Water is the most common reagent used in the laboratory, and while water quality can often be overlooked, the grade of water being used in an application is critical. Minute traces of salts or biological contaminants can result in unfortunate consequences when culturing cells or performing analytical measurements of biological macromolecules.

1. Laboratory Reagent
Water
By
Vinay Kumar Shukla

2. Introduction
Water’s ability to dissolve compounds, along with its polarity,
bonding, melting, boiling and freezing points, heat absorption, and
vaporization characteristics arguably make it the most versatile
substance
Water is the most common reagent used in the laboratory, and while
water quality can often be overlooked, the grade of water being
used in an application is critical. Minute traces of salts or biological
contaminants can result in unfortunate consequences when culturing
cells or performing analytical measurements of biological
macromolecules.
Reagent grade water (RGW) is water that is suitable for use in a
specified procedure such that it does not interfere with the
specificity, accuracy, and precision of the procedure. Process
definitions alone (e.g. “RO”, “DI”, “Distilled”) do not in themselves
adequately define required water quality

3. The reagent grade water shall be of the following three
grades depending on their extent of purity:
Type- 1
Type- 2
Type- 3
Type of Reagent water Grade

4. For use in test methods requiring minimum interference and
maximum precision and accuracy such as trace analysis. Reagent
water may be prepared by distillation of feed water having
maximum conductivity of 20 ymhos/cm at 25°C followed by
polishing with mixed bed deionizers and passage through 0.45 pm
membrane filter.
Type 1 laboratory grade water is required for critical laboratory
applications such as HPLC mobile phase preparation, blanks and
sample dilution in GC, HPLC, AA, ICP-MS and other advanced
analytical techniques; preparation of buffers and culture media for
mammalian cell culture and IVF; production of reagents for
molecular biology applications (DNA sequencing, PCR); and
preparation of solutions for electrophoresis and blotting.
Grade 1

5. For use in the general laboratory analytical procedures
where freedom from organic impurities is of more
significance. It is not applicable for biological or medical
analysis and organic trace analysis.
Type 2 laboratory grade water is used in general
laboratory applications such as buffers, pH solutions and
microbiological culture media preparation; as feed to
Type 1 water systems, clinical analyzers, cell culture
incubators and weathering test chambers; and for
preparation of reagents for chemical analysis or
synthesis.
Grade 2

6. For use in washing of glassware's, preliminary rinsing of
glassware and feed water for production of higher grade
waters or where large quantity of water is required of low
purity for make up of synthetic test solutions.
Type 3 water is the lowest laboratory water grade,
recommended for glassware rinsing, heating baths and
filling autoclaves, or to feed Type 1 lab water systems.
Grade 3

7. Contamination of water during storage may arise mainly
from dissolution of suitable constituents from glass or
plastic containers or absorption of atmospheric carbon
dioxide and or any other impurities present in the laboratory
atmosphere.
For this reason, the storage of grade 1 and grade 2 water is
not recommended. It is desirable to prepare, as required for
immediate use.
NOTE - In case it is required to store grade 1 and grade 2 water it can
be done with proper storage systems having carbon dioxide breather
and with seal arrangement.
Storage of Reagent Water

8. However, grade 2 water may be prepared in reasonable
quantity and stored in suitable, inert, clear, tight full
containers which have been thoroughly pre-washed with
water of same grade.
The storage of grade 3 water possess little problem, and
should be stored in containers that do not affect the quality
for the desired purpose. Let the containers and storage
conditions should be the same as those used for the storage
of grade 2 water.
NOTE - It is recommended that storage container be
reserved exclusively for the storage of grade 3 water.
Storage of Reagent Water

9. Parameters Grade-1 Grade-2 Grade-3
1. Specific Conductivity 0.1 1.0 5.0
umhos/cm at 25oC, Max
2. pH, at 25oC NS NS 5.0-8.0
3. Total solids or non volatile NS 1.0 2.0
residue at 105oC, mg/l Max
4. Silica (as SiO2), mg/l, Max 0.01 0.1 1.0
5. Colour retention of KMnO4,
at 24 x 2oC Minutes 60 10 10
Note: 1. The requirement for specific conductivity for grade 1 and grade 2 apply to
freshly prepared water. During storage it is possible for contaminants like atmospheric
carbon dioxide and alkalies from glass containers. to, be dissolved leading to changes
in conductivity.
2. Because of the difficulties associated with measurement of the pH value of high
purity water, and ‘the doubt- ful significance of the value obtained, limits for the pH of
grade 1 and grade 2 water have not been specified.
3. The limit for oxidizable matter and residue after evaporation for grade 1 water is not
specified because of the difficulty of testing for compliance at this level of purity, the
quality of grade 1 water is, however, assured, by compliance with the other
requirements and by its method of preparation.
Reagent Water Specifications

10. Parameters Value Unit
1. Bacteria <10 CFU/ml
2. Resistivity >10 MΩ.cm
3. TOC <500 PPB
4. Particles 0.2um filtration or better
Clinical Laboratory Reagent Water (CLRW) Specification

11. The containers shall bear legibly and indelibly the following
information:
a) Name of the material;
b) Indication of source of manufacture;
c) Volume of the material in litres;
d) Date of manufacture; and
e) Batch number.
The material shall be packed in clean glass, polyethylene or other
suitable plastic container which do not affect the quality of water.
The containers shall be securely closed.
Packing and Marking

12. It is more expensive to produce Type 1 water than Type 2
or Type 3 water but often a lab will chose a higher grade of
water than they need because they feel there is less chance
of compromising their experiments.
It is to the labs benefit both economically and technically to
determine the grade of water required most frequently
before selecting the water treatment or point of use system.
Where and when a higher grade water is required, a smaller
point of use system can be installed for those specific
applications.
For laboratory Use

13. Thanks