as many other ambient ionization
techniques, have been applied widely in food safety,19,22 environmental
analysis,23 forensics24 and clinical diagnosis.25
As for dyes, several ambient ionization methods have been
used for identi?cation or quanti?cation.However, this
technique is useful also in the food safety domain, because of
the complexity of food matrix and the requirement for trace
residue detection.33-37 Some azo dyes at mgmL1 level have been
examined in powdered chili pepper by PS-MS/MS.38
In this paper, we establish a PS-MS/MS method using
multiple reaction monitoring (MRM) to quantify illegal dyes in
liquid beverages.This
technique requires neither elevated temperature nor gas ?ow so
it is particularly appropriate for use with portable MS systems.30
In several application areas, it has been demonstrated to allow
aDepartment of Chemistry, Purdue University, West Lafayette, IN 47907, USA
bCollege of Life Science and Technology, Beijing University of Chemical Technology,
Beijing 100029, China
+ Electronic supplementary information (ESI) available.Experimental
2.1 Reagents and materials
Analytical standards of Sudan I, Sudan II, Malachite Green (as
chloride), Rhodamine B (as chloride), Crystal Violet (as chloride)
and Methylene Blue (as chloride) were purchased as
powders from Sigma-Aldrich (St.PS was performed
using a toothless copper clip, obtained from Muller
Electric (Akron, OH), and Whatman 1ET-Chr paper, obtained
from GE Healthcare Life Sciences (Chicago, IL).Isotopically labeled IS's have performed
very well in past ambient ionization mass spectrometry
analyses, owing to similar ionization efficiency and similar
transport along the paper substrate.31,32 However, the higher
cost of isotopically labeled standards makes their use for each
individual analyte impractical.LOD and
LOQ at ng mL1 level, accuracy and precision were calculated
using matrix-matched calibration curves.In ambient ionization mass
spectrometry analysis, IS use is highly desirable.39,40 Commonly,
two types of IS are used: structural analogues and stable-isotope
labeled forms of analyte.Samples where diluted in methanol and isotopically-labeled internal standards were utilized to
obtain linear responses over ranges of two or three orders of magnitude.Two representatives
Sudan dyes (Table 1) were selected to assess the
performance of rapid determination by paper spray (PS) as
described in this article.2.3 Instruments and devices
All MS experiments were performed using PS ionization
combined with triple quadruple mass spectrometry.Paper spray (PS) ionization combined with
tandem mass spectrometry (MS/MS) allowed detection of particular dyes in times of less than 1 min per
sample.See DOI:
10.1039/c7ay02241g
Cite this: DOI: 10.1039/c7ay02241g
Received 18th September 2017
Accepted 25th October 2017
DOI: 10.1039/c7ay02241g
rsc.li/methods
This journal is (C) The Royal Society of Chemistry 2017 Anal.The D5 isotopically labeled internal standards Sudan Id5
and Malachite Green-d5 picrate were also purchased from
Sigma-Aldrich, and stored at 20 C. Twenty-one colorless and twenty
colored beverage samples were purchased from three local
supermarkets in West Lafayette, Indiana (Walmart, Meijer,
Fresh City Market).View Article Online
View Journal
quanti?cation of analytes in complex matrices with the use of
internal standards.31,32 Most such applications focus on biological
samples, such as blood, urine and tissue.HPLC grade methanol,
acetic acid, and ammonium acetate were purchased from
Sigma-Aldrich (St.2.2 Analytes and internal standards
Illegal food dyes of two molecular types were investigated:
neutral molecules and salts with complex cations.Internal standards (IS) were used to improve accuracy and
precision, as well as to establish the robustness of the method
for the various complex matrices.Most neutral
molecule dyes produce the protonated molecule [M + H]+ in
positive ion mode, while not being ionized in the negative ion
mode.The complex cation salt dyes, ionize
with relatively high efficiency to give the intact cation [C]+ in the
positive mode.High voltage was supplied to the clip
and the solvent methanol was pipetted onto the paper triangle.The mass spectrometer used was a TSQ Quantum Access
MAX (Thermo Fisher Scienti?c, Waltham, MA).Four Sudan dyes in
chili powder were quanti?ed at the mgmL1 level by DART-MS,26
although DART has proved less successful with those dyes
which are salts.Compounds of this neutral molecule class, including
Sudan and related dyes, are usually oil-soluble.The examples of this class examined were Malachite
Green, Rhodamine B, Crystal Violet and Methylene Blue
(Table 1).Introduction
Dyes are added to speci?c products to provide attractive visual
effects.As a ?nal test, real
samples were screened to ensure the usefulness of this method.The IS Sudan I-d5 was
chosen for the neutral molecule class and Malachite Green-d5
picrate for the complex salt class (Table 1).The LODs were below 1.5 ng
mL1 and LOQs were below 5 ng mL1.Matrix effects were evaluated for different samples but had little effect given the
use of the dilution method.Methods
Analytical
Methods
PAPER
Published on 26 October 2017.Minimal sample preparation was used and
internal standards were investigated and optimized.beverages, energy
beverages, alcoholic beverages, teas, and fruit juices.Therefore, two IS's were selected
to allow detection of multiple analytes.The paper
substrate was cut into a small triangle (8 10, mm), and clamped
with the copper clip.Quality control experiments gave accuracies that ranged
between 80% and 120%.Downloaded by Beijing University of Chemical Technology on 07/11/2017 15:34:28. Louis, MO), and stored at
20 C.Louis, MO).They included so?1.2.
Paper spray ionization mass spectrometry for rapid
quantification of illegal beverage dyes†
Tianyang Guo,ab Zezhen Zhang,a Karen E. Yannell,a Yiyang Dongb
and R. Graham Cooks *a
A rapid method is described for quantification of six illegal colorants in beverages, i.e., soft beverages,
energy beverages, alcoholic beverages, teas, and fruit juices. Paper spray (PS) ionization combined with
tandem mass spectrometry (MS/MS) allowed detection of particular dyes in times of less than 1 min per
sample. Samples where diluted in methanol and isotopically-labeled internal standards were utilized to
obtain linear responses over ranges of two or three orders of magnitude. The LODs were below 1.5 ng
mL1 and LOQs were below 5 ng mL1. Quality control experiments gave accuracies that ranged
between 80% and 120%. Matrix effects were evaluated for different samples but had little effect given the
use of the dilution method.
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