Dextran in Beet Raw Juice and Thick Juice by Spectrometry using a Modified Alcohol Haze Method – Accepted

ICUMSA Method GS8-19 (2026)

 

1 Warnings

Warning and Safety Precautions
Persons using this ICUMSA^® Method should be familiar with normal laboratory practice. This standard does not address all the safety issues, if any, associated with its use. It is the responsibility of the user to establish appropriate safety and health practices and to ensure compliance with any national regulatory conditions.

Disclaimer
The mention of specific companies or of certain manufacturers’ products does not imply that they are endorsed or recommended by the International Commission for Uniform Methods of Sugar Analysis (ICUMSA^®) in preference to others of a similar nature that are not mentioned.

General advice
ICUMSA^® Methods are updated from time to time following scientific and technical development. Please check on https://www.icumsa.org, whether the Method in hand is the current version.
This method edition cancels and replaces the previous (ICUMSA^® Method GS8-19 (2017)), which has been rewritten.

 

2 Scope and field of application

2.1 Scope
This method is used for the spectrophotometric determination of dextran content in beet raw juice and thick juice after removal of protein [1,2]. Beet raw juice or thick juice can be tested as fresh or thawed material that has been previously frozen.

2.2 Field of application
This method is applicable to beet raw juice and thick juice only, as these are the only sample types it has been validated for [2, 3]. In general, it may be appropriate for monitoring the dextran content in juices and syrups during beet processing or in stored thick juice. The raw juice or thick samples may need to be diluted to fall within the calibration curve’s dextran content range. Whether an adaptation by dilution is necessary must be determined prior to the analysis for each sample material.
Dextran in beet raw juice causes severe processing problems, especially in filtration. If dextran content measurements for raw juices of deteriorated beet are to be determined using this method, the precipitation of other higher molecular mass substances could result in over-estimation of dextran content.

 

3 Normative references

The following documents are referred to in the text in such a way that some or all of their content constitutes requirements of this document. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies.

• ICUMSA^® Specification and Standard SPS-11 (2022) Sampling, Sample Handling and Sample Preparation of Liquid Sugar Products, Syrups and Molasses [4]
• ICUMSA^® GS2-15 (2007) The Determination of Sugar Moisture by Loss on Drying [5]
• ICUMSA^® Method GS4-12 (1998) The Determination of Moisture by the Karl Fischer Procedure in Molasses – Official and in Cane Raw Sugars, Speciality Sugars and Syrups – Tentative [6]

 

4 Terms and definitions

ICUMSA^® maintains a terminological database (under development) for use in standardisation https://www.icumsa.org.
For the purposes of this document, the following terms and definitions drawn from the database apply:

4.1 Dextran is defined as a high molecular mass and predominantly straight-chained glucose polymer with a majority of α-(1→6) glucosidic linkages, formed by the action of certain species of bacteria, especially Leuconostoc mesenteroides, on sucrose.

4.2 Raw juice and thick juice are liquids from sugar beet processing. Raw juice is obtained after the extraction step of the beet cossettes. Thick juice is obtained by concentrating the juice after juice purification using lime and carbon dioxide. Raw and thick juice used in this method are needed for calibration (4.3, 4.4) and serve as samples for analysis (8.2, 8.3). Collection of any liquid sample should be performed to align with local procedures for sampling, to ensure that the material collected is representative of the process being sampled.

4.3 Calibration raw juice. Calibration raw juice is an unaltered raw juice (4.2) that has been collected during periods with no frost, which is usually the case at the beginning of the processing period (to ensure unmeasurable or no native dextran content). Preparation and storage of calibration raw juice should follow the same procedure as for samples (8.2). If analysis is not performed soon after collecting the calibration raw juice, it must be frozen (8.2.2). Subsamples of calibration raw juice are used to prepare solutions (4.9, 4.11, 4.13) to establish a calibration curve (9.1.2). The sugar (sucrose) and non-sugar (non-sucrose) contents in the calibration raw juice should be similar to those of samples under test (8.2).

4.4 Calibration thick juice. Calibration thick juice is an unaltered thick juice (4.2) that has been collected during periods with no frost, which is usually the case at the beginning of the processing period (to ensure unmeasurable or no native dextran content). It is recommended that the collected calibration thick juice is diluted (e.g. 1:1 by mass) and frozen if analysis is not performed soon after collection. Subsamples of calibration thick juice are used to prepare sample solutions (4.10, 4.11, 4.13) for preparing a calibration curve (9.1.3). The sugar (sucrose) and non-sugar (non-sucrose) contents in the calibration thick juice should be similar to those of the samples under test (8.3).

4.5 Transmittance of a solution. If P_λ⁰ represents the radiant energy incident upon the first surface of the solution and P_λ represents the radiant energy leaving the second surface of the solution. Then:

\displaystyle T(l)=\frac{{{{P}_{\lambda }}}}{{P_{\lambda }^{0}}}     internal transmittance of the solution with pathlength l

4.6 Transmittance ratio. Let T_soln represent the transmittance of a cell containing the solution and let T_solv represent the transmittance of the same or duplicate cell containing the pure solvent. Then:

\displaystyle {{R}_{\text{T}}}=\frac{{{{T}_{{\text{soln}}}}}}{{{{T}_{{\text{solv}}}}}}     transmittance ratio of the solution

4.7 Absorbance

A(λ)_S = –lg R_T(λ)_S       asorbance of the solution, λ = wavelength in nm

4.8 Absorption coefficient. Let l represent the length (cm) of the absorbing path between the boundary layers of the solution and let β represent the mass concentration (g/mL) of the sugar solution. Then:

\displaystyle \varepsilon {{(\lambda )}_{\text{S}}}=\frac{{A{{{(\lambda )}}_{\text{S}}}}}{{l\cdot \beta }}     absorption coefficient of the solution

4.9 Calibration raw juice solution. The calibration raw juice solution is the solution prepared from the calibration raw juice (4.3) being mixed with TCA solution (6.4) and a defined amount of standard dextran solution (6.2), prepared as described in section 9.1.2.2. A range of calibration raw juice solutions is required to determine the dextran content in any given raw juice sample.

4.10 Calibration thick juice solution. The calibration thick juice solution is the solution prepared from the calibration thick juice (4.4) being mixed with TCA solution (6.4) and a defined amount of standard dextran solution (6.2), prepared as described in section 9.1.3.2. A range of calibration thick juice solutions is required to determine the dextran content in any given thick juice sample.

4.11 Sample blank. A solution that is essentially the sample matrix with no analyte added. For this method the sample blank is a solution prepared using the sample solution to be analysed (8.2. or 8.3), that is mixed with TCA solution (6.4) and diluted to volume described in section 9.2.2, with no addition of DAA (6.7).

4.12 Reagent blank. A mixture of any solvent(s) and/or reagent(s) that would be presented to the detector for analysis of a sample. A reagent blank is analysed to determine if it contributes to the measurement signal and is often used with techniques such as spectrophotometry to zero the instrument before measuring test samples. For this method the reagent blank is the sample blank (4.11) that has had DAA (6.7) added to it as described in sections 9.1.2.3, 9.1.3.3.

4.13 Calibration blank. A solution that is essentially the calibration matrix without precipitant, DAA (6.7), added. For this method the calibration blank is a solution prepared using either calibration raw juice (4.3) or calibration thick juice (4.4), that is mixed with TCA solution (6.4) and standard dextran solution (6.2) and diluted to volume as described in sections 9.1.2.2 and 9.1.3.2.

4.14 Dextran content in raw juice. The dextran content determined by this method for a sample of raw juice (4.2 and 8.2) is expressed as a mass concentration (β_RJ,Dx) in mg/L. A calibration curve (9.1) is used to determine the dextran content of a raw juice sample (9.2.3). Raw juice produced from sugar beet collected after frost conditions and a subsequent thawing period is highly likely to contain dextran.

4.15 Dextran content in thick juice. The dextran content determined by this method for a sample of thick juice (4.2) is expressed as a mass concentration (β_TJ,Dx) in mg/L. A calibration curve is used to determine the dextran content of a thick juice sample (9.2.3). Thick juice that has been produced from sugar beet collected after frost conditions and a subsequent thawing period is highly likely to contain dextran.

 

5 Principle

This method measures the haze formed by dextran-like polysaccharides when alcohol is added to a raw juice or a diluted thick juice. Protein is removed by precipitation with trichloroacetic acid solution followed by filtration with acid-washed filter aid. The dextran haze is produced by diluting an aliquot of treated solution to twice the aliquot volume, by addition of ethanol. The absorbance of the turbidity produced by the dextran haze is measured at a wavelength of 720 nm and is proportional to the dextran content. The method is standardised against a commercially available dextran.

 

6 Reagents

General advice. During the analysis, unless otherwise stated, use reagents only of analytical grade and only deionised or distilled water.

Warning and Safety Precautions. Users of this method are advised to consult their national health and safety legislation before handling trichloroacetic acid, denatured absolute ethanol and concentrated hydrochloric acid.

6.1 Standard dextran – (C₆H₁₀O₅)_n, T110 or T500 (CAS#9004-54-0), technical quality (e.g. D5376). The performance of different sources of dextran may vary so it is important to test how it performs in the calibration range desired (see section 9). The standard dextran contains bound water, whose content must be determined.

6.2 Standard dextran solution – 2 mg/mL. Quickly weigh (7.1) a quantity of standard dextran (wet, 6.1), that contains approximately 0.4 g of dried dextran and record the mass (m_Dx,STD,wet) to ±0.0001 g (6.1). The wet mass required must be calculated (10.1.2) after the moisture content of dextran (w_Dx,STD,W) has been determined (see section 9.1). Transfer the wet dextran into a 100 mL beaker (7.2) and dissolve the dextran by adding 2 mL to 4 mL of water to form a slurry. Allow the particles to become uniformly hydrated by standing for about 10 min (7.3) with occasional mixing. At the end of the standing period, add more water in small aliquots, with gentle mixing. When about 50 mL has been added and a gel is no longer present, wash the slurry into a 200 mL volumetric flask (7.2) with water, to a volume of about 100 mL. Place the flask in a boiling water bath (7.4) for 30 min. Cool to room temperature (22 ± 2 °C) in a cold-water bath (7.5), then make up the mark with water. Prepare a standard dextran solution daily, do not store prepared solutions overnight.

6.3 Trichloroacetic acid (TCA) – C₂HCl₃O₂, analytical reagent grade (CAS# 76-03-9).

6.4 Trichloroacetic acid (TCA) solution – 100 g/L. Dissolve (20 ± 0.1) g of trichloroacetic acid (6.3) in distilled water and dilute to 200 mL using a volumetric flask (7.2). The shelf life of TCA solution is two weeks, if it is stored in a light-shielded bottle, in the fridge.

6.5 Absolute ethanol – C₂H₆O, >98.8 %, analytical reagent grade (CAS# 64-17-5).

6.6 Methanol – CH₄O, >99.9 %, analytical reagent grade (CAS# 67-56-1).

6.7 Denatured absolute alcohol (DAA) – absolute ethanol (6.5) with (2.0 ± 0.2) g/100 g of methanol (6.6), and with a water content of less than 0.5 g/100 g. If particulate matter is visible, filter through a filter paper (6.15). Store in an airtight container.

6.8 Acid-washed filter aid – inert, silica-based/diatomaceous earth powder with high porosity, that has been acid-washed (e.g. kieselguhr, SiO₂ (CAS#61790-53-2).

 

7 Apparatus

7.1 Analytical balance – readable to 0.0001 g.

7.2 Glassware (for the preparation of samples and calibration solutions) – beakers of approximately 150 mL capacity, volumetric flasks and associated closures (25, 100 and 200 mL), light-shielded/brown glass storage bottle (≥200 mL), funnel (to fit a 100 mL volumetric flask), bulb pipettes (1, 2, 5 and 10 mL), graduated pipettes (1, 5 and 10 mL), burettes (25 or 50 mL), conical flasks (200 mL). An automatic dispensing pipette may be used for TCA solution (6.4).

7.3 Stopwatch – readout in seconds and minutes.

7.4 Boiling water bath – for preparation of the standard dextran solution (6.2).

7.5 Cooling water bath (20 °C) – a water bath containing cold tap water for cooling, but ideally a temperature-controlled cold-water bath.

7.6 Thermometer – suitable for measurement of solution temperature to ensure it is at room temperature (22 ± 2 °C).

7.7 Sieve – for removal of solids from raw juice samples collected from the process. The material of the sieve should be chosen to be suitable for this task and the recommended mesh size should be 5 mm.

7.8 Storage containers (for samples to be frozen) – Sample containers of low volume (≤100 mL is recommended) that are suitable for freezing and storage of prepared standard raw juice (4.3) and standard thick juice (4.4) samples.

7.9 Vacuum pump or vacuum supply – suitable for applying low-pressure for filtration.

7.10 Filter paper – filter paper with medium retention and flow rate, suitable for retention of coarse particles or gelatinous precipitates. The paper must be suitably sized for use with the vacuum apparatus (7.11). (e.g. 5.5 cm Whatman Grade 5 or equivalent).

7.11 Filter funnel and flask – rerequired for filtration of sample and calibration solutions (section 9), with a recommended flask capacity of ≥250 mL and funnel diameter appropriate for the filter paper (7.11) in use.

7.12 Spectrophotometer – double beam instrument capable of light transmission measurement at a wavelength of (720 ± 10) nm with the narrowest practical bandwidth. The device needs to be suitable for use with matched 2 cm or 5 cm cells. The spectrophotometer should comply with the following specificationss:
• Wavelength reproducibility ± 0.5 nm,
• Absorbance reproducibility ± 0.003 at 1.0 absorbance.

7.13 Associated optical cells – use two matched cells of either 2 cm or 5 cm. One cell is intended to contain the solution to be examined (4.9, 4.10, 4.14 or 4.15), and the other cell is intended to contain the reference solution (4.11, 4.12 or 4.13) for compensation.

 

8 Sampling, sample handling and sample preparation

8.1 Sampling procedure. For sampling of beet raw or thick juice (4.2), the guidelines in SPS-11 (2022), Sampling, Sample Handling and Sample Preparation of Liquid Sugar Products, Syrups and Molasses should be followed. Local procedures for sampling of raw juice or thick juice samples containing visible solids should be followed, ensuring that the material collected is representative of the process being sampled.

8.2 Preparation of raw juice samples. After collecting raw juice in bulk, ensure each sample is thoroughly mixed before sub-sampling.

8.2.1 Preparation of fresh raw juice samples. The collected bulk raw juice sample (8.2) must be sieved to remove the majority of the solids from the juice (e.g. pulp, soil, etc.). It is recommended that a fine-mesh sieve (7.7) is used to do this. The raw juice sample may be used directly to determine dextran content (9.2) or frozen for later analysis. If the raw juice sample is to be stored, it should be portioned into containers (7.8) and frozen immediately.

8.2.2 Preparation of thawed raw juice samples. Samples of frozen raw juice (8.2.1) must be thawed quickly, but gently, to ensure that the warming process doesn’t degrade the sample. It is recommended to warm the frozen samples in a 20 °C water bath (7.5). The samples should be well-mixed once defrosted, prior to proceeding with determination of dextran content (9.2).

8.3 Preparation of thick juice samples. After collecting thick juice in bulk, ensure each sample is thoroughly mixed before sub-sampling.

8.3.1 Preparation of fresh thick juice samples. It is recommended that the collected and mixed thick juice (8.3) is diluted prior determination of dextran content (9.2). The dilution may need to be determined, depending on the thick juice sampled (8.1), but a 1:1 dilution should be suitable [3]. If the thick juice is to be stored, it should be portioned into containers (7.8) and frozen immediately.

8.3.2 Preparation of thawed thick juice samples. The same procedure as described for frozen raw juice samples (8.2.2) should be followed to thaw thick juice samples (8.3.1). The samples should be well-mixed once defrosted, prior to proceeding with determination of dextran content (9.2).

 

9 Procedure

9.1 Preparation of the calibration curve. Separate procedures (9.1.2 and 9.1.3) are required for preparation of calibration curves for raw juice and thick juice analysis. The moisture content of the standard dextran (6.1) must first be determined (9.1.1) to enable calculation of the dry dextran content of the standard dextran solution (6.2). There is no definitive evidence for the validity of the calibration curve [3], but standard laboratory procedures should be used as a guideline for suitability of the use of the calibration curve (e.g. change in reagents used in samples, change in expected response for samples, etc. would indicate that the calibration curve may be invalid or it should be revalidated).

9.1.1 Determination of moisture content of standard dextran. Determine the moisture content of the standard dextran (6.1, 10.1.1), w_Dx,STD,W, in duplicate, to two decimal places, by following the same procedure that is used for determination of moisture content of white sugar (Method GS2-15 (2007)). Individual determinations must be within 1 % of the mean of the moisture content. Karl Fischer moisture content determination is recommended for a quick and precise value for moisture content, and Method GS4-12 (1998) could be adapted for this purpose.

9.1.2 Calibration curve for raw juice. Samples of prepared (8.2.1 and 8.2.2) calibration raw juice (4.3) should be at room temperature before proceeding with the determination. The moisture content of the dextran used in the standard dextran solution (6.2) must be determined (10.1.1) before the dry dextran content in the calibration solutions (β_{RJ,Dx,STD,dry}) can be calculated (10.1.3) and used to prepare the calibration curve for raw juice.

9.1.2.1 Determination of suitability of calibration raw juice for dextran analysis. The suitability of the calibration raw juice (4.3) for dextran analysis must be checked prior to proceeding with the determination. This is done by measuring the absorbance of the reagent blank solution diluted to volume (4.12, 9.1.2.2), A(720)_RJ,RBl, against the absorbance of the calibration blank solution (4.13) (without any standard dextran solution added) diluted to volume, A(720)_RJ,CALBl. (Line 1 in Table 1 with and without DAA added). The corrected absorbance (A(720)_RJ,RBl – A(720)_RJ,CALBl) should not exceed the following values, which depend upon the optical cell (7.13) in use: 0.003 for a 2 cm cell or 0.008 for a 5 cm cell.

9.1.2.2 Preparation of calibration raw juice solutions for absorbance measurement. Prepare the calibration raw juice solutions using the calibration raw juice (4.3), TCA solution (6.4) and the standard dextran solution (6.2) using 100 mL volumetric flasks (7.2) and following the preparation details shown in Table 1. A set of calibration raw juice solutions is required for a calibration curve. The dextran content (β_{RJ,Dx,STD,wet}) listed in Table 1 needs to be corrected to obtain the dried dextran content β_{RJ,Dx,STD,dry} (10.1.3).

 

Table 1: Calibration raw juice solutions

Flask No.	Calibration raw juice (4.3) (mL)	TCA solution (6.4) (mL)	Standard dextran solution (6.2) (mL)	Wet dextran content in calibration raw juice (mg/L) βRJ,Dx,STD,wet
1	50	10	0.0	0
2	50	10	2.5	100
3	50	10	5.0	200
4	50	10	10.0	400
5	50	10	15.0	500

 

After addition of the TCA solution, cool the flasks in a 20 °C cooling water bath (7.5) before making up to volume with de-ionised water. Stopper the flasks and mix the solutions thoroughly.
Pour each prepared calibration raw juice solution into a 150 mL beaker (7.2). Add approximately (6 to 8) g of acid-washed filter aid (6.8) to each beaker and mix well. Filter the mixture under low-pressure (7.9) through filter paper (7.10) fitted to a funnel connected to a vacuum flask (7.11). Apply vacuum and filter (10 to 15) mL solution. Disconnect the vacuum and use the filtrate to rinse the flask, then dispose of the filtrate. Reconnect the funnel and flask to the vacuum and filter the remaining solution.

9.1.2.3 Absorbance measurement of calibration raw juice solutions. Using a pipette (7.2), transfer an aliquot of 12.5 mL of a calibration raw juice solution filtrate (9.1.2.2) to each of two dry 25 mL volumetric flasks (7.2). Slowly add DAA (6.7) to one of the 25 mL flasks from a burette (7.2) up to the 25 mL mark while gently swirling the flask. Use a stopwatch (7.3) to ensure that the time taken to add the DAA is 30 to 60 s. Stopper the flask and mix the contents by inverting gently three times. Start the stopwatch (7.3) immediately after the mixing step is complete.
Approximately 17 min after the completion of the mixing step, rinse one of the reference optical cells (7.13) three times with the solution and then fill the cell. Clean the optical faces of the cells with a tissue and check that the solution in the cell is uniform (striations are absent). and place in the sample position of the instrument. This is a calibration raw juice solution.
Make up the second 25 mL flask to volume with water. Rinse the other optical cell (7.13) three times with the solution and then fill the cell. Clean the optical faces of the cell with a tissue and check that the solution in the cell is uniform (striations are absent), place this cell in the reference position of the instrument. This is the calibration raw juice blank solution.
At 20 min ± 10 seconds after completion of the mixing step read and record the corrected absorbance, A(720)_{RJ, CAL,Corr}, to 0.001 absorbance units.
Repeat the above steps for each calibration raw juice solution prepared in Table 1, using the calibration raw juice blank solution in the reference position for each calibration thick juice solution.

 

NOTE 1 – Alcohol must be added within 20 min of addition of dextran solution to the juice/TCA solution.
NOTE 2 – As the absorbance must be read at a precise time after the mixing step it is recommended that alcohol be added to the dextran standards at uniform time intervals (3 min or 4 min).
NOTE 3 – The filtrate of Flask No. 1 (Table 1, no dextran addition) is the calibration blank (4.13, 9.1.2.2) after diluting the filtrate with de-ionised water to volume and mixing.

 

9.1.2.4 Calculation of calibration curve for raw juice. Calculate the dry dextran content values for the calibration raw juice solutions (β_{RJ,Dx,STD,dry}) (10.1.3) and plot these against the corresponding corrected absorbance values (A(720)_RJ,CAL,Corr) to produce a raw juice calibration curve. The calibration curve should be evaluated using standard laboratory protocols and also include the determination of the coefficient of determination R² (Note that the coefficient of determination R² should be greater than 0.9900).

9.1.3 Calibration curve for thick juice. Samples of prepared (8.3.1 and 8.3.2) calibration thick juice (4.4) should be at room temperature before proceeding with the determination. The moisture content of the dextran used in the standard dextran solution (6.2) must be determined (10.1.1) before the dry dextran content in the standard solutions (β_{TJ,Dx,STD,dry}) can be calculated (10.1.3) and used to prepare the calibration curve for thick juice.

9.1.3.1 Determination of suitability of calibration thick juice for dextran analysis. The suitability of the calibration thick juice (4.4) for dextran analysis must be checked prior to proceeding with the determination. This is done by measuring the absorbance of the reagent blank solution diluted to volume (4.12, 9.1.2.2), A(720)_TJ,RBl, against the absorbance of the calibration blank solution (4.13, 9.1.3.3) (without dextran solution added) diluted to volume, A(720)_TJ,CALBl. The corrected absorbance (A(720)_TJ,RBl – A(720)_TJ,CALBl) should not exceed the following values, which depend upon the optical cell (7.13) in use: 0.003 for a 2 cm cell or 0.008 for a 5 cm cell.

9.1.3.2 Preparation of calibration thick juice solutions for absorbance measurement. Prepare the calibration thick juice solutions using the calibration thick juice (4.4), TCA solution (6.4) and the standard dextran solution (6.2) using 100 mL volumetric flasks (7.2) and following the preparation details shown in Table 2. A set of calibration thick juice solutions is required for each calibration curve. The wet dextran content (β_{TJ,Dx,STD,wet}) listed in Table 2 needs to be corrected to obtain the dry dextran content (10.1.3).

 

Table 2: Calibration thick juice solutions

Flask No.	Calibration thick juice (4.3) (mL)	TCA solution (6.4) (mL)	Standard dextran solution (6.2) (mL)	Wet dextran content in calibration thick juice (mg/L) βTJ,Dx,STD,wet
1	50	10	0.0	0
2	50	10	1.25	50
3	50	10	2.5	100
4	50	10	5.0	200
5	50	10	7.5	300

 

After addition of the TCA solution, cool the flasks in a 20 °C in a cooling water bath (7.5) before making up to volume with de-ionised water. Stopper the flasks and mix the solutions thoroughly.
Pour each prepared calibration thick juice solution into a 150 mL beaker (7.2). Add approximately (6 to 8) g of acid-washed filter aid (6.8) to each beaker and mix well. Filter the mixture under low pressure (7.9) through a filter paper (7.10) fitted to a funnel connected to a vacuum flask (7.11). Apply vacuum and filter (10 to 15) mL solution. Disconnect the vacuum and use the filtrate to rinse the flask, then dispose of the filtrate. Reconnect the funnel and flask to the vacuum and filter the remaining solution.

9.1.3.3 Absorbance measurement of calibration thick juice solutions. Using a pipette (7.2), transfer an aliquot of 12.5 mL of a calibration thick juice solution filtrate (9.1.3.2) to each of two dry 25 mL volumetric flasks (7.2). Slowly add DAA (6.7) to one of the 25 mL flasks from a burette (7.2) up to the 25 mL mark while gently swirling the flask. Use a stopwatch (7.3) to ensure that the time taken to add the alcohol is 30 to 60 s. Stopper the flask and mix the contents by inverting gently three times. Start the stopwatch (7.3) immediately after the mixing step is complete.
Approximately 17 min after the completion of the mixing step, rinse one of the optical cells (7.13) three times with the solution and then fill the cell. Clean the optical faces of the cells with a tissue and check that the solutions in the cell are uniform (striations are absent) and place in the sample position of the instrument. This is a calibration thick juice solution.
Make up the second 25 mL flask to volume with water. Rinse the other optical cell (7.13) three times with the solution and then fill the cell. Clean the optical faces of the cell with a tissue and check that the solution in the cell is uniform (striations are absent), place this cell in the reference position of the instrument. This is the calibration thick juice blank solution.
At 20 min ± 10 seconds after completion of the mixing step, read and record the corrected absorbance A(720)_TJ,CALCorr), to 0.001 absorbance units.
Repeat the above steps for each calibration thick juice solution prepared in Table 2, using the calibration thick juice blank solution in the reference position for each calibration thick juice solution.

 

NOTE 4 – Alcohol must be added within 20 min of addition of dextran solution to the juice/TCA solution.
NOTE 5 – As the absorbance must be read at a precise time after the mixing step it is recommended that alcohol be added to the dextran standards at uniform time intervals (3 min or 4 min).
NOTE 6 – The filtrate of Flask No. 1 (Table 2, no dextran addition) is the calibration blank (4.13, 9.1.3.2) after diluting the filtrate with de-ionised water to volume and mixing.

 

9.1.3.4 Calculation of calibration curve for thick juice. Calculate the dry dextran content values for the calibration thick juice solutions (β_{TJ,Dx, STD,dry}) (10.1.3) and plot these against the corresponding corrected absorbance values (A(720)_TJ,CALCorr) values to produce a thick juice calibration curve. The calibration curve should be evaluated using standard laboratory protocols and also include the determination of the coefficient of determination R² (Note that the coefficient of determination R² should be greater than 0.9900).

9.2 Determination of dextran content in raw juice and thick juice samples. Raw juice samples (8.2) and thick juice samples (8.3) must be at room temperature prior to preparation of the sample solutions for determination of dextran content. Each sample should be prepared in duplicate.

9.2.1 Preparation and filtration of the raw juice and thick juice samples. Transfer 50 mL of the sample into a pair of 100 mL volumetric flasks (7.2). Add (10.0 ± 0.1) mL of TCA solution (6.4) from a pipette or dispenser (7.2) to each flask and make up to the mark using deionised water, stopper and mix well.
For each flask, pour the solution into a 150 mL beaker (7.2). Add approximately (6 to 8) g of acid-washed filter aid (6.8) and mix well. Filter the mixture under low pressure (7.9) through a filter paper (7.10) fitted to a funnel connected to a vacuum flask (7.11). Apply vacuum and filter (10 to 15) mL solution. Disconnect the vacuum and use the filtrate to rinse the flask, then dispose of the filtrate. Reconnect the funnel and flask to the vacuum and filter the remaining solution.

9.2.2 Preparation of the sample solution for absorbance measurement. Using a pipette (7.2), transfer 12.5 mL of the sample solution filtrate (9.2.1) to each of two clean, dry 25 mL volumetric flasks (7.2). Cool flasks to 20 °C in a cold-water bath (7.5).
Add DAA (6.7) slowly from a 50 mL burette (7. 2) to the 25 mL mark of one flask, gently swirling the flask. Use a stopwatch (7.3) to ensure that the time taken to add the alcohol is (30 to 60) s. Stopper the flask and mix the contents by inverting gently three times. Start the stopwatch (7.3) immediately after the mixing step is complete. To the other flask, add distilled water to the 25 mL mark, stopper, and mix. This is the sample blank solution (4.11).

 

NOTE 7 – Add alcohol within 20 min of addition of the TCA solution.
NOTE 8 – Avoid vigorous shaking of the flask as it may cause coagulation of the dextran haze.

 

9.2.3 Determination of the absorbance for the sample solution. Approximately 17 min after the completion of the mixing step in 9.2.2, rinse one of the optical cells (7.13) three times with the sample blank solution (4.11) and then fill the cell. In a similar way, rinse and fill the other optical cell with the sample solution (9.2.2). Clean the optical faces of the cells with a tissue and check that the solution in the cell is uniform (striations are absent) and place in the sample position of the instrument.
Rinse the other optical cell (7.13) three times with the sample blank solution (9.2.2) and then fill the cell. Clean the optical faces of the cell with a tissue and check that the solution in the cell is uniform (striations are absent), place this cell in the reference position of the instrument.
At 20 min ± 10 seconds after the completion of the mixing step read and record the corrected absorbance of the sample solution, A(720)_SA,Corr, to 0.001 absorbance units. Immediately after reading the absorbance, visually inspect the contents of the test solution cell to check for flocculation. If the haze has flocculated, repeat the analysis.
If the corrected absorbance of the sample solution, A(720)_SA,Corr, is higher than the upper limit of the calibration graph (9.1.2.4 or 9.1.3.4), repeat the determination by diluting the appropriate raw juice or thick juice sample with water (8.2 or 8.3).

 

10 Calculation and expression of results

10.1 Calculation of calibration curve and dextran content of samples.
10.1.1 Calculation of moisture content of dextran. The moisture content of the standard dextran (w_Dx,STD,W) (9.1.1) is determined by gravimetry (e.g. Method GS2-15 (2007)) or by Karl Fischer (e.g. Method GS4-12 (1998)). The calculation required for gravimetric determination of moisture content of the standard dextran is as follows:

\displaystyle {{w}_{{\text{Dx}\text{,STD}\text{,W}}}}\text{=100}\cdot \frac{{\left( {{{m}_{\text{2}}}-{{m}_{\text{3}}}} \right)}}{{\left( {{{m}_{\text{2}}}-{{m}_{\text{1}}}} \right)}}

wDx,STD,W	Moisture content of the standard dextran used to prepare the standard dextran solution (g/100 g)
m1	Mass of the dish (g)
m2	Mass of the dish + dextran, before drying (g)
m3	Mass of the dish + dextran, after drying (g)

 

Example:
Moisture content of wet standard dextran:

\displaystyle {{w}_{{\text{Dx}\text{,STD}\text{,W}}}}\text{=100}\cdot \frac{{\left( {\text{10}\text{.0064}-\text{10}\text{.0061}} \right)}}{{\left( {\text{10}\text{.0064}-\text{10}\text{.0009}} \right)}}

w_Dx,STD,W = 5.45 g/100 g

m₁ = 10.0009 g
m₂ = 10.0064 g
m₃ = 10.0061 g

 

10.1.2 Calculation of the wet mass of standard dextran required to prepare the standard dextran solutions. The moisture content of the standard dextran (w_Dx,STD,W) is required to determine the mass of wet dextran (m_Dx,STD,wet) to use in preparation of the standard dextran solution (6.2).

\displaystyle {{m}_{{\text{Dx}\text{,STD}\text{,wet}}}}\text{=}\frac{{\left( {{{m}_{{\text{Dx}\text{,STD}\text{,dry}}}}\cdot 100} \right)}}{{\left( {\text{100}-{{w}_{{\text{Dx}\text{,STD}\text{,W}}}}} \right)}}

mDx,STD,wet	Mass of wet standard dextran standard required for the standard dextran solution (g)
mDx,STD,dry	Mass of dry standard dextran required for the standard dextran solution (g)
mDx,STD,W	Moisture content of the standard dextran used in the standard dextran solution (g/100 g)

 

Example:
Mass of wet standard dextran required for an equivalent 0.4 g dried dextran:

\displaystyle {{m}_{{\text{Dx}\text{,STD}\text{,wet}}}}\text{=}\frac{{0.4\cdot 100}}{{100-5.45}}

m_Dx,STD,wet = 0.4231 g
m_Dx,STD,dry = 0.4 g
m_Dx,STD,W = 5.45 g/100 g (see example in 10.1.2)

 

10.1.3 Calculation of the dextran content in calibration solutions. Calculate the content of dry dextran in each flask for Tables 1 (β_{RJ,Dx,STD,dry}) and 2 (β_{TJ,Dx,STD,dry}) using content of wet dextran for the flask (β_{RJ,Dx,STD,wet} or β_{TJ,Dx,STD,wet}), the wet mass required to prepare the standard dextran (m_Dx,STD,wet) and the calculated dry mass (m_Dx,STD,dry). These dextran contents should then be used in the calibration curve (9.1.2.4 or 9.1.3.4).

\displaystyle {{\beta }_{{\text{RJ}\text{,Dx}\text{,STD}\text{,dry}}}}\text{=}{{\beta }_{{\text{RJ}\text{,Dx}\text{,STD}\text{,wet}}}}\cdot \frac{{{{m}_{{\text{Dx}\text{,STD}\text{,dry}}}}}}{{{{m}_{{\text{Dx}\text{,STD}\text{,wet}}}}}}

or

\displaystyle {{\beta }_{{\text{TJ}\text{,Dx}\text{,STD}\text{,dry}}}}\text{=}{{\beta }_{{\text{TJ}\text{,Dx}\text{,STD}\text{,wet}}}}\cdot \frac{{{{m}_{{\text{Dx}\text{,STD}\text{,dry}}}}}}{{{{m}_{{\text{Dx}\text{,STD}\text{,wet}}}}}}

β_{RJ,Dx,STD,dry}     Content of dry dextran in the calibration raw juice solution (mg/L)
β_{RJ,Dx,STD,wet}    Content of wet dextran in the calibration raw juice solution (mg/L)
β_{TJ,Dx,STD,dry}     Content of dry dextran in the calibration thick juice solution (mg/L)
β_{TJ,Dx,STD,wet}    Content of wet dextran in the calibration thick juice solution (mg/L)
m_Dx,STD,wet      Mass of wet dextran in the standard dextran solution (g)
m_Dx,STD,dry      Mass of dry dextran in the standard dextran solution (g)

 

Example:
Dry dextran mass in flask 5, for Table 1 (β_{RJ,Dx,STD,wet})

\displaystyle {{\beta }_{{\text{RJ}\text{,Dx}\text{,STD}\text{,dry}}}}=600\cdot \frac{{0.4017}}{{0.4231}}

β_{RJ,Dx,STD,dry} = 569.71 mg/L
β_{RJ,Dx,STD,wet} = 600 mg/L
m_Dx,STD,dry = 0.4017 g
m_Dx,STD,wet = 0.4231 g

 

10.1.4 Calculation of the dextran content of samples. The content of dextran in juice samples (β_RJ,Dx or β_TJ,Dx) is calculated directly from the relevant calibration curve (9.1.2.4 or 9.1.3.4), using corrected absorbance of the sample solution (A(720)_SA,Corr). This calculation may be done by using plot of the calibration curve or the equation for the line fitted to the data, following standard laboratory practice for the preparation and interpretation of calibration curves.

10.2 Expression of results. Express results as content of dextran in juice in mg/L as a whole number (no decimal place).

10.3 Test report. The test report shall include information on at least the following aspects of the test:

• the sample
• the ICUMSA® Method used (including its year of publication)
• the method used (if the standard includes several)
• the result(s), including a reference to the “Calculation” clause
• any deviations from the procedure
• any unusual features observed
• the date of the test.

 

11 Precision

11.1 Interlaboratory test. Interlaboratory tests were conducted for both frozen raw juice samples and frozen thick juice samples. The results for thick juice were reported at the 33^rd Session of ICUMSA^®, 2023 [2] and the results for raw juice were reported at the 34^th Session of ICUMSA^® in 2025 [3].

11.2 Repeatability and reproducibility. The absolute difference between two results obtained under repeatability (r) and reproducibility conditions (R) should not be greater than the values given in Table 3 and Table 4. These values were determined from the collaborative studies [2, 3], using an average value for r and R, for the data available.

 

Table 3: Repeatability and reproducibility for raw juice

	Content of dextran [mg/L]
	50	100	200	300	400	500
Repeatability r (mg/L)	37	30	24	25	10	43
Reproducibility R (mg/L)	53	125	69	195	135	76

 

Table 4: Repeatability and reproducibility for thick juice

	Content of dextran [mg/L]
	12.5	25	50	100	200
Repeatability r (mg/L)	6	14	15	16	22
Reproducibility R (mg/L)	9	14	17	44	68

 

12 Bibliography

1. Huet J.M. (2008): Referee’s Report on General Subject 8, Beet Sugar Processing, Proc. ICUMSA^® 26^th Session, Delray Beach, Verlag Dr. Albert Bartens KG, Berlin, 90–145
2. Emerstorfer F. (2023): Referee’s Report on General Subject 8, Beet Processing, Proc. ICUMSA^® 33^rd Session, Vienna, Verlag Dr. Albert Bartens KG, Berlin, 139–152
3. Emerstorfer F. (2025): Referee’s Report on General Subject 8, Beet Processing, Proc. ICUMSA^® 33^rd Session, Delhi, Verlag Dr. Albert Bartens KG, Berlin, 115–141

 

13 Historical

Method GS1-15 (2005) was modified and adapted to include process juices and presented at the 25^th Session of ICUMSA^® in 2006 Session. The Method, GS1/2/8-15 was renamed “The Determination of Dextran in Sugars and Juices by a Modified Alcohol Haze Method – Official for Raw Sugars – Accepted for Beet Sugar Processing” and published in 2007. The Method was revised and presented at the 26^th Session of ICUMSA^® in 2008, as Method GS8-19, “The Determination of Higher Molecular Mass Substances in Beet Raw Juice and Thick Juice by a Modified Alcohol Haze Method”. This Method was adopted with Tentative status and published in 2009. Minor corrections were made, and the Method was presented at the 30^th Session in 2016, with the recommendation for additional work to be done to improve and validate the Method. Further work was undertaken and presented at the 32^nd and 33^rd Sessions of ICUMSA^® in 2021 and 2023, with a focus on raw juice samples. The Method was rewritten in 2025 to align to the new ICUMSA^® Method format and include the validation and preparation data determined from collaborative studies performed.

 

Annex 1 – Collaborative Study data

Table A1 was reported in the 33^rd ICUMSA^® Proceedings [2] but it has been reproduced here, as it is relevant to the performance of the method. The sample material used in the study was standard thick juice that had been spiked with known contents of dextran.

 

Table A1: Performance data for dextran content of thick juice determined using Method GS8-19 (2026) (outliers removed)

Material No.	1	2	3	4	5
No. of laboratories in study, L	8	8	8	9	9
No. of samples analysed, k	16	16	16	18	18
Mean value value of dextran content, x̄ (mg/L)	21	27	42	75	167
Repeatability standard deviation, sr (mg/L)	2	5	5	6	8
Reproducibility standard deviation, sR (mg/L)	3	5	6	15	24
Relative repeatability standard deviation, RSDr (%)	10	19	12	8	5
Relative reproducibility standard deviation, RSDR (%)	15	18	14	21	14
Repeatability limit, r (mg/L)	6	14	15	16	22
Reproducibility limit, R (mg/L)	9	14	17	44	68

 

Annex 2 – Collaborative Study data

Table A2 was reported in the 34^th ICUMSA^® Proceedings [3] but it has been reproduced here, as it is relevant to the performance of the method. The sample material used in the study was standard raw juice that had been spiked with known contents of dextran.

 

Table A2: Performance data for dextran content of raw juice determined using Method GS8-19 (2026) (outliers removed)

Material No.	1	2	3	4	5	6
No. of laboratories in study, L	13	13	13	13	13	13
No. of samples analysed, k	20	20	20	22	22	20
Mean value value of dextran content, x̄ (mg/L)	68	100	219	309	394	494
Repeatability standard deviation, sr (mg/L)	11	9	7	10	7	13
Reproducibility standard deviation, sR (mg/L)	28	14	25	61	56	52
Relative repeatability standard deviation, RSDr (%)	16	9	3	3	2	3
Relative reproducibility standard deviation, RSDR (%)	41	14	11	20	14	10
Repeatability limit, r (mg/L)	30	27	20	28	20	37
Reproducibility limit, R (mg/L)	79	39	69	171	159	146