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Determination of Sugar Solutions Color According to ICUMSA / Application Note Analytical Chemistry Chapter · January 2018
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Multiparameter Application NoteNote Multiparameter Application
Determination of Sugar Solutions Color According to ICUMSA 1. Introduction The measurement of both degrees Brix and solution color are required in sugar manufacturing industries, as a mean to grade product concentration, quality, or in blending operations. In fact, the formation of color is the result of specific steps of the manufacturing processes. In sugar refining, the more processing steps raw sugar is subjected to, the more color is removed: pigmentation of the product reflects the degree of refining to which it has been subjected. It helps to think of the process of refining as a series of steps, going from higher-colored raw sugar with lower market value, to very low-colored, refined white sugar with the highest market value.
Fig. 1: Sugar refining is a series of steps (left to right), where color and non-sugars are concentrated to the left, while pure sugar is concentrated to the right. Raw sugar comes into the process to left-of-center, not at one end.
Several scales for degrees Brix and color measurement are used in the food industry. ICUMSA (International Commission for Uniform Methods of Sugar Analysis) is a world-wide industry body that is concerned with analytical methods for the sugar industry. Many ICUMSA methods are used for the determination of sugar color in solution [1-5], and those are summarized in Table 1.
Table 1: ICUMSA methods for determine the color of sugar solutions.
Method
Title
Year Official
GS1/3-7
Determination of the solution colour of raw sugars, brown sugars and coloured syrups at pH 7.0
2011
GS2/3-9
The determination of sugar solution colour at pH 7.0
2005
GS2/3-10
The determination of white sugar solution colour
2011
GS2/3-18
The determination of the turbidity of white sugar solutions
2013
GS9/1/2/3-8
The determination of sugar solution colour at pH 7.0
2011
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1.1. Dictionary of sugars Raw sugar is the portion from the initial production process. It has a very high color value. Raw sugar must be refined and purified prior to consumption. Brown sugar and natural sugar are purified versions, and the color still has a high value. Refined and white sugar comes from raw sugar that is refined to extract impurities, and so has a low color value. This type of sugar is widely consumed among general households and used as raw material in food manufacturing plants, where moderately-purified sugar is needed, e.g. for drinks, sweetened condensed milk, etc. Super-refined sugars are those having undergone the refining process as white sugar, but they are purified to even a greater extent, resulting in a very low color value. The outstanding characteristic is its high purity. Most superrefined sugars are used in the food & beverage and in pharmaceutical industries. Molasses is a valuable by-product of sugar processing. It is mainly used as a raw material for the production of ethanol, liquor, animal feed, etc.
2. Material & methods 2.1. Instruments and accessories
®
Fig. 2: LabX enables the variety of laboratory line instruments to seamlessly integrate into a multi-parameter platform. The highlighted instrumentation is used in this work.
RM40 refractometer (51337003), UV5 Spectrophotometer (30254725), S400 pH meter (30046240) and XP 205 Analytical balance (11106027)
LabX software (30247984)
Quartz cuvettes 1.0 cm (30258738), 2.0 and 4.0 cm (commercial brand) and 5.0 cm (30258739)
Cellulose nitrate membrane filters, pore size 0.45 µm, vacuum pump and Büchner funnel
®
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2.2. Samples and reagents
Different sugar samples
Neutralizing agents:
o
For GS1/3-7: Hydrochloric acid (HCl; aq) (0.1M), Sodium hydroxide (NaOH; aq) (0.1M)
o
For GS2/3-9: Triethanolamine (TEA)/Hydrocholoric acid (HCl) pH 7.0 buffer solution (aq)
o
For GS9/1/2/3-8: MOPS (3-(N-morpholino)propanesulphonic acid)/Sodium hydroxide (NaOH) pH 7.00 buffer solution (aq)
Deionized water
3. Measurement Table 2: Differences in the various ICUMSA methods
20
Method
Cell length (b) in cm
pH regulation
Filtration
nD (SPS-3)
ICUMSA units
Transmittance in %
Rounding
GS1/3-7
1; 2; 5
HCl/NaOH
Yes
RDS
250
20 T 80
10
GS2/3-9
4
TEA/HCl
Yes
RDS*0.989
600
–
1
GS2/3-10
4
–
Yes
RDS
500
–
1
GS2/3-18
5
–
No
RDS
500
–
1
1; 2; 4; 5
MOPS
Yes
–
16 000
15 T 80
1 or 10
GS9/1/2/3-8
3.1. Sample preparation Weigh quantities of sugar or syrup and water, as shown in the corresponding ICUMSA method tables. Depending on the color range, sample aliquots and cell path lengths are selected that provide a transmittance in a preferred range. Dissolve samples by swirling at room temperature. Depending on the method (see Table 2), pH is measured and adjusted by using buffering or neutralizing agents, prior to color determination. Pass the sample solution through an appropriate filter. If necessary, deaerate the filtered solution at room temperature.
3.2. Procedure The principles of UV/VIS spectroscopy are described by The Lambert-Beer law: absorbance (A) maintains a linear relationship with the path length of the instrument (b), the concentration of the analyte (c), and the molar extinction coefficient of the analyte (a). The subscript denotes wavelength dependence. This relationship is mathematically expressed as:
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A = a b c
(Eq.1)
a is also known as absorbancy index of the analyte. The value of the absorbancy index (extinction index) at the wavelength of 420 nm is multiplied by 1000, and is subsequently reported as ICUMSA color. The resulting values are designated as ICUMSA Units (IU). If the solution is adjusted to pH 7.0, the units are designated as ICUMSA Units at pH 7.0 (IU7.0). Initially measure the refractometric refractive index of the dry substance (RDS) of the solution, using the ICUMSA method [6]. With the RM40 refractometer, the ICUMSA table is already built-in. Use the RDS to obtain the density of 3 th the test solution, in kg/m , from ICUMSA equation [7]. This is a polynomial equation of 6 order, and can be conveniently programmed as a calculation method function. The concentration of the test solution is described by:
5
c = (RDS)/10 g/mL
(Eq.2)
A UV5 spectrophotometer is used to determine absorbance (A420nm) of the sample with water serving as reference standard for zero absorbance (blank).
From the definition, ICUMSA Color = 1000a420nm, we obtain:
8
ICUMSA Color = (1000A420nm)/(bc) = (10 A420nm)/(bRDS)
(Eq. 3)
Finally, color is calculated in ICUMSA units and expressed to the nearest 1 or 10 IU.
3.3. RM40 parameters Measuring temperature
20.00 °C
Result 1
T[Brix_ICUMSA_nD(nD)]
Result 2
Density calculation via ICUMSA formula [6]
Result 3
Concentration calculation via ICUMSA formula [7]
3.4. UV5 method parameters Method
Fixed wavelength
Path lengths
1.0, 2.0, 4.0, or 5.0 cm
Measurement time
3s
Wavelength
420.0 nm
Background correction
1-point at 720.0 nm
Result 4
ICUMSA color: (A420*1000)/(b*concentration) [IU]
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4. Results 4.1. Method GS1/3-7 The method can be applied to raw sugars, affined raw sugars, higher color plantation white sugars and partly refined and brown sugars. Is designed for sugars having colors greater than 250 ICUMSA units at pH 7.0 (IU 7.0).
Table 3: Optical cells, reagents, settings, range and limits for method GS1/3-7. 20
Cell length (b) in cm
pH regulation
Filtration
nD (SPS-3)
IU
Transmittance in %
Rounding
1; 2; 5
HCl/NaOH
yes
RDS
250
20 T 80
10
In Table 3 and Fig. 3 the raw data (nD of a raw brown sugar are reported.
20
and A) and the ICUMSA color of five consecutively investigated specimen
20
Table 4: Repeatability of refractive index (nD ) and absorbance (A) measurements for raw brown sugar (n=5). Measurement
nD
20
A
1
1.38060
0.2016
2
1.38050
0.2018
3
1.38050
0.2029
4
1.38050
0.2014
5
1.38050
0.2018
Mean
1.38050
0.2019
S
0.00004
0.0006
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Multiparameter Application Note Fig. 3: Repeatability of ICUMSA color results for raw brown sugar measurements (n=5). Dotted lines indicate the maximum absolute difference allowed for measurement repeatability, as indicated in ICUMSA GS 1/3-7 (110 IU7.0 for this sample and color range).
ICUMSA defines the maximal permitted repeatability as the absolute difference between two results obtained under repeatability conditions. In Table 5 and Fig. 4 results of ICUMSA color determination of five consecutively investigated specimen of whiteand brown raw sugars are reported. Due to different solution concentrations, varying cell path lengths are used. For White 1 and White 2 raw sugar samples, path lengths of 5 and 2 cm are used, respectively. For brown raw sugars, cuvettes maintaining 1 cm path lengths are used.
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Table 5: Mean values from measured raw sugar samples (n=5) for ICUMSA color along with repeatability data for both the measured and permitted repeatability. All values are presented in IU7.0.
Sample
Color
Measured repeatability
Max. permitted repeatability*
White 1
3280
30
N/A
White 2
3340
30
N/A
Brown 1
3600
20
110
Brown 2
4080
10
300
Brown 3
5310
30
300
* as defined by ICUSMA Method GS1/3-7
Fig. 4: ICUMSA color results for white and brown raw sugars (n=5). Dotted lines indicate the maximum absolute difference allowed for measurement repeatability, as indicated in ICUMSA GS 1/3-7 for sample and color range.
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4.2. Method GS2/3-9 This method can be applied to all crystalline, powdered white sugars and very pure syrups of color values up to 600 IU7.0.
Table 6: Optical cells, reagents, settings, range and limits for method GS2/3-9. 20
Cell length (b) in cm
pH regulation
Filtration
nD (SPS-3)
IU
Transmittance in %
Rounding
4
TEA/HCl
yes
RDS*0.989
600
–
1
In Table 7, results of ICUMSA color for two white sugars are reported. Sugars are dissolved in TEA/HCl buffer solutions adjusted to a pH of 7.0. The concentration of sample solids in solution is calculated from the RDS measurements. To allow for concentration of TEA/HCl buffer in the test solution, measured RDS values must be multiplied by 0.989, in order to express the "corrected RDS". For this measurement, a cuvette maintaining a 5 cm path lengths was used.
Table 7: Mean values from measured white sugar samples (n=5): ICUMSA color along with data for both the measured and permitted repeatability. All values in IU7.0.
Sample
Color
Measured repeatability
Max. permitted repeatability*
White 1
386
3
49
White 2
362
0
44
* as defined by ICUSMA Method GS2/3-9
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4.3. Methods GS2/3-10, GS2/3-18 GS1/3-10 is the method used for determining white sugar solution color values for all crystalline or powdered white sugars, and very pure syrups. GS2/3-18 is the method for determining turbidity in white sugar, and is a derived of ICUMSA method GS1/3-10. Turbidity is calculated as the difference in color of white sugar solutions before and after filtration. For both methods, a pH adjustment is not needed. Resulting values are designated as ICUMSA Units (IU).
Table 8: Optical cells, reagents, settings, range and limits for method GS1/3-7. 20
Cell length (b) in cm
pH regulation
Filtration
nD (SPS-3)
IU
Transmittance in %
Rounding
GS2/3-10
4
–
yes
RDS
500
–
1
GS2/3-18
5
–
no
RDS
500
–
1
Method
Fig. 5: ICUMSA color repeatability results for crystalline white sugar measurements (n=5). Dotted lines indicate the maximum absolute difference allowed for measurement repeatability, as indicated in ICUMSA GS 2/3-10 (3 IU).
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4.4. Method GS9/1/2/3-8 This method is applicable for plantation white sugars, raw sugars, affined raw sugars and very low-color sugars. It is designed for all sugars in the solution color value up to 16 000 ICUMSA Units at pH 7.0 (IU7.0).
Table 9: Optical cells, reagents, settings, range and limits for method GS9/1/2/3-8. 20
Cell length (b) in cm
pH regulation
Filtration
nD (SPS-3)
IU
Transmittance in %
Rounding*
1; 2; 4; 5
MOPS
yes
–
16 000
15 T 80
10 or 1
* for results over 1000 IU7.0, nearest 10 IU7.0; for results under 1000 IU7.0, nearest 1 IU7.0.
In Table 10 and Fig. 6, results for ICUMSA color of five consecutively-investigated specimen of different white-, brown- and raw sugars are reported. Due to differences in solution concentrations, varying cell path lengths were used. For white 1, 2 and 3 sugars, a 5 cm path length was used. For brown 1 and 2 sugars, cuvettes maintaining a 2 cm path length were used. For raw 1 and 2 sugars, cuvettes with 1 cm path length were used.
Table 10: Mean values (n=5) from measured white, brown and raw sugar samples: ICUMSA color, measured and permitted repeatability. All values in IU7.0.
Sample
Color
Measured repeatability
Max. permitted repeatability*
White 1
3397
32
339
White 2
3183
33
339
White 3
3264
34
323
Brown 1
3673
12
348
Brown 2
3732
39
348
Raw
1
3070
10
220
Raw
2
3590
10
220
Raw
3
3850
10
220
* as defined by ICUSMA Method GS9/1/2/3-8
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Multiparameter Application Note Fig. 6: ICUMSA color results for different white, brown and raw sugars (n=5). Dotted lines indicate the maximum absolute difference allowed for measurement repeatability, as indicated in ICUMSA GS9/1/2/3-8 for sample and color range.
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4.5. The Effect of pH It is well known that absorbance values of sugar solutions are pH-dependent. In turn, this can modify the ICUMSA Color [8], especially at high color values. To illustrate this observation, an experiment on brown sugar samples was performed to determine subsequent ICUMSA color as function of differing pH of the solutions. After sugar dissolution, a pH of 6.43 was obtained. As described in method GS1/3-7, the appropriate pH adjustment is performed with the addition of either HCl or NaOH. The upper diagram in Fig. 7 demonstrates the pH-dependence of the ICUMSA color profile of a brown sugar solution. The relationship is sigmoidal in nature: an absorbance increase occurs when the pH value increased, whereas the reverse trend is observed by lowering pH. An interpolation of data points is carried out using a generalized logistic functional form. The latter is used for illustrative purposes, and serves to calculate the first derivative. The lower diagram of Fig. 7 depicts the first derivative of the respective color by pH of 0.1 units. The absolute value of ICUMSA color for this brown sugar solution at pH 7.0 was 610 IU7.0. At a pH of 6.43, just after sugar dissolution and without adjusting pH, a color value of 525 IU was obtained. The difference between the two results amounted to 85 IU. Analysis of the first derivative indicated that, at around pH 7 0.1 units, a corresponding difference of 18.2 IU was observed. Based on these two observations, the following conclusions were reached: pH values substantially impact measurements, and accurate determination of pH is essential to repeatable and reproducible data. Thus, ICUMSA color values are inherently sensitive to even small variations and errors in pH determination or pH solution adjustment.
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Multiparameter Application Note Fig. 7: The upper diagram depicts brown sugar solution ICUMSA color as function of pH. The lower diagram reports first derivative values as a function of 0.1 pH.
It has been reported that results from the lower color range up to about 60 IU are nearly equal in both pH corrected and non-corrected methods [9], and are well-correlated. The results obtained for higher color ranges show greater deviations between both methods, and those results obtained without pH adjustment were much lower. Therefore, for color values over 60 IU, solution color is pH-dependent. If a sugar has a solution color less than 60–65 IU, then the simpler, more eco–friendly method, GS2/3-10, can be used. Irrespective of the sugar solution color measured, the recommendation is to always state the method used in determining the color value.
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4.6. The relationship between path length and linearity For optimal measurements that comply with the Lambert–Beer law, absorbance must be in the linear range, and is therefore dependent on concentration of the analyte path length and instrument performance. Absorbance A of a sample is calculated via transmittance, and is expressed as -log10T. This is why some ICUMSA methods specify working with a given path length or transmittance interval. For low IU sugars, cell lengths no smaller than 4 or 5 cm are recommended (GS2/3-9, GS2/3-10 and GS2/3-18). However, a fundamental question still remains: how to achieve results within the range of linearity for a given path length and instrument selected to measure a specific sample concentration? One solution available to resolve this challenge is to simply measure a series of the same sample with varying concentrations. Absorbance values for sugar solutions are measured using a fixed path length and a series of diluted samples, as shown in Fig. 8. The initial stock concentration used is indicated as c0/1, and successive 50-50% solution–water dilutions were measured sequentially. As shown in the graph, all sample concentrations tested returned measured absorbance values within the range of linearity of the path length and instrument used. According to the linear relationship described by The Lambert–Beer law, the absorbancy index a is an intrinsic property of the absorbing species, and this value does not change unless the chemical nature of the species changes. Hence, decreasing the concentration will have the same resultant effect as increasing path length. Thus, in the linear range, dividing c0 by a factor, corresponds to multiply the path length b0 by the same amount, as represented in the alternative x–axis (top of the diagram).
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Multiparameter Application Note Fig. 8: Lambert–Beer linearity: measured values are in the linear range of absorbance.
For this experiment, c0 corresponds to 0.5859 g/mL, and b0 is 1.0 cm. Fig. 8 demonstrates absorbance values with a very high degree of linearity across large concentration and path length ranges. By varying these two parameters, sample concentrations can be maintained within the range of linearity for the instrument, as described by the Lambert–Beer relation. This is further indicated by the linear interpolation, y = 1.090x - 0.01, where the 2 slope and offset are close to unity and zero respectively. The correlation coefficient (R ) approaches 1.000. These data support the use of standard-sized cuvettes (1.0 cm) as sufficient for determining ICUMSA colors. Using larger path lengths does not improve quality of the results.
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4.7. Miscellany Software associated with instrumentation is programmed to process the results algebraically from concentrations to ICUMSA color calculations. Results are generated fully automatically, avoiding tedious data transcription and manual calculations. RDS and are also automatically retrieved, either by table lookup or using a polynome. Depending on the ICUMSA method, aliquots of sugar and water will differ, and must be within specified tolerances. The same is true for pH, transmittance and cell length. Repeatability thresholds are also tabulated. Automatic parameter selection, tolerances control and documentation are operations carried out by software associated with the instrumentation.
4.8. An infrastructure for quality control Harvesting sugarcane or sugar beet is a seasonal activity, and therefore occurs during a defined and limited time of the year. Often very large harvest quantities are involved. The juncture between these two factors poses major challenges to sugar manufacturers, since the incoming raw sugar must be rapidly processed. Effective TM automatization solutions like InMotion and SC30 autosampler can be used to increase testing throughput. Additionally, software support is also available to process the flow of results for quality control purposes. Statistical evaluations can be routinely used to test results against a target value or tolerance, generating automatic Pass/Fail reports.
5. Conclusions ICUMSA color of sugar solutions is calculated as function of sugar concentration and UV/VIS absorbance: refractive index of the solution and absorption at 420 nm are measured. Final results are given in ICUMSA Units (IU or IU7.0, rounded or not), and calculated from the sugar concentration, the absorption value and the path length used for ® measurement. The sugar concentration is determined using Brix value and specific ICUMSA formulae. With LabX , the laboratory Software for full user guidance and automatic data handling, the data access method function can be used to transfer all results to a calculation method. Thus, the ICUMSA color is fully calculated automatically within ® LabX . ®
Using LabX PC Software, limits can be easily set for ICMUMSA color, with easy implementation of automatic tolerance checks of measurement results. In the result overview, customized text and background color codes can be used for easy identification of sugars for pass/fail tolerance checks. For all investigated sugar solutions, repeatability of the measurements was excellent and well within the maximallytolerated repeatability criteria. In summary, we establish a highly reproducible method, and offering fully automated ICUMSA color calculations.
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References [1] ICUMSA Method GS1/3-7 Determination of the Solution Colour of Raw Sugars, Brown Sugars and Coloured Syrups at pH 7.0 – Official (2011) [2] ICUMSA Method GS2/3-9 The determination of sugar solution colour at pH 7.0 – Official (2005) [3] ICUMSA Method GS2/3-10 The determination of white sugar solution colour – Official (2011) [4] ICUMSA Method GS2/3-18 The determination of the turbidity of white sugar solutions – Official (2013) [5] ICUMSA Method GS9/1/2/3-8 The determination of sugar solution colour at pH 7.0 – Official (2011) [6] ICUMSA Specification and Standards SPS-3 Refractometry and Tables – Official (2000) [7] ICUMSA Specification and Standards SPS-4 Densimetry and Tables – Official (1998) [8] K. Sing et al., Recent developments in white sugar solution colorimetry, Sugar Industry (3), 2013:159–163. [9] H. Puke, C. Lakenbrink, A treatise on ICUMSA solution colour – Part 2: Studies and conclusions, Sugar Industry (3), 2013:153–158.
Disclaimer This report presents one or more examples of analysis. The experiments were conducted with the utmost care using the instruments specified in the description of each application. The results have been evaluated according to the current state of our knowledge. This does not however absolve you from personally testing the suitability of the examples for your own methods, instruments and purposes. Since the transfer and use of an application is beyond our control, we cannot of course accept any responsibility. When chemicals, solvents and gases are used, the general safety rules and the instructions given by the manufacturer or supplier must be observed.
Further information www.mt.com/DERE www.mt.com/UVVIS https://youtu.be/lZWj_Y9PrWg Video-URL
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