Extraction procedures

Since lupin seeds are used in some areas in cattle feeding, it is of practical as well as theoretical interest to determine the stage at which the seeds will be rich in the alkaloidal material responsible for toxicity. It has also been important to devise methods for the removal of alkaloids from the seeds so that the detoxified or debittered material can still be used as feed (111). Extraction procedures which accent the recovery of non-alkaloidal material have less interest to the alkaloid chemist than those which provide for the isolation of the pure organic bases. Given below are typical examples of the extraction procedures employed for the isolation of the lupin alkaloids lupinine, cytisine, Z-sparteine, d-lupanine, and anagyrine. The methods selected are representative of those utilized for the isolation of the less abundant or well-known lupin alkaloids as well. These methods are also representative of the different quantities of materials which are handled. One of the methods was selected (for anagyrine) to indicate some of the complexities of separation when there are a number of alkaloids present in a plant, rather than only one main alkaloidal constituent. The techniques of fractional distillation of the bases, fractional crystallization of alkaloid salts, such as perchlorates and picrates, and extractions dependent upon differential solubility have been employed for the isolation of pure individual alkaloids from a mixture. 

Lupinine (78). Lupinua pdlmeri was air-dried and ground, and 60.9 kg. was extracted with ethanol. The solvent was distilled and the residual extract was boiled with successive portions of water until all soluble matter was removed. The concentrated aqueous solutions were treated with an excess of a mixture of neutral and basic lead acetates, filtered, and freed from the excess of lead with hydrogen sulfide. The filtrate was concentrated, made alkaline with sodium hydroxide, and extracted with chloroform. The solvent was distilled from the chloroform extract, leaving the 

Cytisine (12, 38). The ground tops and seeds of Laburnum vuLgare were air-dried and extracted in a soxhlet with 50 % aqueous ethanol to which had been added 2 % of acetic acid. The extract after concentration was cleared with lead acetate, concentrated, made alkaline with sodium hydroxide, and extracted six times with an equal volume of chloroform. This left only a trace of cytisine in the aqueous layer. When the chloroform was removed, the residual alkaloidal material crystallized in part. No clear-cut separation of components was accomplished by fractional crystallization of the free base or the crude picrate. The isolation of cytisine as the benzenesulfonyl derivative was effected according to the method of Ing (5), and benzenesulfonyl-cytisine was collected and recrystallized from ethanol as glistening prisms, m.p. 261 . Microchemical tests and picrate formation and identification were also used to establish the presence of cytisine in a particular plant. 

Some anagyrine was carried over with cytisine in fraction IV but the major portion of it was found in fraction V, which was dissolved in methanol and the solution was made just acid to Congo red by the cautious addition of 65 % perchloric acid. The crystalline anagyrine perchloroate thus obtained was recrystallized several times from boiling methanol, from which it separates as colorless needles m.p. 315°. An aqueous solution of the perchlorate was made ammoniacal and extracted with chloroform. The base, obtained by removal of the chloroform, was dissolved in methanol and added to a methanolic solution of picric acid. Anagyrine picrate separated as yellow needles m.p. 252°. 

Column Preparation a. In a large beaker, swirl about 100 g of Dowex 1-X8 resin, 50-100 mesh, (for resin 2) chloride form, with about 500 mL of 1N acetic acid. 

NOTE Steps b, c, and e may be facilitated with the use of a sintered glass filter. 

Fortify the blank tissue by adding 5 mL of the HPLC standard solution (section D.2.C.). This will yield a recovery of 100 ng/g. 

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A suspension of 3.90 g (19.6 mmol) of p-(bromomethyl)benzaldehyde (2.8) and 4.00 g (31.7 mmol) of sodium sulfite in 40 ml of water was refluxed for two hours, after which a clear solution was obtained. The reaction mixture was cooled on an ice bath resulting in precipitation of some sodium sulfite. After filtration, the solvent was evaporated. Ethanol was added to the remaining solid and the suspension was refluxed for 10 minutes. After filtering the hot solution, the filtrate was allowed to cool down slowly to -18 °C whereupon sodium (p-oxomethylphenyl)methylsulfonate (2.9) separated as colourless crystals. The extraction procedure was repeated two more times, affording 2.29 g (10.3 mmol, 53%) of the desired product. H-NMR (200 MH D2O) 5(ppm) =4.10 (s,2H) 7.44 (d,2H) 7,76 (d,2H) 9.75 (s,lH). 

Note 2. The separation of the layers may give some difficulties, owing to the presence of aluminium hydroxide. Too vigorous shaking during the extraction procedure should be avoided. The best way to separate the layers is to run off as much of the aqueous layer as possible and subsequently decant the ethereal layer. 

Miscellaneous Pharmaceutical Processes. Solvent extraction is used for the preparation of many products that ate either isolated from naturally occurring materials or purified during synthesis. Among these are sulfa dmgs, methaqualone [72-44-6] phenobarbital [50-06-6] antihistamines, cortisone [53-06-5] estrogens and other hormones (qv), and reserpine [50-55-5] and alkaloids (qv). Common solvents for these appHcations are chloroform, isoamyl alcohol, diethyl ether, and methylene chloride. Distribution coefficient data for dmg species are important for the design of solvent extraction procedures. These can be determined with a laboratory continuous extraction system (AKUEVE) (244). 

Direct attack by hot 70—80 wt % hydrofluoric acid, sometimes with nitric acid (qv), is effective for processiag columbites and tantalo-columbites. Yields are >90 wt%. This method, used in the first commercial separation of tantalum and niobium, is used commercially as a lead-in to solvent extraction procedures. The method is not suited to direct processiag of pyrochlores because of the large alkaU and alkaline-earth oxide content therein, ie, ca 30 wt %, and the corresponding high consumption of acid. 

The batch and fed-batch procedures are used for most commercial antibiotic fermentations. A typical batch fermentor may hold over 150,000 Hters. When a maximum yield of antibiotic is obtained, the fermentation broth is processed by purification procedures tailored for the specific antibiotic being produced. Nonpolar antibiotics are usually purified by solvent extraction procedures water-soluble compounds are commonly purified by ion-exchange methods. Chromatography procedures can readily provide high quaHty material, but for economic reasons chromatography steps are avoided if possible. 

Solvent extraction in batch or continuous systems is used to recover most of the residual oil from the presscake. Heptane, hexane, or a mixture of these solvents is used to recover the oil. The solvent-extracted presscake is steam stripped to recover solvent and a residual meal known as castor pomace, containing 1% residual oil. The solvent extracted oil is also processed for solvent recovery (qv). The oil from the extraction procedure is darker than the mechanically pressed oil and has a higher free fatty acid content. It is sometimes referred to as a No. 3 castor oil and is used for blending with higher quaUty oils that are well above No. 1 specifications. 

The tissue to be analyzed is placed directiy onto the gel. Using the tissue itself and not tissue extracts has advanced the study of proteins that are difficult to extract from tissue, or are damaged by the extraction procedure. Dtif is an important advancement in the area of sample handling and appHcation where direct appHcation of a soHd to a gel matrix may actually enhance resolution. 

Microwave extraction realized at 120 °C for 30 min with Hexane -Acetone (3 2 V/V) as the extraction solvent was identified as the most effective extraction procedure for isolation of TPH from biotic matrices. The aim of this research is to develop a silica gel and alumina fractionation procedure for plant sample extraction. Column chromatography with two solvents (chloroform and hexane dichloromethane) as a mobile phase were used for clean-up of extract. In this research the efficiency of recovery received from chloroform as a mobile phase. 

The behavior of elements (toxicity, bioavailability, and distribution) in the environment depends strongly on their chemical forms and type of binding and cannot be reliably predicted on the basis of the total concentration. In order to assess the mobility and reactivity of heavy metal (HM) species in solid samples (soils and sediments), batch sequential extraction procedures are used. HM are fractionated into operationally defined forms under the action of selective leaching reagents. 

The ability to identify and quantify cyanobacterial toxins in animal and human clinical material following (suspected) intoxications or illnesses associated with contact with toxic cyanobacteria is an increasing requirement. The recoveries of anatoxin-a from animal stomach material and of microcystins from sheep rumen contents are relatively straightforward. However, the recovery of microcystin from liver and tissue samples cannot be expected to be complete without the application of proteolytic digestion and extraction procedures. This is likely because microcystins bind covalently to a cysteine residue in protein phosphatase. Unless an effective procedure is applied for the extraction of covalently bound microcystins (and nodiilarins), then a negative result in analysis cannot be taken to indicate the absence of toxins in clinical specimens. Furthermore, any positive result may be an underestimate of the true amount of microcystin in the material and would only represent free toxin, not bound to the protein phosphatases. Optimized procedures for the extraction of bound microcystins and nodiilarins from organ and tissue samples are needed. 

It has been demonstrated that a solvent-extraction procedure with N-methyl pyrrolidone is capable of producing coal-derived extract pitches with low-ash contents. Moreover, the properties of the pitches can be varied by partial hydrogenation of the coal prior to extraction. The yield of the pitches along with the physical and chemical properties of the cokes and graphites vai in an understandable fashion. 

Physical and chemical tests of the final product may need to address two concerns (1) whether the solidified waste exhibits any RCRA defined toxicity characteristics or could be delisted and (2) the potential long term fate of treated materials in the disposal environment. Three tests are available which address the first concern. These are the Extraction Procedure (EP Tox) (40 CFR 261, Appendix II, 1980) and the Toxicity Characteristic Leaching Procedure (TCLP) (40 CFR 261, Appendix II, 1986), and the Multiple Extraction Procedure Test (40 CFR 261, Appendix II, January 1989). It is important to note that these tests are not indicators of expected leachate quality but of potentials. A solidified product which cannot pass the appropriate test (EP Tox or TCLP) would be subject to classification as a hazardous waste. 

Federal Register - Multiple Extraction Procedure Test, Revised 40 CFR 261, Appendix II, January 1989. 

Noller, H. F., Hoffarth, V, and Zimniak, L., 1992. Unusual resistance of peptidyl transferase to protein extraction procedures. Science 256 1416-1419. 

Thallium is likewise recovered from flue dusts emitted during sulfide roasting for H2SO4 manufacture, and from the smelting of Zn/Pb ores. Extraction procedures are complicated because of the need to recover Cd at the same time. There are no major commercial uses for T1 metal world production in 1983 was estimated to be 5-15 tonnes p.a. and the price ranged from 60 to 80 per kg depending on purity and amount purchased. 

The usual extraction procedure is to roast the crushed ore, or vanadium residue, with NaCl or Na2C03 at 850°C. This produces sodium vanadate, NaV03, which is leached out with water. Acidification with sulfuric acid to pH 2-3 precipitates red cake , a polyvanadate which, on fusing at 700°C, gives a black, technical grade vanadium pentoxide. Reduction is then necessary to obtain the metal, but, since about 80% of vanadium produced is used as an additive to steel, it is usual to effect the reduction in an electric furnace in the presence of iron or iron ore to produce ferrovanadium, which can then be used without further refinement. Carbon was formerly used as the reductant, but it is difficult to avoid the formation of an intractable carbide, and so it has been superseded by aluminium or, more commonly, ferrosilicon (p. 330) in which case lime is also added to remove the silica as a slag of calcium silicate. If pure vanadium metal is required it can... 

The use of SPME for CE has not (yet) been studied widely. Li and Weber (170) reported an off-line SPME-CE approach for the determination of barbiturates in urine and serum, utilizing a sorbent of plasticized PVC coated around a stainless steel rod. Eor extraction, the coated rod was inserted for 4 min in a Teflon tube containing 50 p.1 of sample, and next the rod was repeatedly desorbed in another Teflon tube which each time contained 5 p.1 of desorption solution. This solution was transferred to an injection vial and an aliquot was injected into the CE system (Eigure 11.19). The extraction procedure appeared to be selective and effectively allowed the handling of very small samples. 

In the second and following passes through the press, water was added to the pulp to increase the efficiency of the extraction procedure. The crude juice was screened to remove the coarse particles. Hydrogen sulfide gas was bled into the collected juice to partially saturate it. 

The cationic nature of the copper(I) catalyst means that it is immobilized in the ionic liquid. This permits the PMMA product to be obtained, with negligible copper contamination, by a simple extraction procedure with toluene (in which the ionic liquid is not miscible) as the solvent. The ionic liquid/catalyst solution was subsequently reused. 

The initial solution is characterized by a high level of acidity that excludes the possibility of selective extraction. Such solutions can be treated only using the collective extraction procedure. 

Automation of solvent extraction. Although automatic methods of analysis do not fall within the scope of the present text, it is appropriate to emphasise here that solvent extraction methods offer considerable scope for automation. A fully automated solvent extraction procedure, using APDC, for the determination of... 

In the extraction procedure the yellow solution is allowed to stand for 10 minutes, and then extracted with 3 mL portions of a 3 1 mixture by volume of pentan-l-ol and ethyl acetate until the last extract is colourless. Make up the combined extracts to a definite volume (10 mL or 25 mL) with the organic solvent, and determine the transmittance (460 nm) at once. Construct the calibration curve by extracting known amounts of bismuth under the same conditions as the sample. 

Procedure. Prepare four test solutions of phenol by placing 200 mL of boiled and cooled distilled water in each of four stoppered, 500 mL bottles, and adding to each 5g of sodium chloride this assists the extraction procedure by salting out the phenol. Add respectively 5.0, 10.0, 15.0 and 20.0 mL of the standard phenol solution to the four bottles, then adjust the pH of each solution to about 5 by the careful addition of 5M hydrochloric acid (use a test-paper). Add distilled... 

Extraction of the analyte or of the interfering element(s) is an obvious method of overcoming the effect of interferences . It is frequently sufficient to perform a simple solvent extraction to remove the major portion of an interfering substance so that, at the concentration at which it then exists in the solution, the interference becomes negligible. If necessary, repeated solvent extraction will reduce the effect of the interference even further and, equally, a quantitative solvent extraction procedure may be carried out so as to isolate the substance to be determined from interfering substances. 

Note. If lead caprate is not available, standard lead solutions can be prepared from aqueous solutions containing known weights of lead nitrate and following through the extraction procedure as detailed for the final extraction of lead into methyl isobutyl ketone for the alloy. It should also be noted that steps should be taken to avoid excessive inhalation of the vapour of the methyl isobutyl ketone, which can cause a headache. 

Unsubstituted phthalocyanines can readily be purified by sublimation or by dissolution in concentrated sulfuric acid followed by precipitation in water. These classical methods of purification are applicable to phthalocyanines due to their high stability towards heat and acid. Simple washing or extraction procedures using water and organic solvents can also be used. 

The increased solubility of substituted phthalocyanines (vide infra) enables more common purifications as used for other organic compounds. Usually the purification is done by chromatography either on alumina or silica gel, but recrystallization and extraction procedures can also be used. In some cases, the methods used for unsubstituted phthalocyanines can also be practiced, although the increased molecular weight accompanied by a reduced thermal stability makes sublimation more difficult.97 98 However, for substituted phthalocyanines, the stability towards acid may be reduced97 and, therefore, purification by treatment with sulfuric acid cannot generally be recommended. 

The tightly bound chromophore could be extracted from the protein with methanol [186], and the major component of the extract was determined to have the enediyne structure 116 (Figure 11.21), related to chromophores of other chromoprotein antitumor agents such as neocarzinostatin. Additional minor components were extracted, variously containing an OH group instead of OMe attached to the enediyne core, with Cl instead of OMe when chloride was present in the buffer salt, or with OEt instead of OMe when ethanol was used for the extraction. Another byproduct was isolated in the form of structure 117, consistent with a facile cy-doaromatization reaction as observed for all other enediyne antibiotics. Surprisingly, 117 also displayed antibiotic and antitumor activity, perhaps due to alkylation of DNA or protein by the aziridine. The interpretation of these results was that 116 and the other enediyne byproducts were merely artifacts of the extraction procedure and that the true structure of the maduropeptin chromophore is the aziridine 118. 

Complex ( + )-(FeS)-3 is obtained by extraction of the diastereomeric mixture with 50 mL of pentane in portions at r.t. The specific rotation of the solid product obtained from the first fraction is [a]jJ6 +50 (o = 0.001. benzene). The rotation of the remaining mixture becomes more negative as the (+)-isomer is removed. After concentration of the combined fractions, the resulting material, enriched in the ( + )-isomcr, is further purified by extraction with 10 mL of pentane in portions. Repetition of the pentane extraction procedure affords ( + )-(FeS)-3 yield-. 750 mg (30%) mp 120"C (dee) [a] +72 (c = 0.001, benzene). The residue, enriched in (—)-(FeR)-3 is extracted for 1 h with boiling pentane. After separation from the extraction liquor, a yellow-orange solid is obtained yield I -35 g (50%) mp 120°C (dec) — 120... 

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