oil extraction and analysis phần 2 pps

Chapter 2
Soxtec: Its Principles and Applications
Shirley Anderson
Foss North America, Eden Prairie, MN 55344
Abstract
The classical Soxhlet method provides the fundamental basis for a modern-day sol-
vent extraction system, the Soxtec
TM
. Using the Randall modification, sometimes
called the submersion method, the Soxtec provides a faster approach to solvent
extraction for the gravimetric quantitation of fat and oil. Typically, the Soxtec meth-
ods require only 20–25% of the time required for traditional Soxhlet extraction.
Sample preparation, general extraction procedures, method considerations, and opti-
mization are addressed. By definition, the procedure to determine “crude fat” is an
empirical method in which the result is determined by the conditions of the procedure.
Several aspects of the extraction process, such as solvent type, time, and temperature,
are explored. Several standardized Soxtec methods are discussed, including the
recently approved AOAC method for determining crude fat in feeds, cereal grains,
and forages. Many Soxtec applications are routinely used in food, feed, industrial, and
environmental laboratories for the measurement of fats, oils, semivolatiles, and other
solvent “extractables.” For the determination of crude fat, descriptions are given for
various sample pretreatment and extraction procedures. Practical guidelines for han-
dling challenging samples as well as general suggestions are presented.
History
The foundation of today’s automated solvent extraction systems can be traced to
1879 to a German chemist, Franz Von Soxhlet. He devised a liquid/solid extrac-
tion apparatus in which a sample is placed in a cellulose thimble and stationed
over boiling solvent. Condensed solvent would then drip into the sample, solubiliz-
ing extractable material and then siphon back into the boiling solvent, where this
cycle would then repeat. After several cycles over many hours, the apparatus is
disassembled and the solvent, now containing extract (fat), is evaporated off, leav-

ing the residue for further analysis. The Soxhlet procedure remains the most
exhaustive extraction technique, and today it is still widely used.
Over the years, there had been some improvements to the basic technique but
the procedure remained long, tedious, and prone to variability. In the early 1970s,
Edward Randall (1) developed an accelerated extraction technique that cut the
extraction time to as little as 30 min. In the Randall method, the sample is lowered
Copyright © 2004 AOCS Press
and totally immersed in the boiling solvent. The simple principle is that the materi-
al to be extracted, in this case, fats and waxes, is more soluble in hot solvent than
in cold or room temperature solvent. His procedure included this new boiling step
followed by a rinsing step to flush residual extract from the sample (Fig. 2.1).
Demonstrating excellent agreement with Soxhlet and improved precision, the
Randall method has become the basis of many automated extraction systems.
In 1975, Tecator AB of Höganäs Sweden acquired the rights to what had
become known as the “Randall modification” of the Soxhlet method. This was first
commercialized as the RaFaTec and later became the Soxtec systems of today.
Figure 2.2 is a photograph of the Soxtec
TM
Avanti, from Foss-Tecator, an automat-
ed Soxhlet extraction system using the Randall submersion technique.
Procedural Overview of Soxtec, Automated Soxhlet
for Crude Fat
Automated extraction methods using the Soxtec have gained widespread accep-
tance and have a number of regulatory agency approvals worldwide. The Soxtec
method has been used on literally hundreds of different sample types for many
extracts. For our purposes, this discussion will be limited mainly to crude fat
extraction. Crude fat by solvent extraction is classified as an empirical method (2).
This means that the final result can be arrived at only according to the terms or
variables of the method. It therefore becomes critical that all aspects of the proce-
dure be followed strictly.

Sample Preparation. Proper handling of the sample and attention to detail are
extremely important parts of the analytical process. An improperly or sloppily pre-
FIG. 2.1. Original Soxhlet (left)
and Randall Extraction Apparatus.
(a) Condenser (b) sample thimble
(c) solvent flask (d) siphon tube (e)
solvent vapor tube (f) thimble
positioning mechanism (g) heater
(not shown on the Soxhlet). In the
original Randall method, the thim-
ble is positioned by use of the
slide rod (f). Lowering the thimble
(b) into the boiling solvent for the
boiling step, then raising it out of
the solvent for the rinsing step. In
both stages, condensed solvent is
flowing continuously through the
sample and thimble back into the
boiling solvent.
Copyright © 2004 AOCS Press
pared sample can invalidate even the most carefully performed extraction proce-
dure. Depending on the type and nature of the sample, the sample preparation may
incorporate different procedures. For grinding and weighing, the sample should be
homogenous and finely ground, usually to pass through a 1-mm sieve (~18 mesh).
Particular attention should be paid to the type of grinding mill used. The mill or
milling process should not contribute to any loss of moisture or fat from the sample.
Samples should be weighed using a calibrated 4-place analytical balance and
in most cases, can be weighed directly into the cellulose thimble. The weight of the
sample is dependent on its approximate fat content. Table 2.1 can be used a guide-
line. Because only a few grams of sample are normally used for the analysis, it is

critical that this small sample be representative of the larger sample lot.
Pretreatment
Drying. Most samples should be predried to optimize the fat extraction. Water in
the sample can decrease the efficiency of the solvent extraction, resulting in low fat
recoveries. Conversely, water-soluble components in the samples such as urea, car-
bohydrates, salts, and glycerol can be extracted with fat yielding falsely high
recoveries.
Samples are weighed into the extraction thimbles and are then typically dried
at 102 ± 2°C for 1–2 h (3–5,7). Because the samples are weighed before drying,
FIG. 2.2. The Soxtec
TM
Avanti 2050 automated extraction system.
Copyright © 2004 AOCS Press
the results are on an as-is basis. Results expressed on a dry matter basis must be
calculated from a separate moisture determination. For very moist samples, sand
may be mixed with the sample before drying. This prevents the sample from
becoming caked during drying and improves the solvent flow for optimal extrac-
tion (see below: Crude fat in meat and meat products).
Hydrolysis. Samples that have been processed, cooked, or extruded often have fat
that is bound to proteins, carbohydrates, and/or minerals, making it unavailable for
solubilization. Acid hydrolysis, in which a sample is boiled with hydrochloric acid,
breaks these bonds, allowing the fat to be solvent extracted (see below: Total fat).
Water Rinse. Samples that contain a large amount of water-soluble components
may exhibit poor solvent extraction efficiency. A preextraction with water, fol-
lowed by a thorough drying step can be used to obtain an acceptable recovery by
removing these water-soluble components. The procedure specifies washing the
weighed sample with 5 aliquots of 20 mL of deionized water. The sample is dried
and the extraction procedure is carried out as usual. (3,4)
Solvent Extraction
Once the samples have been properly prepared and pretreated, they can now be

placed in the Soxtec for fat extraction. The weights of clean and dry extraction
cups must also be obtained for later use in the final calculation. The samples and
extraction cups are then positioned in the extractor. The solvent is added through a
closed-loop addition process and the extraction begins. The steps of boiling, rins-
ing, and evaporation/solvent recovery then proceed in an automated manner. At the
end of the cycle, an alarm signals completion.
Boiling. In this step, the sample and thimble are lowered and totally immersed in
the boiling solvent contained in the extraction cup. The solvent vapor refluxes
against a water-jacketed cooling column and the condensed solvent flows back
continuously through the sample returning to the boiling solvent (Fig. 2.3). The
boiling step is the key to accelerating the extraction process compared with the
Soxhlet method. The solvent simply solubilizes the extract faster in hot solvent,
thus decreasing the time required for extraction.
TABLE 2.1
Expected Fat Content and Sample Weights
Fat content (%) Sample weight (g)
0–10 2–3
10–25 1–2
>25 0.5–1
Copyright © 2004 AOCS Press
To ensure optimal extraction, the level of boiling solvent must be higher than
the sample in the thimble. A plug of defatted cotton is frequently placed on top of
the sample to keep it in the thimble during extraction. With the Soxtec Avanti sys-
tem, 70–90 mL of solvent is used. Typical extraction times range from 20 to 40
min depending on the solvent and sample characteristics.
Immediately after the boiling step, the rinsing step begins. The sample is raised
and suspended over the boiling solvent. During rinsing, residual traces of the
extractable material are flushed out of the sample and are retained in the extraction
cup. This step is usually 10–20 min longer than the boiling step to ensure complete
extraction.

The last step in the crude fat extraction process is evaporation/solvent recovery.
The condensed solvent continues to boil and evaporate and, using an internal valve,
the condensate is redirected out of the condenser. The evaporation step is complete
when all solvent is driven from the cup, concentrating the extract. This usually
requires 7–10 min depending on the solvent. Excessive drying may oxidize the
extract, causing weight changes and erroneous readings. The Soxtec Avanti stores
the evaporated solvent in a common collection tank for reuse. The Soxtec Avanti
offers an optional fourth, cup predrying step, i.e., the extraction cups are raised a few
millimeters off the heating surface allowing radiant heat to complete the drying
cycle. This step is used in applications in which the extract is extremely heat labile.
Postextraction. Once the extraction process is completed, the cups are taken off
the Soxtec and placed in a drying oven at 103°C for 30 min to drive off any mois-
ture or solvent residuals. Extended drying, especially at higher temperatures,
should be avoided because it can cause oxidation of the fat extract and falsely high
results. Extraction cups are cooled completely to room temperature in a desiccator
before final weights are taken.
FIG. 2.3. Three-step extraction
procedure. The Foss-Tecator
Soxtec Avanti automated
extraction system is based on
the Randall modification of the
Soxhlet technique. In the boil-
ing and rinsing steps, solvent is
refluxed within the condenser.
During evaporation, solvent
flow is blocked from returning
to the extraction cup and flows
out tube (a) into a collection
tank (not shown).
Copyright © 2004 AOCS Press

Calculation of Results. Crude fat by solvent extraction is a gravimetric method.
The final result is calculated from the original sample weight and the weights of
the extraction cup before and after the extraction.
% Fat = (W
2
– W
1
)/W
3
× 100
where W
1
= weight of the extraction cup
W
2
= weight of the extraction cup + extract
W
3
= weight of the sample
All weights should be recorded to 0.1 mg (0.0001g).
Optimizing the Extraction Process
The Soxtec/Soxhlet extraction method for crude fat relies on separating sample
components on the basis of physical and chemical (solubility) properties. There are
several factors that influence the extraction; the most significant of these is the spe-
cific solvent that is being used. Nevertheless, the influence of sample preparation,
extraction timing, and general Soxtec operating conditions is important. Keeping in
mind the empirical nature of the analysis, consistency with all aspects of the proce-
dure is strongly recommended. An often overlooked aspect of a fat extraction
method is the predrying of the sample. As mentioned earlier, water in the sample
can contribute to error in two ways, i.e., it can act as a physical barrier preventing

dissolution of the fat into the solvent, thus generating low fat recoveries; it can also
contribute to falsely high apparent fat recoveries by allowing water-soluble compo-
nents such as urea or carbohydrates to be co-extracted with the fat. Unfortunately,
in the interest of time and productivity, many laboratories do not predry samples.
In these instances, the error potential for each type of sample should be investigat-
ed fully by carrying out extractions both with and without predrying.
Figure 2.4 illustrates apparent fat recovery on dried vs. undried samples.
Moisture in the samples ranges from 5 to 25%. Some samples show a “water
effect” more than others. Samples such as the texturized feeds, which contain
molasses, and the feedlot concentrate, which contains urea, are examples in which
failing to predry the sample can have a marked effect on recovery. Note that this is
also dependent on the solvent used. [%Recovery is defined as (%crude fat from the
undried sample/% crude fat from the dried sample) × 100].
Solvent Choices. The versatility of the Soxhlet/Soxtec extraction method allows
for the use of various classes of organic solvents. These include ethers, aliphatic,
aromatic, and chlorinated hydrocarbons, as well as alcohols. Due to the different
solubility characteristics of various solvents, a sample extraction will have some-
what different fat yields depending on the solvent. For crude fat extractions,
diethyl ether and petroleum ether are most commonly used. The peroxide-forming
Copyright © 2004 AOCS Press
nature of diethyl ether causes it to be a less than desirable choice for routine use in
the laboratory. This has caused many laboratories to look for an alternative solvent.
Commonly, petroleum ether is directly substituted. However, petroleum ethers or
ligroin, are not true ethers but mixtures of aliphatic hydrocarbons and can be pur-
chased in various formulations and boiling point ranges. Further complicating the
“pet ether” issue is that solvents are often recycled and reused in the Soxtec. This
can cause a change in the properties of petroleum ether because its more volatile
components may be driven off. This can cause a change or drift in the fat results.
Considering the innate variability of petroleum ether and the relative lower
recovery of plant-based lipids, it is not a suitable substitution for diethyl ether. An

experiment was undertaken (3,4) to compare the recovery of three common sol-
vents, petroleum ether, hexanes, and pentane, to that of diethyl ether in terms of
crude fat recovery. The objective was to find a solvent that is safer and has recov-
ery statistically equivalent to diethyl ether. The results are shown in Table 2.2.
From these results, it can be seen that hexanes yield a recovery equivalent to
that of diethyl ether with an R
2
of 0.9925. It should also be noted that for meat and
bone meal, petroleum ether also yields a good recovery. This is consistent with the
use of petroleum ether in the AOAC method 991.36 (7) for crude fat in meat and
meat products.
Extraction Times. In Soxtec extraction procedures, the timing for the boiling and
the rinsing steps is important. If the boiling or rinsing step is too short, the extrac-
tion will likely not adequately recover the fat in the sample. Most method develop-
ment protocols will define the extraction times at which the results closely match
FIG. 2.4. Apparent fat recovery from dried vs. undried samples with moisture content
ranging from 5 to 25%.
Dehydrated Alfalfa
Corn Silage
Mixed Bird Seed
Texturized Feed
(Molasses)
Fat Supplement
Medicated Goat Feed
Feedlot Conc. (Urea)
Calf Starter, Medicated
Calf Feed, Medicated
Meat Meal/Hulls Mixture
Swine Feed
Broiler Starter

High Oil Corn
Diethyl ether
Hexanes
% Recovery
Undried vs. Dried
Copyright © 2004 AOCS Press
those obtained by classical Soxhlet methods using suitable reference materials. The
automation of the Soxtec offers consistent extraction timing for each batch of sam-
ples.
Extraction Temperature. The temperature of the extraction system should be set
to the recommendations provided by the manufacturer. This helps ensure optimal
condensation or reflex rates. Ideally, this is typically 3–5 drops/s coming off the
condenser.
Condenser Temperature. The temperature of the condenser cooling water plays
an important role in establishing the condensation or reflux rate of the solvent.
Cold tap water, <20°C at 2 L/min, should be used so that the condensers feel cool
to the touch. If the water temperature is too warm, it will usually cause slow reflux
rates and can result in low fat recovery. In some cases, warm condensers can cause
the loss of solvent during the boiling and rinsing stages. In areas in which cold tap
water is seasonal or not obtainable or in which water conservation is an issue,
refrigerated circulating water baths are a convenient way to regulate the tempera-
ture and flow.
Robustness of a Method. Using the statistical model described by Youden (8) the
“ruggedness” of the extraction method was evaluated. The purpose of this exercise
was to determine whether the method can tolerate minor variations in the proce-
dure that might be encountered in the laboratory.
In this model, several variables or factors are identified and are selectively
used in a defined protocol or matrix. This is described in Table 2.3. For the extrac-
tion of crude fat, sample handling, weight, extraction parameters, and solvent types
were defined as variables. For example, sample weight of 1 g is described as vari-

able “E” and 3 g as “e.” Using the schedule in Table 2.3, and implementing the
variables in this manner allows the evaluation of the ruggedness of the method
while minimizing the number of determinations.
TABLE 2.2
Crude Fat Recovery of Four Common Solvents
Diethyl ether Petroleum ether Hexanes Pentane
Sample (% Crude fat)
Alfalfa hay 1.29 1.00 1.36 0.97
Beet pulp 0.30 0.24 0.25 0.19
Meat/bone meal 10.52 10.40 10.69 10.54
Cattle protein supplement 3.10 2.65 2.88 2.54
Corn 3.59 3.06 3.16 3.00
Average 3.76 3.47 3.67 3.45
R
2
— 0.9878 0.9925 0.9880
Copyright © 2004 AOCS Press
The results from three different samples, cattle, swine, and mixed feeds, from
three different laboratories are seen in Table 2.4. Values for the individual factors
are averages from all results in which this variable was used. Considering the aver-
ages of the differences, the most significant variable is the choice of solvent. Other
variables in the method do not contribute to a significant variation, thus making the
method robust.
Common Applications
Crude Fat in Meat and Meat Products: AOAC Method 991.36. A 2-g sample of
homogenized meat is mixed with acid-washed sand and dried at 125°C for 1 h. The
sand is added directly to the thimble at approximately double the sample weight. A
glass stir rod is used to thoroughly mix the sample and sand together. The glass rod is
left in the sample during drying and subsequently used to break up any sample/sand
clumps before extraction. The sand is used to maintain porosity of the sample after

drying for optimal solvent penetration. A petroleum ether Soxtec extraction is then
performed with 25-min boil and 35-min rinse periods. Table 2.5 compares recovery
and repeatability of classical Soxhlet to Soxtec (6) showing that the Soxtec offers bet-
ter precision with the same results as the classical Soxhlet method.
Crude Fat in Feed, Cereal Grains, and Forages: AOAC Methods 2003.05 and
2003.06. A 1- to 5-g ground sample is weighed into an extraction thimble. If the
sample contains quantities of water-soluble components such as >5% carbohy-
drates, >15% glycerol, lactic acid, or amino salts, or >10% of other water-soluble
TABLE 2.3
Variables Used to Determine Method Robustness
Combination or determination number
Factor value 1 2 3 4 5 6 7 8
A or a A A A A a a a a
B or b B B b b B B b b
C or c C c C c C c C c
D or d D D d d d d D D
E or e E e E e e E e E
F or f F f f F F f f F
G or g G g g G g G G g
Observed result S T U V W X Y Z
The chosen variables (factors):
A Sample predry 103°C, 2 h a Sample predry 103°C, 4 h
B Boil time, 20 min b Boil time, 40 min
C Diethyl ether c Petroleum ether
D Rinse time, 30 min d Rinse time, 60 min
E Sample weight, 1 g e Sample weight, 3 g
F Cup dry, 103°C, 2 h f Cup dry, 103°C, 4 h
G Solvent drip rate, 2/s g Solvent drip rate, 6/s
Copyright © 2004 AOCS Press
components, the sample is washed with 5 aliquots of 20 mL deionized water. The

sample is dried at 102°C for 2 h. Extraction is performed with either diethyl ether
or hexanes using a 20-min boil and 40-min rinse cycle. The Soxtec method was
compared with the AOAC Soxhlet method on data from 90 AAFCO check sam-
ples. Regression analysis generated an R
2
correlation coefficient of 0.9946, slope
TABLE 2.4
Evaluating the Robustness of the Crude Fat Method Using Cattle, Swine, and Mixed Feeds
from Three Different Laboratories
Laboratory 1 Laboratory 2 Laboratory 3
(% crude fat)
Feed type Cattle Swine Mixed Cattle Swine Mixed Cattle Swine Mixed Average
S 11.61 2.45 10.50 11.82 2.41 11.16 11.99 2.99 10.97
T 11.30 2.10 10.24 11.63 2.35 10.25 11.55 2.49 10.64
U 11.79 2.46 10.60 11.98 2.81 10.84 11.84 2.93 11.05
V 11.02 2.17 9.88 11.76 2.70 10.78 11.57 2.46 10.50
W 10.87 2.60 10.67 11.61 2.52 9.86 11.87 4.13 10.94
X 11.11 2.00 10.00 11.28 2.10 10.28 11.24 2.38 10.43
Y 11.62 2.55 10.60 11.49 2.64 10.53 11.94 2.82 10.91
Z 10.73 1.91 9.82 11.19 2.10 10.01 11.74 2.51 10.73
Predry
2 h 11.43 2.29 10.30 11.80 2.57 10.76 11.74 2.72 10.79
4 h 11.08 2.26 10.28 11.39 2.34 10.17 11.70 2.96 10.75
Difference 0.34 0.03 0.03 0.41 0.23 0.59 0.04 –0.24 0.04 0.16
Boil time
20 min 11.22 2.29 10.35 11.59 2.35 10.39 11.66 3.00 10.75
40 min 11.29 2.27 10.23 11.61 2.56 10.54 11.77 2.68 10.80
Difference –0.06 0.02 0.13 –0.02 –0.22 –0.15 –0.11 0.32 –0.05 –0.02
Ether
Diethyl 11.47 2.52 10.59 11.73 2.60 10.60 11.91 3.22 10.97

Petroleum 11.04 2.04 9.99 11.47 2.31 10.33 11.53 2.46 10.58
Difference 0.43 0.47 0.61 0.26 0.28 0.27 0.38 0.76 0.39 0.43
Rinse time
30 min 11.32 2.25 10.29 11.53 2.38 10.49 11.81 2.70 10.81
60 min 11.20 2.31 10.29 11.66 2.53 10.44 11.63 2.98 10.73
Difference 0.12 –0.06 0.00 –0.13 –0.16 0.05 0.17 –0.27 0.08 –0.02
Sample wt
1 g 11.31 2.20 10.23 11.57 2.36 10.57 11.70 2.70 10.80
3 g 11.20 2.35 10.35 11.62 2.55 10.36 11.73 2.98 10.75
Difference 0.11 –0.15 –0.12 –0.06 –0.20 0.22 –0.03 –0.27 0.05 –0.05
Cup dry
30 m 11.06 2.28 10.22 11.60 2.43 10.45 11.79 3.02 10.79
2 hr 11.46 2.28 10.36 11.60 2.48 10.48 11.64 2.66 10.76
Difference –0.40 0.01 –0.14 0.00 –0.04 –0.02 0.15 0.37 0.03 0.00
Drop rate
2/s 11.34 2.29 10.24 11.59 2.46 10.69 11.69 2.66 10.70
6/s 11.17 2.27 10.33 11.60 2.45 10.24 11.75 3.02 10.84
Difference 0.17 0.02 –0.09 –0.01 0.02 0.45 –0.06 –0.35 –0.14 0.00
Copyright © 2004 AOCS Press
of 1.00062 and a y-intercept of 0.137. On the basis of these data, the methods
appear to be comparable (3,4).
Environmental EPA 3541, SW 846. Of note, the Soxtec extraction procedure is
used widely in environmental laboratories to extract organic compounds such as
pesticides or PCB from soils, sludge, sediments, and hazardous waste samples
(9,10). In these applications, the Soxtec is used as a sample preparation device.
The sample is weighed directly into a cellulose or fritted glass thimble and placed
in the Soxtec where the extraction of semivolatile organics is done using a mixture
of hexane and acetone (1:1). The process is stopped during the evaporation/solvent
recovery step while there is still 15–20 mL of solvent (containing the semi-
volatiles) left in the cup. The extract/solvent mixture is further concentrated and

the final analysis performed by GC or GC/MS techniques. The Soxtec method with
a 2-h extraction replaces the traditional Soxhlet, which takes 8–24 h.
Total Fat. In samples that are baked, extruded, or with some commercial process-
ing, the fat becomes bound to other components in the sample such as proteins,
carbohydrates, and minerals. An acid hydrolysis before the solvent extraction step
is needed to “free” the fat in the sample, making it available for solvent extraction.
Typically 1–2 g of sample is boiled with strong hydrochloric acid solutions. The
sample is then rinsed, dried and then extracted. The SoxCap
TM
(Fig. 2.5) for acid
hydrolysis from Foss Tecator enables the acid hydrolysis step and extraction step
to occur in the same sample vessel, thus eliminating any sample transfer errors.
Industrial Applications. Soxtec extraction methods were found to be suitable for
use in industrial applications. A summary of some of these applications appears in
Table 2.6. Application Sub Notes (ASN) are available from Foss Tecator.
TABLE 2.5
Average Recoveries and Relative Standard Deviations (RSD) for Meat Samples with
Petroleum Ether Extraction
Average recovery RSD
r
Average recovery RSD
r
Soxhlet
a
Soxtec
b
Meat sample (%)
1 4.6 2.63 4.34 2.44
2 28.35 8.75 27.29 1.95
3 28.21 5.52 27.95 2.32

4 34.98 2.09 34.51 2.21
5 34.10 2.25 33.57 1.01
6 26.81 1.74 26.20 1.55
Average 26.18 3.83 25.64 1.91
a
Soxhlet: 4-h extraction, 2-h drying time.
b
Soxtec: 55-min extraction, 30-min drying time.
Copyright © 2004 AOCS Press
TABLE 2.6
Summary of Soxtec Extraction Methods Used in Industrial Applications
a
ASN 3516 Extraction of aromatic hydrocarbons in soil
ASN 3602 Extraction of resins from paper pulp
ASN 3603 Extraction of finish from textiles
ASN 3604 Extraction of starch containing finish from textiles
ASN 3605 Extraction of surfactant from detergents
ASN 3606 Extraction of paraffin from detergent
ASN 3607 Extractable matter in leather
ASN 3608 Extraction of petroleum source rock
ASN 3611 Extraction of explosives and propellants
ASN 3612 Extraction of plastics and polymers
ASN 3613 Extraction of rubber and rubber compounds
ASN 3614 Extraction of finish oils from textiles and synthetic fibers
ASN 3615 Extraction of migration components in plastic packaging
ASN 3616 Extraction of organic dyestuffs
ASN 3617 Extraction of leather
ASN 3618 Extraction of core material in petroleum exploration
ASN 3619 Extraction of tobacco
ASN 3622 Extraction of solubles in paper pulp

ASN 3700 Extraction of fecal fat
a
Application sub notes (ASN) are available from Foss Tecator.
FIG. 2.5. The 2047 SoxCap
TM
hydrolysis system.
Copyright © 2004 AOCS Press
Difficult Samples. Samples that represent special challenges and handling are fre-
quently encountered. Table 2.7 summarizes some common approaches to aid the
analyst in extracting crude fat from these types of samples.
Quality Control. Evaluating the performance of the extraction process is normal-
ly achieved by running a reference or check sample. Many commercial check sam-
TABLE 2.7
Common Approaches to Help Extract Crude Fat from Difficult Samples
High-fat samples that • Place the thimble containing the sample into a preweighed
melt during drying extraction cup. Any sample that melts out of the thimble will be
retained in the cup.
Incomplete fat recovery, • A two-phase extraction protocol is used: After the first extraction
high-fat seeds boil and rinse, the samples are removed from the thimbles,
ground with a mortar and pestle and returned to the thimble for
a secondary extraction. The results are added together.
Sample becomes impacted • Mix equal volumes of acid-washed sand or Celite and sample in
during extraction the extraction thimble. This allows for better solvent flow
through the sample.
Solvent overboiling • Use 3–5 boiling beads in the extraction cups.
• Decrease the temperature setting on the extractor.
Moist samples • Mix sample with sand and predry.
• Mix sample with equal weight of sodium sulfate to bind water.
Semisolid • Depending on the nature of the sample, mix with either sand
and dry or mix with sodium sulfate.

Nonhomogeneous samples • Optimize sample preparation step.
• Use larger sample size to obtain a representative sample.
• Do replicate analysis to generate reportable results.
Low-fat samples • Use larger sample weights.
General practice • Place a plug of defatted cotton on top of the sample to ensure
that the sample is retained in the thimble.
• Wear gloves during handling of thimbles and cups to avoid
errors.
• Weigh cups at room temperature. Weighing errors will result
from warm cups.
• When using recovered petroleum ether, supplement with fresh
ether to help maintain desired boiling point range.
• Diethyl ether can be purchased with stabilizers to minimize the
formation of peroxides. Such ether should be used with the
label guidelines.
• Test strips are available to check for peroxide formation in
diethyl ether.
Copyright © 2004 AOCS Press
ple services are available for different sample types. Results should be compared
only to those from similar instruments running the same extraction procedure and
sample protocol.
Conclusion
The long history of solvent extraction has led to automated Soxhlet extraction sys-
tems, such as the Soxtec
TM
Avanti. They offer convenient and useful tools with
which to improve productivity in the laboratory. Modern instrumentation provides
application flexibility and improved economy as well as enhancement of the preci-
sion and recovery of the extraction.
References

1. Randall, E.L. (1974) Improved Method for Fat and Oil Analysis by a New Process of
Extraction, JAOAC 57: 1165–1168.
2. Codex Alimentarius Commission (1986) Procedural Manual, 6th edn., p. 139, Food
and Agricultural Organization, Rome, Italy.
3. Thiex, N., Anderson, S., and Gildemeister, B. (2003) Crude Fat, Hexanes Extraction, in
Feed, Cereal Grain, & Forage (Randall/Soxtec/Submersion Method): A Collaborative
Study, JAOAC Int. 86: 888–898.
4. Thiex, N., Anderson, S., and Gildemeister, B. (2003) Crude Fat, Diethyl Ether
Extraction, in Feed, Cereal Grain, & Forage (Randall/ Soxtec/Submersion Method): A
Collaborative Study, JAOAC Int. 86: 899–908.
5. Official Methods of Analysis of AOAC International, 16th ed., Chapter 4, p. 25, section
4.5.01. AOAC Official Method 920.39, Fat (Crude) or Ether Extract in Animal Feed,
AOAC International, Gaithersburg, MD, 1997.
6. Foster, M.L., and Gonzales, S.E. (1992) Soxtec Fat Analyzer for Determination of Total
Fat in Meat: Collaborative Study, Kansas State Board of Agriculture, JAOAC Int. 75:
288–292.
7. Official Methods of Analysis of AOAC International, 16th ed., Chapter 39, p. 3, section
39.1.08. AOAC Official Method 991.36, Fat (Crude) in Meats and Meat Products,
AOAC International, Gaithersburg, MD, 1997.
8. Youden, W.J., and Steiner, E.H. (1975) Statistical Manual of the AOAC, Association of
the Official Analytical Chemists, Arlington, VA.
9. Lopez-Avila, V. (Beckert, W., Project Officer) (1991) Development of a Soxtec Extraction
Procedure for Extracting Organic Compounds from Soils and Sediments, EPA600/X-
91/140. U.S. EPA, Environmental Monitoring Systems Laboratory, Las Vegas.
10. Test Methods for Evaluation of Solid Waste, Physical/Chemical Methods (1996) SW-846,
Method 3541. U.S. Environmental Protection Agency, Office of Solid Waste, Washington,
DC.
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