This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles

for the
Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

Designation: E1721 − 01 (Reapproved 2020)

Standard Test Method for

Determination of Acid-Insoluble Residue in Biomass1
This standard is issued under the fixed designation E1721; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.

INTRODUCTION

Biomass is composed largely of the following: cellulose, a polymer of glucose; hemicellulose, a

complex polymer, the main chain of which consists of xylans or glucomannans; and lignin, a complex
phenolic polymer. The lignin is mostly insoluble in mineral acids, unlike the other cell wall
components of biomass. For this reason, lignin can be analyzed gravimetrically after hydrolyzing the
cellulose and hemicellulose fractions with sulfuric acid.

1. Scope 1.6 This international standard was developed in accor-

1.1 This test method covers determination of the acid- dance with internationally recognized principles on standard-
insoluble residue of hard and soft woods, herbaceous materials ization established in the Decision on Principles for the
(such as switchgrass and sericea), agricultural residues (such as Development of International Standards, Guides and Recom-
corn stover, wheat straw, and bagasse), wastepaper (such as mendations issued by the World Trade Organization Technical
Barriers to Trade (TBT) Committee.
iTeh Standards
office waste, boxboard, and newsprint), acid and alkaline
pretreated biomass, and the solid fraction of fermentation 2. Referenced Documents
residues. All results are reported relative to the 105 °C oven-
dried weight of the sample. (https://standards.iteh.ai)
1.2 The residue collected contains the acid-insoluble lignin
2.1 ASTM Standards:
E1690 Test Method for Determination of Ethanol Extrac-
2

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tives in Biomass
and any condensed proteins from the original sample. An E1756 Test Method for Determination of Total Solids in
independent nitrogen analysis would be required to determine Biomass
the acid-insoluble lignin content separate from the condensed E1757 Practice for Preparation of Biomass for Composi-
protein fraction and is outside the scope of thisASTM E1721-01(2020)
test method. tional Analysis
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1.3 A portion of the lignin in some biomass samples will
3. Terminology
remain soluble during this procedure. The total lignin in a
biomass sample includes both acid-soluble lignin and lignin in 3.1 Definitions:
the acid insoluble residue. 3.1.1 acid-insoluble residue—the solid residue, corrected
1.4 The values stated in SI units are to be regarded as for acid-insoluble ash, retained on a medium-porosity filter
standard. No other units of measurement are included in this crucible after the primary 72 % and secondary 4 % H2SO4
standard. hydrolysis described in this test method. This material is
primarily acid-insoluble lignin and any condensed proteins.
1.5 This standard does not purport to address all of the
safety concerns, if any, associated with its use. It is the 3.1.2 prepared biomass—material that has been treated in
responsibility of the user of this standard to establish appro- accordance with Practice E1757 in order to raise the total solids
priate safety, health, and environmental practices and deter- content above 85 %, based on an oven-dried solids weight.
mine the applicability of regulatory limitations prior to use.
4. Significance and Use
Specific hazards statements are given in Section 8 and Note 2
and Note 4. 4.1 The acid-insoluble residue content is used in conjunc-
tion with other assays to determine the total composition of
biomass samples.
1
This test method is under the jurisdiction of ASTM Committee E48 on
Bioenergy and Industrial Chemicals from Biomass and is the direct responsibility of
2
Subcommittee E48.05 on Biomass Conversion. For referenced ASTM standards, visit the ASTM website, www.astm.org, or
Current edition approved July 1, 2020. Published August 2020. Originally contact ASTM Customer Service at [email protected]. For Annual Book of ASTM
approved in 1995. Last previous edition approved in 2015 as E1721 – 01(2015). Standards volume information, refer to the standard’s Document Summary page on
DOI: 10.1520/E1721-01R20. the ASTM website.

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1
 E1721 − 01 (2020)
5. Interferences 9.3 The samples for total solids determination should be
5.1 The results of acid-insoluble residue analysis are af- weighed out at the same time as those for acid-insoluble
fected by the incomplete hydrolysis of biomass. The results residue determination. If this is performed later, it can intro-
will be biased high unless the sample is hydrolyzed completely. duce an error in the calculation because ground biomass can
Take care to mix the acid/biomass slurry thoroughly during the gain or lose moisture rapidly when exposed to the atmosphere.
concentrated acid hydrolysis.
10. Procedure
5.2 The results of acid-insoluble residue analysis are af-
fected by the timing of the acid digestion steps. The insoluble 10.1 Label the crucibles needed for analysis individually,
residue will dissolve slowly into solution in an irreproducible and ignite them at 575 6 25 °C to achieve a constant weight of
fashion. The timing within this test method must be followed 60.3 mg. Store the ignited crucibles in a desiccator until
closely. needed.
NOTE 1—In order to determine the absolute amounts of acid-insoluble
6. Apparatus residue and acid-insoluble ash, for quality control purposes, it is useful to
6.1 Analytical Balance, readable to 0.1 mg. weigh and record the ignited crucible to the nearest 0.1 mg.

6.2 Convection Oven, with a temperature control of 105 6 10.2 Weigh a 0.3 6 0.01-g sample to the nearest 0.1 mg, and
3 °C. place it in a test tube. Record the initial weight as W1.

6.3 Muffle Furnace—An electric furnace is recommended NOTE 2—Warning: 72 % sulfuric acid is very corrosive and should be
handled only by trained personnel.
for igniting the sample. The furnace should be fitted with an
indicating pyrometer or thermocouple so that the required 10.3 Add 3.00 6 0.01 mL (4.92 6 0.01 g) of 72 % H2SO4,
temperature of 575 6 25 °C can be maintained. and stir for 1 min or until mixed thoroughly.
6.4 Autoclave, capable of maintaining 121 6 3 °C. 10.4 Place the test tube in the water bath controlled to 30 6
1 °C, and hydrolyze for 2 h.
6.5 Water Bath, set at 30 6 1 °C.
NOTE 3—The hydrolysis time may be reduced to 1 h if the dried sample

iTeh Standards
6.6 Desiccator, using anhydrous calcium sulfate. has been milled and sieved to pass through a 20-mesh sieve and be
retained on a 80-mesh sieve.
7. Reagents and Materials

(https://standards.iteh.ai)
10.5 Stir the sample every 15 min to ensure complete
7.1 Chemicals: mixing and wetting.
7.1.1 72 % H SO , specific gravity 1.6389 6 0.0012 at
2 4

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15.6 °C ⁄15.6 °C or 12.00 6 0.02 M. 10.6 Transfer the hydrolyzate to a glass bottle, and dilute to
7.1.2 Water, 18 MΩ deionized. a 4 % acid concentration by adding 84.00 6 0.04 mL water or
by bringing the combined weight of sample, acid, and water up
7.2 Materials: to 89.22 6 0.04 g. Be careful to transfer all of the residual
7.2.1 Glass Serum Bottles, 125 mL, crimp ASTM
top style,E1721-01(2020)
with solids along with the hydrolysis liquor.
rubber stoppers and aluminum seals to fit.
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7.2.2 Glass Filtering Crucible, 50 mL, medium porosity, 10.7 Stopper each of the bottles, and crimp the aluminum
with a nominal maximum pore size of 10 µm. seals into place.
7.2.3 Vacuum Adapter for Crucibles. 10.8 Set the autoclave to a liquid vent cycle to prevent loss
of sample from the bottle in the event of a loose crimp seal.
8. Hazards Autoclave the samples in their sealed bottles for 1 h at 121 6
8.1 Handle the sulfuric acid carefully. 3 °C.
8.2 Use caution when handling glass bottles after the NOTE 4—Warning: Handle sealed bottles with caution after the
autoclave step since they may become pressurized. autoclave step since they may become pressurized.
10.9 After completion of the autoclave cycle, allow the
9. Sampling, Test Specimens, and Test Units
samples to cool for approximately 20 min at room temperature
9.1 Test specimens suitable for analysis with this procedure before removing the seals and stoppers.
are as follows:
10.10 Vacuum filter the hydrolysis solution through a pre-
9.1.1 Prepared biomass samples that have been treated in
viously ignited filtering crucible.
accordance with Practice E1757.
9.1.2 Extractives-free material prepared in accordance with 10.11 If a carbohydrate analysis or acid-soluble lignin
Test Method E1690. analysis, or both, is desired, decant 15 to 25 mL of filtrate into
a resealable container. If the aliquot is not used immediately for
9.2 The test specimen shall consist of approximately 0.3 g
further analysis, store it in a refrigerator at 4 °C.
of sample obtained in such a manner to ensure that it is
representative of the entire lot of material being tested. NOTE 5—Acid-soluble lignin should be analyzed within 24 h and
Prepared biomass is used in this test, but the weight of the preferably within 6 h of hydrolysis.
material must be corrected to 105 °C dry weight by using the 10.12 Use hot water to wash any particles clinging to the
percent total solids value determined in accordance with Test glass bottle into the crucible and to wash the filtered residue
Method E1756, prior to calculating the acid-insoluble residue. free of acid using vacuum filtration.