Government of Canada Gouvernement du Canada

Laboratory Procedure MFLP-66

June 2006

HEALTH PRODUCTS AND FOOD BRANCH

OTTAWA

DETERMINATION OF WATER ACTIVITY USING THE DECAGON AQUALAB CX-2 AND SERIES 3

Patti Wilson Irene Iugovaz

Canadian Food Inspection Agency Food Directorate
Atlantic Region Health Canada-Quebec Region
P.O. Box 1060 1001 St-Laurent west
Dartmouth, Nova Scotia Longueuil, Quebec
B2Y 3Z7 J4K 1C7
wilsonpa@inspection.gc.ca irene_iugovaz@hc-sc.gc.ca

1. APPLICATION

This method is applicable to the measurement of water activity in foods and food ingredients to
determine compliance with the requirements of Sections 4 and 7 of the Food and Drugs Act,
Section B.27.001 of the regulations of the Food and Drugs Act and any other federal regulation.

This revised method replaces MFLP-66, dated October 2001.

2. PRINCIPLE

Water activity plays an important role in the preservation of food and the control of microbial
growth, especially pathogens. Decagon AquaLab models CX-2 and Series 3 use the chilled-mirror
dewpoint technique to measure the water activity of a product.

3. DEFINITION OF TERMS

See Appendix A of Volume 3, items 1,2,3,4.

Water activity (aw) : is the ratio of the water vapour pressure in any food system to the water
vapour pressure of pure water at the same temperature.

Equilibration: When the aw values of any two readings are less than .001 apart, the instrument
has reached equilibrium and the measurement process stops.

Published on the Food Directorate’s (Health Canada's) website at :

http://www.hc-sc.gc.ca/fn-an/res-rech/analy-meth/microbio/index_e.html
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Humidity standard (slush) : is a saturated solution of a salt of a known % relative humidity at a

designated temperature.

Commercial humidity standard: is an unsaturated salt solution of a specific molality representing a

know Aw.

4. COLLECTION OF SAMPLES

See Appendix B of Volume 3.

Keep samples sealed to prevent loss or gain of moisture during transport.

5. APPARATUS AND MATERIALS

5.1 Apparatus

5.1.1 AquaLab Models CX-2 and Series 3 are available from Decagon Devices Inc., P.O.Box
835, 950 NE Nelson Court, Pullman, Washington USA 99163. The Canadian distributor is
Meyer Service and Supply Ltd., 119 Warner Drive RR1 Long Sault, Ont. K0C 1P0.

5.2 Materials

5.2.1 Reference salts: ACS reagent grade, fine crystal, available from scientific supply houses.
See Table 1 for a list of the reference salts which may be used.

5.2.2 Reference humidity standards, available from Decagon distributors.

5.2.3 Disposable sample cups from Decagon or equivalent.

5.2.4 Test containers - 120 mL or 240mL Mason-type glass jar with screw-caps and gaskets or
equilavent

5.2.5 AquaLab cleaning kit (optional)

6. PROCEDURE

Each sample unit may be analyzed individually or the analytical units may be combined. Carry out
the test in accordance with the following instructions:

6.1 Handling of Samples Units

6.1.1 In the laboratory prior to analysis, except for shelf-stable foods, keep sample units
refrigerated or frozen, depending on the nature of the product. Thaw frozen samples in a
refrigerator, or under time and temperature conditions which prevent microbial growth or
death. Maintain package integrity until the point of measurement to prevent water loss,
which may alter the water activity.

6.1.2 Analyze sample units as soon as possible after their receipt in the laboratory.

6.2 Reference Humidity Standards

6.2.1 Certified humidity standards are preferred and may be purchased.

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6.2.2 To prepare in-house reference humidity standards, place selected reference salt in a
labeled test container to a depth of about 4 cm. Add distilled water in gradual increments
with constant stirring until no more water is absorbed and there is some free liquid on top
of the slush. At this point they are ready to use and will keep indefinitely, if contained in
manner to prevent substantial evaporation losses.

6.2.3 To use the in-house reference humidity standard for the AquaLab, use the liquid portion
of the mixture only.

6.2.4 Replace either of the purchased or in-house humidity standards if they become
contaminated or expired.

QUALITY CONTROL

The aw reading obtained by measuring the Decagon Devices humidity standards must be
within ±0.003 aw units of the stated value of the standard.

To determine the accuracy of uncertified humidity standards, use a method that is independent
of the calibration methodology in order to have a meaningful check on the standard material.
Options include:

1. Send blind samples to another facility to be checked on the same type of aw apparatus.

2. Test blind samples on a type of apparatus different than the one than you are using.

3. Send blind samples to an external calibration laboratory to certify the standards.

Certified humidity standards are best, if available.

6.3 Calibration

6.3.1 Calibration should be performed daily before sample measurements.

6.3.2 Before calibrating, locate the AquaLab in an area with a stable ambient temperature and
have the reference humidity standards at ambient temperature.

6.3.3 Always, use new sample cups for the reference humidity standards. For routine analysis
of food, use 6M NaCl, which has aw = 0.760 or choose a humidity standard with a value
higher than 6M NaCl as long as it is of a lower value than the anticipated aw of the sample
being measured. Only samples within the aw range of this humidity standard and the aw of
distilled water can be reported as a numerical value rather than > the upper point or < the
lower point.

6.3.4 Measure in duplicate the aw of 6M NaCl or of another chosen humidity standard as

described in 6.3.3, comparing the readings and temperatures with theoretical values, as
shown in table 1.

6.3.5 If readings of the humidity standard are within the stated ranges, take duplicate readings
of distilled water. If the readings for distilled water are within 1.000 ± 0.003, the calibration
is complete.
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If readings of distilled water are not correct, clean the sensor as described in section 6.6
and repeat the calibration from 6.3.4.

Hint : Always start calibration / verification with the lowest aw slush.

Hint : If the aw value of water is not correct (1.000), use steam distilled water; that will always read
1.000.

6.3.6 If readings of the humidity standard are outside the stated ranges, even after re-checking,
adjust the meter reading to its correct value. For AquaLab model CX-2, carefully adjust
the potentiometer as needed with a small screwdriver to the correct aw value while the
meter is still beeping. For Aqualab Series 3, move through the linear offset menu as
directed and, using the function keys, correct the aw value.

Repeat the calibration from 6.3.4.

6.3.7 To check the linearity of the calibration for each sensor, measure the aw of 4 humidity
standards: the upper and lower humidity standards plus 2 other humidity standards which
have an aw between the upper and lower humidity standards. When these upper and
lower humidity standards are adjusted as described above, measure the other 2 humidity
standards in duplicate. The verification of the linearity should be performed periodically.

6.3.8 If the humidity standards can not be adjusted to be linear, refer the Troubleshooting
section of the AquaLab Operator’s Manual. If non-linearity continues, contact Decagon.

6.4 Sample Preparation

6.4.1 Samples should be left intact if possible to ensure that the aw is not altered. If necessary,
cut the sample material to fit the sampling cup. Fill the plastic sample cups half full in a
such manner to cover most of the bottom of the cup. Take care not to use your hands to
directly handle the sample material. If needed, clean the exterior surfaces (sides and rim)
of the cup to avoid contamination of the reading chamber.

6.4.2 Samples that cannot be analysed immediately upon receipt may be stored. Once
prepared, the sample unit should be analyzed the same day. If not, tape the cup/lid
junction of the the filled plastic sample cups and place them in sealed plastic bags.
Remove excess air and store them at an appropriate temperature.

For long term storage, glass jars or mylar bags should be used. Care should be taken in
minimizing headspace to reduce water vapor and condensation effects. The containers
should be stored in a freezer.

Prewarm refrigerated and frozen samples to ambient temperature to reduce the

equilibration time and to give a more accurate reading. Care should be taken to avoid loss
of water from the sample.

6.4.3 Special preparation is required for materials, like coated foods, dried foods, highly viscous
water-in-oil (butter), high fat and volatiles. Refer to the chapter on Sample Preparation in
the AquaLab Operator’s Manual.
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6.4.4 For products in oil, drain the product through sieves (or similar devices) with appropriate
mesh size to retain solid portion of the product. Let drain sufficiently to remove as much
oil as possible; if oil is still visible, use an absorbent paper towel and let it absorb the
excess oil. Do not Blot. Measure aw on the solid portion of the sample.

6.4.5 For products such as butter or other emulsion containing water in oil (pesto or
equivalent), the equilibration times are dramaticaly prolonged due to the presence of
lipids acting as a barrier to vapor diffusion. Sample should be run in a chamber that has a
lower aw than the sample or that has been dried with desiccant before measurement. It
may need to take 3 readings with this type of product to be in the ± 0.003 range.

6.5 Sample Measurement

6.5.1 Turn the sample drawer knob to the “Open/Load” position and open the sample drawer.
Place the prepared sample cup in the drawer and slide the drawer closed being careful
not to spill or splash liquid samples.

6.5.2 Turn the knob to the “Read” position to seal the sample cup with the chamber and start
the read cycle. When the instrument begins to beep and/ or the decimal points or the
LCD flash, the reading is complete. The display will show a final aw and temperature
measurement.

6.5.3 Measure the aw of each sample in duplicate by taking 2 reading of the same sample cup. [
to replace: Each sample should be analysed (aw measured) in duplicate].

6.5.4 For cautions, tips and display messages, refer to the chapter on “Taking
a Reading” in the AquaLab Operator’s Manual.

6.5.5 Record sample aw reading and the temperature on the appropriate lab sheet.

Hint: To minimise reading time, always start the readings with the dryer food and finish with the most
humid.

Hint: Check stability of calibration by measuring the aw of distilled water after each food type and/or at
the end of the day.

6.5.6 Report sample aw values using all three decimal places. Determine the mean of multiple
readings using three decimal places, then report the final mean value rounded to two
decimal places.

Note that, only samples within the aw range of the tested humidity standard and the aw of
distilled water can be reported as a numerical value rather than > the upper point or < the
lower point.

6.6 Cleaning and Maintenance

6.6.1 For procedures to clean the sensor chamber and fan filter, refer to the chapter on
“Cleaning and Maintenance” in the AquaLab Operator’s Manual.

6.6.2 Recheck the calibration after cleaning as described in section 6.3.

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7. References

7.1 AquaLab model CX-2, Operator’s Manual. Decagon Devices Inc.,Pullman,

Washington USA 99163.

7.2 AquaLab model Series 3, Operator’s Manual. Decagon Devices Inc.,Pullman, Washington
USA 99163.

7.3 Decagon Device inc, 2004, Sample preparation , Water Activity News. p.1,6.

7.4 Fontana, Anthony J., Decagon Device Inc, Personnal communication.

7.5 Greenspan, L. 1977. Humidity fixed points of binary saturated aqueous solutions. Journal
of Research of the National Bureau of Standards- A physics and Chemistry 81A:89-96.

7.6 Measurement of water activity (Aw) and acidity , 1992. Compendium of methods
for the microbiological examination of foods. American Public Health
Association,3rd Edition.135-154.

7.7 Robinson, R.A. and R.H. Stokes. 1965 Electrolyte solutions: the Measurement and
Interpretation of Conductance, Chemical Potential, and Diffusion in Solution of Simple
Electrolytes. London. Butterworth.

7.8 Stoloff, L. 1978. Calibration of Water Activity Measuring Instruments and Devices:
Collaborative Study. JAOAC. 61:1166-1178.

7.9 Stroup W.H. and Peeler J. T. 1987. Evaluation of Precision Estimates for Fiber-
Dimensional and Electrical Hygrometers for Water Activity Determinations. JAOAC.
70:955-957.

7.10 Troller J.A. 1977. Statistical Analysis of aw Measurements Obtained with the Sina Scope.
Journal of Food Science. 42:86-90.

7.11 Water Activity of Canned Vegetables. 1990. Official Methods of Analysis - AOAC - 15th
Edition. 987-988.
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TABLE I. Water Activity of Reference Salt and Slushes. (7.1,7.2,7.3, 7.6, 7.7, 7.11)

Salt aw at 20°C aw at 25°C

LiCl 0.113 ± 0.003 0.113 ± 0.003

13.41M LiCl 0.245 ± 0.003 0.250 ± 0.003

MgCl2 0.331 ± 0.002 0.328 ± 0.002

K2CO3 0.432 ± 0.003 0.432 ± 0.004

8.57M LiCl 0.496 ± 0.003 0.500 ± 0.003

Mg(NO3)2 0.544 ± 0.002 0.529 ± 0.002

NaBr 0.591 ± 0.004 0.576 ± 0.004

6.0M NaCl 0.760 ± 0.003 0.760 ± 0.003

KBr 0.817 ± 0.002 0.809 ± 0.002

KCl 0.851 ± 0.003 0.843 ± 0.003

KNO3 0.946 ± 0.007 0.936 ± 0.006

K2SO4 0.976 ± 0.005 0.973 ± 0.005

0.5M KCl 0.984 ± 0.003 0.984 ± 0.003

Distilled H2O 1.000 1.000

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MFLP-66 DETERMINATION OF WATER ACTIVITY USING DECAGON AQUALAB

Calibration Standard and samples at room

temperature

Step 6.3 Calibration using:

1. A reference humidity standard selected

to encompass the anticipated aw value of the
sample (usually 0.760)

2. Distilled water aw= 1.000±0.003

Duplicate readings of the reference humidity

standard and duplicate readings of water are
within acceptable ranges defined in
manufacturer’s instructions.

Calibration OK Calibration not

Proceed with sample
acceptable

Preparation of sample step 6.4 or Clean cell as in 6.6

as per Operator’s manual

Sample measurement in duplicate step 6.5

Duplicate: Duplicate readings of the same

Reporting : Report sample aw using all 3 decimal places.

Determination of mean of multiple readings using all 3 decimal places.

- Rounding to 2 decimal places for the final mean value or final result.
- aw of sample that are comprised within the aw range of the humidity standard used and the
aw of the water are reported as a numerical value. Values outside this range should be