soxtec: its principles and applications

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 SoxtecTM. Using the Randall modification, sometimescalled the submersion method, the Soxtec provides a faster approach to solventextraction 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 anempirical 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 therecently approved AOAC method for determining crude fat in feeds, cereal grains,and forages. Many Soxtec applications are routinely used in food, feed, industrial, andenvironmental laboratories for the measurement of fats, oils, semivolatiles, and othersolvent “extractables.” For the determination of crude fat, descriptions are given forvarious 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 to1879 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 stationedover boiling solvent. Condensed solvent would then drip into the sample, solubiliz-ing extractable material and then siphon back into the boiling solvent, where thiscycle would then repeat. After several cycles over many hours, the apparatus isdisassembled and the solvent, now containing extract (fat), is evaporated off, leav-ing the residue for further analysis. The Soxhlet procedure remains the mostexhaustive extraction technique, and today it is still widely used.

Over the years, there had been some improvements to the basic technique butthe procedure remained long, tedious, and prone to variability. In the early 1970s,Edward Randall (1) developed an accelerated extraction technique that cut theextraction 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 thanin cold or room temperature solvent. His procedure included this new boiling stepfollowed by a rinsing step to flush residual extract from the sample (Fig. 2.1).Demonstrating excellent agreement with Soxhlet and improved precision, theRandall method has become the basis of many automated extraction systems.

In 1975, Tecator AB of Höganäs Sweden acquired the rights to what hadbecome known as the “Randall modification” of the Soxhlet method. This was firstcommercialized as the RaFaTec and later became the Soxtec systems of today.Figure 2.2 is a photograph of the SoxtecTM Avanti, from Foss-Tecator, an automat-ed Soxhlet extraction system using the Randall submersion technique.

Procedural Overview of Soxtec, Automated Soxhletfor Crude Fat

Automated extraction methods using the Soxtec have gained widespread accep-tance and have a number of regulatory agency approvals worldwide. The Soxtecmethod has been used on literally hundreds of different sample types for manyextracts. For our purposes, this discussion will be limited mainly to crude fatextraction. 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 orvariables 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 areextremely 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) thimblepositioning mechanism (g) heater(not shown on the Soxhlet). In theoriginal Randall method, the thim-ble is positioned by use of theslide rod (f). Lowering the thimble(b) into the boiling solvent for theboiling step, then raising it out ofthe solvent for the rinsing step. Inboth stages, condensed solvent isflowing continuously through thesample and thimble back into theboiling solvent.


pared sample can invalidate even the most carefully performed extraction proce-dure. Depending on the type and nature of the sample, the sample preparation mayincorporate different procedures. For grinding and weighing, the sample should behomogenous 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 ormilling 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 andin most cases, can be weighed directly into the cellulose thimble. The weight of thesample 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 iscritical that this small sample be representative of the larger sample lot.

Pretreatment

Drying. Most samples should be predried to optimize the fat extraction. Water inthe sample can decrease the efficiency of the solvent extraction, resulting in low fatrecoveries. Conversely, water-soluble components in the samples such as urea, car-bohydrates, salts, and glycerol can be extracted with fat yielding falsely highrecoveries.

Samples are weighed into the extraction thimbles and are then typically driedat 102 ± 2°C for 1–2 h (3–5,7). Because the samples are weighed before drying,

FIG. 2.2. The SoxtecTM Avanti 2050 automated extraction system.


the results are on an as-is basis. Results expressed on a dry matter basis must becalculated from a separate moisture determination. For very moist samples, sandmay be mixed with the sample before drying. This prevents the sample frombecoming 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 fatthat is bound to proteins, carbohydrates, and/or minerals, making it unavailable forsolubilization. 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 componentsmay exhibit poor solvent extraction efficiency. A preextraction with water, fol-lowed by a thorough drying step can be used to obtain an acceptable recovery byremoving these water-soluble components. The procedure specifies washing theweighed sample with 5 aliquots of 20 mL of deionized water. The sample is driedand the extraction procedure is carried out as usual. (3,4)

Solvent Extraction

Once the samples have been properly prepared and pretreated, they can now beplaced in the Soxtec for fat extraction. The weights of clean and dry extractioncups must also be obtained for later use in the final calculation. The samples andextraction cups are then positioned in the extractor. The solvent is added through aclosed-loop addition process and the extraction begins. The steps of boiling, rins-ing, and evaporation/solvent recovery then proceed in an automated manner. At theend of the cycle, an alarm signals completion.

Boiling. In this step, the sample and thimble are lowered and totally immersed inthe boiling solvent contained in the extraction cup. The solvent vapor refluxesagainst a water-jacketed cooling column and the condensed solvent flows backcontinuously through the sample returning to the boiling solvent (Fig. 2.3). Theboiling step is the key to accelerating the extraction process compared with theSoxhlet 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 2–310–25 1–2>25 0.5–1


To ensure optimal extraction, the level of boiling solvent must be higher thanthe sample in the thimble. A plug of defatted cotton is frequently placed on top ofthe 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 40min depending on the solvent and sample characteristics.

Immediately after the boiling step, the rinsing step begins. The sample is raisedand suspended over the boiling solvent. During rinsing, residual traces of theextractable material are flushed out of the sample and are retained in the extractioncup. This step is usually 10–20 min longer than the boiling step to ensure completeextraction.

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 completewhen all solvent is driven from the cup, concentrating the extract. This usuallyrequires 7–10 min depending on the solvent. Excessive drying may oxidize theextract, causing weight changes and erroneous readings. The Soxtec Avanti storesthe evaporated solvent in a common collection tank for reuse. The Soxtec Avantioffers an optional fourth, cup predrying step, i.e., the extraction cups are raised a fewmillimeters off the heating surface allowing radiant heat to complete the dryingcycle. 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 offthe 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 highresults. Extraction cups are cooled completely to room temperature in a desiccatorbefore final weights are taken.

FIG. 2.3. Three-step extractionprocedure. The Foss-TecatorSoxtec Avanti automatedextraction system is based onthe Randall modification of theSoxhlet technique. In the boil-ing and rinsing steps, solvent isrefluxed within the condenser.During evaporation, solventflow is blocked from returningto the extraction cup and flowsout tube (a) into a collectiontank (not shown).


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 ofthe extraction cup before and after the extraction.

% Fat = (W2 – W1)/W3 × 100

where W1 = weight of the extraction cupW2 = weight of the extraction cup + extractW3 = 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 samplecomponents on the basis of physical and chemical (solubility) properties. There areseveral 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 inmind the empirical nature of the analysis, consistency with all aspects of the proce-dure is strongly recommended. An often overlooked aspect of a fat extractionmethod is the predrying of the sample. As mentioned earlier, water in the samplecan contribute to error in two ways, i.e., it can act as a physical barrier preventingdissolution of the fat into the solvent, thus generating low fat recoveries; it can alsocontribute 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 “watereffect” more than others. Samples such as the texturized feeds, which containmolasses, and the feedlot concentrate, which contains urea, are examples in whichfailing to predry the sample can have a marked effect on recovery. Note that this isalso dependent on the solvent used. [%Recovery is defined as (%crude fat from theundried sample/% crude fat from the dried sample) × 100].

Solvent Choices. The versatility of the Soxhlet/Soxtec extraction method allowsfor the use of various classes of organic solvents. These include ethers, aliphatic,aromatic, and chlorinated hydrocarbons, as well as alcohols. Due to the differentsolubility 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


nature of diethyl ether causes it to be a less than desirable choice for routine use inthe laboratory. This has caused many laboratories to look for an alternative solvent.Commonly, petroleum ether is directly substituted. However, petroleum ethers orligroin, 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. Thiscan cause a change in the properties of petroleum ether because its more volatilecomponents 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 lowerrecovery of plant-based lipids, it is not a suitable substitution for diethyl ether. Anexperiment 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 ofcrude 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 tothat of diethyl ether with an R2 of 0.9925. It should also be noted that for meat andbone meal, petroleum ether also yields a good recovery. This is consistent with theuse of petroleum ether in the AOAC method 991.36 (7) for crude fat in meat andmeat products.

Extraction Times. In Soxtec extraction procedures, the timing for the boiling andthe 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 contentranging from 5 to 25%.

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those obtained by classical Soxhlet methods using suitable reference materials. Theautomation of the Soxtec offers consistent extraction timing for each batch of sam-ples.

Extraction Temperature. The temperature of the extraction system should be setto the recommendations provided by the manufacturer. This helps ensure optimalcondensation or reflex rates. Ideally, this is typically 3–5 drops/s coming off thecondenser.

Condenser Temperature. The temperature of the condenser cooling water playsan 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 coolto the touch. If the water temperature is too warm, it will usually cause slow refluxrates and can result in low fat recovery. In some cases, warm condensers can causethe loss of solvent during the boiling and rinsing stages. In areas in which cold tapwater 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 exercisewas 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 selectivelyused 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 typeswere 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 thevariables in this manner allows the evaluation of the ruggedness of the methodwhile 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 0.97Beet pulp 0.30 0.24 0.25 0.19Meat/bone meal 10.52 10.40 10.69 10.54Cattle protein supplement 3.10 2.65 2.88 2.54Corn 3.59 3.06 3.16 3.00Average 3.76 3.47 3.67 3.45R2 — 0.9878 0.9925 0.9880


The results from three different samples, cattle, swine, and mixed feeds, fromthree different laboratories are seen in Table 2.4. Values for the individual factorsare 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. Othervariables in the method do not contribute to a significant variation, thus making themethod robust.

Common Applications

Crude Fat in Meat and Meat Products: AOAC Method 991.36. A 2-g sample ofhomogenized meat is mixed with acid-washed sand and dried at 125°C for 1 h. Thesand is added directly to the thimble at approximately double the sample weight. Aglass stir rod is used to thoroughly mix the sample and sand together. The glass rod isleft in the sample during drying and subsequently used to break up any sample/sandclumps before extraction. The sand is used to maintain porosity of the sample afterdrying for optimal solvent penetration. A petroleum ether Soxtec extraction is thenperformed with 25-min boil and 35-min rinse periods. Table 2.5 compares recoveryand 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 and2003.06. A 1- to 5-g ground sample is weighed into an extraction thimble. If thesample 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 aB or b B B b b B B b bC or c C c C c C c C cD or d D D d d d d D DE or e E e E e e E e EF or f F f f F F f f FG or g G g g G g G G gObserved result S T U V W X Y ZThe chosen variables (factors):

A Sample predry 103°C, 2 h a Sample predry 103°C, 4 hB Boil time, 20 min b Boil time, 40 minC Diethyl ether c Petroleum etherD Rinse time, 30 min d Rinse time, 60 minE Sample weight, 1 g e Sample weight, 3 gF Cup dry, 103°C, 2 h f Cup dry, 103°C, 4 hG Solvent drip rate, 2/s g Solvent drip rate, 6/s


components, the sample is washed with 5 aliquots of 20 mL deionized water. Thesample is dried at 102°C for 2 h. Extraction is performed with either diethyl etheror hexanes using a 20-min boil and 40-min rinse cycle. The Soxtec method wascompared with the AOAC Soxhlet method on data from 90 AAFCO check sam-ples. Regression analysis generated an R2 correlation coefficient of 0.9946, slope

TABLE 2.4 Evaluating the Robustness of the Crude Fat Method Using Cattle, Swine, and Mixed Feedsfrom 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 10.97T 11.30 2.10 10.24 11.63 2.35 10.25 11.55 2.49 10.64U 11.79 2.46 10.60 11.98 2.81 10.84 11.84 2.93 11.05V 11.02 2.17 9.88 11.76 2.70 10.78 11.57 2.46 10.50W 10.87 2.60 10.67 11.61 2.52 9.86 11.87 4.13 10.94X 11.11 2.00 10.00 11.28 2.10 10.28 11.24 2.38 10.43Y 11.62 2.55 10.60 11.49 2.64 10.53 11.94 2.82 10.91Z 10.73 1.91 9.82 11.19 2.10 10.01 11.74 2.51 10.73

Predry2 h 11.43 2.29 10.30 11.80 2.57 10.76 11.74 2.72 10.794 h 11.08 2.26 10.28 11.39 2.34 10.17 11.70 2.96 10.75Difference 0.34 0.03 0.03 0.41 0.23 0.59 0.04 –0.24 0.04 0.16

Boil time20 min 11.22 2.29 10.35 11.59 2.35 10.39 11.66 3.00 10.7540 min 11.29 2.27 10.23 11.61 2.56 10.54 11.77 2.68 10.80Difference –0.06 0.02 0.13 –0.02 –0.22 –0.15 –0.11 0.32 –0.05 –0.02

EtherDiethyl 11.47 2.52 10.59 11.73 2.60 10.60 11.91 3.22 10.97Petroleum 11.04 2.04 9.99 11.47 2.31 10.33 11.53 2.46 10.58Difference 0.43 0.47 0.61 0.26 0.28 0.27 0.38 0.76 0.39 0.43

Rinse time30 min 11.32 2.25 10.29 11.53 2.38 10.49 11.81 2.70 10.8160 min 11.20 2.31 10.29 11.66 2.53 10.44 11.63 2.98 10.73Difference 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 10.803 g 11.20 2.35 10.35 11.62 2.55 10.36 11.73 2.98 10.75Difference 0.11 –0.15 –0.12 –0.06 –0.20 0.22 –0.03 –0.27 0.05 –0.05

Cup dry30 m 11.06 2.28 10.22 11.60 2.43 10.45 11.79 3.02 10.792 hr 11.46 2.28 10.36 11.60 2.48 10.48 11.64 2.66 10.76Difference –0.40 0.01 –0.14 0.00 –0.04 –0.02 0.15 0.37 0.03 0.00

Drop rate2/s 11.34 2.29 10.24 11.59 2.46 10.69 11.69 2.66 10.706/s 11.17 2.27 10.33 11.60 2.45 10.24 11.75 3.02 10.84Difference 0.17 0.02 –0.09 –0.01 0.02 0.45 –0.06 –0.35 –0.14 0.00


of 1.00062 and a y-intercept of 0.137. On the basis of these data, the methodsappear to be comparable (3,4).

Environmental EPA 3541, SW 846. Of note, the Soxtec extraction procedure isused widely in environmental laboratories to extract organic compounds such aspesticides 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 placedin the Soxtec where the extraction of semivolatile organics is done using a mixtureof hexane and acetone (1:1). The process is stopped during the evaporation/solventrecovery 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 andthe final analysis performed by GC or GC/MS techniques. The Soxtec method witha 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 stepis 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. Thesample is then rinsed, dried and then extracted. The SoxCapTM (Fig. 2.5) for acidhydrolysis from Foss Tecator enables the acid hydrolysis step and extraction stepto occur in the same sample vessel, thus eliminating any sample transfer errors.

Industrial Applications. Soxtec extraction methods were found to be suitable foruse in industrial applications. A summary of some of these applications appears inTable 2.6. Application Sub Notes (ASN) are available from Foss Tecator.

TABLE 2.5 Average Recoveries and Relative Standard Deviations (RSD) for Meat Samples withPetroleum Ether Extraction

Average recovery RSDr Average recovery RSDr

Soxhleta Soxtecb

Meat sample (%)

1 4.6 2.63 4.34 2.442 28.35 8.75 27.29 1.953 28.21 5.52 27.95 2.324 34.98 2.09 34.51 2.215 34.10 2.25 33.57 1.016 26.81 1.74 26.20 1.55Average 26.18 3.83 25.64 1.91aSoxhlet: 4-h extraction, 2-h drying time.bSoxtec: 55-min extraction, 30-min drying time.


TABLE 2.6 Summary of Soxtec Extraction Methods Used in Industrial Applicationsa

ASN 3516 Extraction of aromatic hydrocarbons in soilASN 3602 Extraction of resins from paper pulpASN 3603 Extraction of finish from textilesASN 3604 Extraction of starch containing finish from textilesASN 3605 Extraction of surfactant from detergentsASN 3606 Extraction of paraffin from detergentASN 3607 Extractable matter in leatherASN 3608 Extraction of petroleum source rockASN 3611 Extraction of explosives and propellantsASN 3612 Extraction of plastics and polymersASN 3613 Extraction of rubber and rubber compoundsASN 3614 Extraction of finish oils from textiles and synthetic fibersASN 3615 Extraction of migration components in plastic packagingASN 3616 Extraction of organic dyestuffsASN 3617 Extraction of leatherASN 3618 Extraction of core material in petroleum explorationASN 3619 Extraction of tobaccoASN 3622 Extraction of solubles in paper pulpASN 3700 Extraction of fecal fataApplication sub notes (ASN) are available from Foss Tecator.

FIG. 2.5. The 2047 SoxCapTM hydrolysis system.


Difficult Samples. Samples that represent special challenges and handling are fre-quently encountered. Table 2.7 summarizes some common approaches to aid theanalyst 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 preweighedmelt 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 extractionhigh-fat seeds boil and rinse, the samples are removed from the thimbles,

ground with a mortar and pestle and returned to the thimble fora secondary extraction. The results are added together.

Sample becomes impacted • Mix equal volumes of acid-washed sand or Celite and sample induring 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 sandand 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 ensurethat the sample is retained in the thimble.

• Wear gloves during handling of thimbles and cups to avoiderrors.

• Weigh cups at room temperature. Weighing errors will resultfrom warm cups.

• When using recovered petroleum ether, supplement with freshether to help maintain desired boiling point range.

• Diethyl ether can be purchased with stabilizers to minimize theformation of peroxides. Such ether should be used with thelabel guidelines.

• Test strips are available to check for peroxide formation indiethyl ether.


ple services are available for different sample types. Results should be comparedonly to those from similar instruments running the same extraction procedure andsample protocol.

Conclusion

The long history of solvent extraction has led to automated Soxhlet extraction sys-tems, such as the SoxtecTM Avanti. They offer convenient and useful tools withwhich to improve productivity in the laboratory. Modern instrumentation providesapplication 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 ofExtraction, JAOAC 57: 1165–1168.

2. Codex Alimentarius Commission (1986) Procedural Manual, 6th edn., p. 139, Foodand Agricultural Organization, Rome, Italy.

3. Thiex, N., Anderson, S., and Gildemeister, B. (2003) Crude Fat, Hexanes Extraction, inFeed, Cereal Grain, & Forage (Randall/Soxtec/Submersion Method): A CollaborativeStudy, JAOAC Int. 86: 888–898.

4. Thiex, N., Anderson, S., and Gildemeister, B. (2003) Crude Fat, Diethyl EtherExtraction, in Feed, Cereal Grain, & Forage (Randall/ Soxtec/Submersion Method): ACollaborative Study, JAOAC Int. 86: 899–908.

5. Official Methods of Analysis of AOAC International, 16th ed., Chapter 4, p. 25, section4.5.01. 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 TotalFat 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, section39.1.08. 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 ofthe Official Analytical Chemists, Arlington, VA.

9. Lopez-Avila, V. (Beckert, W., Project Officer) (1991) Development of a Soxtec ExtractionProcedure 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.


soxtec: its principles and applications

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