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Determination of Sucralose by Controlled UV Photodegradation Followed by

UV Spectrophotometry

Article in Journal of AOAC International · May 2013

DOI: 10.5740/jaoacint.12-143

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 Idris et al.: Journal of AOAC International Vol. 96, No. 3, 2013 603

FOOD COMPOSITION AND ADDITIVES

Determination of Sucralose by Controlled UV

Photodegradation Followed by UV Spectrophotometry
Mohd Idris
Govt. of India, Ministry of Home Affairs, Central Forensic Science Laboratory, Ramanthapur, Hyderabad, India
V. Jayathirtha Rao
Indian Institute of Chemical Technology, Organic Chemistry-II, Tarnaka, Hyderabad, India
Deepak Middha
Govt. of India, Ministry of Home Affairs, Central Forensic Science Laboratory, Ramanthapur, Hyderabad, India
Sudhir Kumar Shukla
Govt. of India, Ministry of Home Affairs, Central Forensic Science Laboratory, Chandigarh, India
Tulsidas Ramachandra Rao Baggi
Osmania University, University College of Science, Department of Chemistry, Forensic Science Unit, Hyderabad, India

A simple and time-efficient UV spectrophotometric the lower range of the spectrum approaching the vacuum UV
method was developed for the determination of region, which is vulnerable to error. It has also been determined
sucralose in artificial sweeteners. The procedure is simultaneously with other non-nutritive sweeteners in foods by
based on the UV spectrophotometric determination HPLC/electrospray ionization-MS (7), HPLC-evaporative light
of the photodegraded product at 270 nm in scattering detector (8), capillary electrophoresis (9), isotope
alkaline medium (pH 12). LOD for this method dilution LC/MS/MS (10), and TLC using amino plates (11)
was 0.02 g/L, and the calibration curve was found with a derivatization reagent (12, 13). These methods are time-
to obey Beer’s law in the concentration range consuming and tedious and need special cleanup procedures
of 0.1 to 1.2 g/L, with a correlation coefficient (5–10) for sample preparation. The method based upon TLC
2
(R ) value of 0.9977. The proposed method using amino plates (11) was simple, and sucralose determination
was studied for accuracy and precision. after postchromatographic derivatization was sensitive, but these
methods require a densitometer for quantitative analysis.
This paper describes the UV irradiation of sucralose in

S
pH 12 solution at 254 nm, followed by the determination of
ucralose (1,6-dichloro-1,6-dideoxy-β-D-fructofuranosyl-
its photodegraded product by UV spectrophotometry. The
4-chloro-4-deoxy-α-D-galactopyranoside; Figure 1) is
developed method was applied for determination of sucralose
made by substituting three hydroxyl groups of sucrose
in commercial sweeteners.
with chlorine atoms. Replacing three of the hydrogen atoms
in the sucrose molecule with chlorine atoms makes sucralose
Experimental
almost nonabsorbable by the intestine, thereby not increasing
the intake of calories in the food ingested. Its importance is
Chemicals, Solutions, Standards, and Samples
based mainly on properties such as its sweetness, immunity
to metabolic degradation, nonhygroscopic nature, and relative All solutions were prepared with double-distilled water
heat resistance. It has a taste profile very similar to that of from a Mono Quartz distillation unit (Borosil, Mumbai, India).
sugar without any aftertaste and is approximately 600 times Buffer solutions were prepared as follows: (pH 14, 1 M NaOH
as sweet as sucrose (1). It has been approved by the European solution; pH 12, 0.1 M NaOH solution; pH 10, 25 mL 0.05 M
Commission’s Scientific Committee on Food, the U.S. Food borax + 43 mL 0.1 M NaOH, in 100 mL; pH 7, 82 mL 1 M
and Drug Administration, and the regulatory agencies of more Na2HPO4 made up to 100 mL by adding 0.1 M citric acid; and
than 60 countries for use in numerous food and pharmaceutical pH 2: 0.01 M HCl solution. All reagents and chemicals used
preparations. The acceptable daily intake in general has been were purchased from S.D. Fine Chem Ltd (Mumbai, India).
agreed to be 15 mg/kg/day. Standard sucralose (98.2%) was purchased from Sigma-Aldrich
Analysis of sucralose by spectrophotometric methods is (Bangaluru, India). Tabletop sweeteners containing sucralose
difficult because it lacks a suitable chromophore. Several methods were purchased from a local market.
have been reported for the analysis of sucralose, including RP-
HPLC coupled with a refractive index detector (2–4) or UV Apparatus
detection at 192 and 200 nm (3, 4), precolumn derivatized HPLC
by treatment with p-nitrobenzoyl chloride (5), and GC/MS after As a UV source for irradiation and photodegradation of
trimethylsilyl derivatization (6). UV detection (3, 4) suffers from sucralose, a 16 W mercury lamp with emission at 254 nm
lack of specificity and sensitivity, as measurements are done at was used. A Chemito Spectrascan UV 2600 double-beam
spectrophotometer (Thermo Fisher Scientific, Mumbai, India)
Received April 2, 2012. Accepted by SG July 10, 2012. with a computer system and Spectrum 6.89.8 software was
Corresponding author’s e-mail: md_idris3@yahoo.com used. The path length of the quartz cell was 1 cm, and the scan
DOI: 10.5740/jaoacint.12-143 speed was 1000 nm/s.
604 Idris et al.: Journal of AOAC International Vol. 96, No. 3, 2013

Figure 1. Sucralose structure.

Preparation of Standards and Sample

Standard solutions.—Stock solution of reference standard

Figure 2. Effect of UV radiation on sucralose in alkaline
sucralose was prepared in buffer solution (pH 12) at a medium.
concentration of 2 g/L. From the stock solution five different
concentrations of sucralose were prepared in the range of 0.1
to 1.2 g/L by diluting with buffer (pH 12) solution.
Sample preparation.—A 100 mg commercial sweetener
sample was transferred to a 100 mL volumetric flask containing
25 mL pH 12 buffer. The solution was sonicated for 10 min and
made up to the mark with pH 12 buffer solution. This solution
was filtered before UV irradiation.

Analytical Procedure

All the above standard and sample solutions were exposed to

Figure 3. Effect of pH on maximum product formation.
254 nm UV light for 1 h. These solutions were then scanned in
the UV range of 200–400 nm in a 1 cm cell against a solvent
blank buffer solution that does not contain sucralose exposed to
the same UV treatment for sucralose determination.

Method Validation

(a) LOD and LOQ.—LOD and LOQ were determined based

on the S/N ratio by scanning diluted solutions in the range of 0.1
to 1.8 g/L (made from stock solution of 2 g/L stock solution).
LOD was calculated as S/N × 3, and LOQ as S/N × 10. To detect
the noise, absorbance of a blank solution (buffer exposed to
Figure 4. Effect of UV exposure time on product formation
the same UV treatment as the samples) was recorded, and the (pH 12).
observed absorbance values for the blank and sucralose were
used to calculate the LOD and LOQ values.
(b) Linearity.—For linearity determination, stock solution by irradiating the sample solution at pH 12 for 1 h. Sucralose
(2 g/L) was diluted with pH 12 buffer to give a final concentration is probably photochemically oxidized to a UV-active carbonyl
of 0.1 g/L. Solutions of varying concentrations were checked compound characterized by a maximum absorbance at 270 nm
after irradiation for conformity to Beer’s law. (Figure 2) when scanned in the range of 200 to 400 nm.
(c) Precision and accuracy.—The precision and accuracy Formation of a carbonyl group was confirmed by reaction with
were determined with three different concentration levels
p-dinitrophenylhydrazine. Detailed spectroscopic (NMR, IR, and
covering the entire linearity range. The precision of the assay
was measured by repeatability (intraday, n = 6) and intermediate mass) studies to establish the photochemical reaction mechanism
precision (interday, n = 3) in terms of RSD for a statistically are in an advanced stage of progress and will be published
significant number of replicate measurements. Data from three separately. Simultaneously, a control analysis was carried out by
different concentration levels (0.4, 0.6, and 1 g/L) covering the keeping the solutions in the dark for 1 h and recording the UV
specified range were obtained (three replicates) for measuring spectrum. The solutions did not show any significant absorbance
accuracy.
in the range of 200 to 400 nm.
The formation of a photodegraded compound was monitored
Results and Discussion
by UV absorption spectrophotometry at the wavelength of
Sucralose is a non-UV-absorbing molecule due to lack of maximum absorbance (270 nm). The efficiency of the formation
a chromophore. UV determination was made possible in the of the UV-absorbing product is governed by several experimental
proposed method without using any derivatization reagents parameters, such as pH conditions and oxidation rate.
 Idris et al.: Journal of AOAC International Vol. 96, No. 3, 2013 605

Table 1. Quantitative parameters/optical characteristics of

the proposed method

Parameter Value

Beer’s Law limit, g/L 0.1–1.2

LOD, g/L 0.02
LOQ, g/L 0.07
Slope 0.0805
Intercept –0.0012
R2 0.9977
Figure 5. Stability of UV-active degradation product of
sucralose at room temperature (pH 12).
Table 2. Accuracy and precision data obtained for the
proposed method for spiked samples

Concentration found, g/L

Sample, amount Mean, Recovery,
of sucralose 1 2 3 ug/L ± RSD, % %

0.4 g/L 0.41 0.43 0.40 0.41 ± 1.62 103.25

0.6 g/L 0.59 0.59 0.58 0.59 ± 0.52 97.66
1.0 g/L 0.99 0.99 0.99 0.99 ± 0.33 99.2

Table 3. Intraday reproducibility data for spiked samples

Figure 6. Bottom spectra obtained for 0.1 g/L; top spectra
obtained for 1.2 g/L concentration of sucralose.
Amount of sucralose Intraday recovery, %
spiked, g/L (n = 6) Mean ± SD
pH Effect 0.40 103.0, 101.9, 104.1, 102.7, 103.0 ± 0.79
103.1
To study the effect of pH on the formation of a UV-active 0.60 97.7, 97.3, 98.9, 95.2, 96.7 97.16 ± 1.50
product, a photodegradation step was carried out at different pH 1.0 99.2, 101.7, 98.8, 102.3, 100.2 100.4 ± 1.38
values (12, 7, and 2). No changes were observed in the absorbance
spectrum of sucralose when UV irradiation was carried out for 1 h Table 4. Interday reproducibility data for spiked samples
at acidic or neutral pH, whereas a distinct band was observed at
270 nm under alkaline conditions (Figure 3). The studies carried Amount of sucralose Interday recovery, %
spiked, g/L (n = 3) Mean ± SD
out at different alkaline pH values indicated the optimum pH of
the photochemical reaction was 12.
0.40 103.0, 105.2, 101.3 103.14 ± 1.98

Oxidation Rate 0.60 97.7, 101.2, 100.0 99.44 ± 2.23

1.0 100.4, 99.0, 104.7 101.3 ± 2.86

The rate of the reaction was optimized by exposing the
same concentration of sucralose to UV radiation for different
intervals of time. It was found that the 1 h irradiation time was 40% (Figure 5). Therefore, it is very important to measure the
sufficient for maximum product formation (Figure 4). It was absorbance within 1 h after the irradiation process.
found in the literature (14) that chemical oxidizing reagents
promote a better reaction, but the oxidation is too drastic; the Validation Studies
UV-absorbing (carbonyl compound) intermediate was weakly
absorbed and quickly oxidized further to carboxylic acid. Thus, The photochemical reaction under optimized conditions
milder photolytic oxidation conditions should be used in order obeys Beer’s law in the concentration range of 0.1 to 1.2 g/L
to provide an appropriate oxidation reaction that allows the 2
(Figure 6) with a correlation coefficient (R ) value of 0.9977
formation of a stable compound with a suitable absorption peak. (Table 1). The LOD and LOQ were found to be 0.02 and
0.07 g/L, respectively. Accuracy of the method was determined
Stability of the UV-Active Degradation Product by the recovery studies, which indicated recoveries better than
97.6%.
Stability of the UV-active photodegradation product was Data related to the amount recovered from the spiked
studied by measuring the absorbance of the solution at room samples are given in Table 2. Reproducibility of the method
temperature at different intervals of time, and it was found that was evaluated by intraday and interday variation studies. For
up to 1 h there was a negligible change in the absorbance of intraday measurements, the reproducibility was SD less than
the solution. With an increase of time, a gradual decrease of 1.5, whereas interday studies showed an SD less than 2.9
absorbance was observed. After 24 h, absorbance decreased by (Tables 3 and 4).
606 Idris et al.: Journal of AOAC International Vol. 96, No. 3, 2013

Table 5. Data obtained by the proposed and reference Acknowledgments

methods for commercial sweetener samples
Mohd Idris would like to thank C.N. Bhattacharya, Director
Amount found by Amount found by
proposed method, reference method,
and Chief Forensic Scientist, Directorate of Forensic Science
Sample, tablet g/L g/L
a
% (± SD) Services, (MHA) Government of India, for providing a research
fellowship. The authors would like to thank A.K. Ganjoo,
1 (No label claim) 0.61 0.61 0.58
director, Central Forensic Science Laboratory, Hyderabad,
2 (No label claim) 0.64 0.64 0.75 India, for providing a necessary facilities for research work.
3 (No label claim) 0.63 0.64 0.63

4 (No label claim) 0.62 0.62 0.39 References

5 (No label claim) 0.65 0.65 0.55
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