Capillary gas chromatographic analysis of amino acids by enantiomer labeling

The optical isomers of amino acids can be easily separated by gas chromatography using capillary columns coated with the chiral polysiloxane peptide, Chirasil-Val. Quantitative trace amino acid analysis in complex mixtures such as biological fluids, sea water, or protein hydrolysates can be achieved by enantiomer labeling: The D-amino acid enantiomers, which do not occur naturally, are added to the sample prior to analysis as internal standards. Because the D-enantiomers show the same physical and chemical properties as the natural L-enantiomers, they are ideal standard references. In routine analysis, the derivatization is achieved with a new automated derivatization robot. The D-standard serves as overall internal standard for the whole analytical procedure from sample enrichment to derivatization, chromatography, and response of the detector.     

Capillary gas chromatographic determination of prometryn and its degradation products in parsley

Residue analysis of the herbicide prometryn (2,4-bis(isopropylamino)-6-methylthio-1,3,5-triazine) is widely known, but an analytical method for determining its metabolities or degradation products in addition to the parent chemical has not yet been reported in the literature. The procedure reported here is for the extraction and determination of prometryn and 2 metabolites, 2-amino-4-isopropylamino-6-methyl-thio-1,3,5-triazine and 2,4-diamino-6-methylthio-1,3,5-triazine, in parsley. Crops were extracted with 2-propanol followed by concentration of the extract and partitioning with a minimum amount of hexane in the presence of a large excess of water to remove most of the green pigment. The aqueous phase was divided into 2 equal halves: (A) One-half portion was partitioned with dichloromethane in the presence of saturated sodium chloride solution, the dichloromethane phase was separated, and the aqueous phase was discarded. The organic solvent was evaporated, and the contents were reconstituted in petroleum ether before prometryn analysis. (B) The other half was made slightly alkaline with ammonium hydroxide solution and was partitioned with ethyl acetate in the presence of saturated sodium chloride solution. The ethyl acetate phase was concentrated, centrifuged to remove any turbidity, and analyzed for the 2 metabolities above. Fused silica capillary gas chromatography (GC) with nitrogen-phosphorus (N-P) detection was used for quantitation. The limit of detection was 0.05 mg/kg for all the compounds examined. Recoveries from fortified parsley samples ranged from 59 to 73% at fortification levels of 0.05 to 1.0 mg/kg.     

Capillary gas chromatographic determination with electron capture detector of beta-propiolactone in biological materials

A method has been developed for the determination of beta-propiolactone by derivatizing it to the volatile N-hexyl-3-heptafluorobutanoyloxypropanamide, which can be separated and identified by a capillary CP-Sil 8 column, and detected by an electron capture detector (ECD). First, beta-propiolactone is reacted with N-hexylamine to yield N-hexyl-3-hydroxypropanamide. The fluorobutanoyl ester derivative is next prepared by using heptafluorobutyric acid anhydride in the presence of trimethylamine. The method is very sensitive, simple, and specific, and can be used to detect and quantitate residual beta-propiolactone in lyophilized biological materials. The limit of detection is 0.2 ppm beta-propiolactone in a 50 mg sample; however, because of variability at low levels, the limit of quantitation is 1 ppm. Detector response was linear for 2-500 mg beta-propiolactone. Recoveries were 98% or greater from lyophilized vaccines spiked at the 2-20 ppm level. No side products or interference peaks were observed in the derivatization reaction.     

Capillary gas chromatographic determination of some chlorobiphenyls in eel fat: interlaboratory study

A method for determination of 6 individual chlorobiphenyls in eel fat, based on saponification of the sample and determination with capillary gas chromatography, was studied collaboratively. Eleven laboratories submitted analytical results in duplicate for 6 individual chlorobiphenyls on 2 naturally contaminated eel fat samples. The reproducibility coefficient of variation was about 14% at the 1 mg/kg level for each chlorobiphenyl compound and about 23% at the 0.1 mg/kg level. For each compound, the mean recovery was about 90% with a standard deviation varying from 7 to 9%.     

Capillary gas chromatographic method for determination of benzylpenicillin and other beta-lactam antibiotics in milk

A capillary gas chromatographic method is described for determining residues of beta-lactam antibiotic residues in milk, with specificity for benzylpenicillin (penicillin G), phenoxymethylpenicillin, methicillin, oxacillin, cloxacillin, dicloxacillin, and nafcillin. Residues are extracted from milk with acetonitrile. Samples are cleaned up by partitioning between aqueous and organic phases at different pH values. The penicillin residues are methylated with diazomethane to render them amenable to determination by gas chromatography on a methyl silicone fused silica column. Samples are introduced by split/splitless injection using a programmed temperature vaporization injector and are detected by nitrogen-selective thermionic detection. Internal standardization is used for quantitation. The limits of detection for all penicillins are well below 1 microgram/kg. Recoveries of spiked samples at 3 and 10 micrograms/kg are in the range of 42-85% (coefficients of variation 2-5%) and 41-92% (coefficients of variation 3-7%), respectively.     

Capillary column gas chromatographic determination of dicamba in water, including mass spectrometric confirmation

A sensitive method is described for determining dicamba at low micrograms/L levels in ground waters by capillary column gas chromatography with electron-capture detection (GC-EC); compound identity is confirmed by gas chromatography-mass spectrometry (GC-MS) using selected ion monitoring. Dicamba residue is hydrolyzed in KOH to form the potassium salt. The sample is then extracted with ethyl ether which is discarded. The aqueous phase is acidified to pH less than 1 and extracted twice with ethyl ether. The combined ethyl ether extracts are concentrated, and the residue is methylated using diazomethane to form the corresponding dicamba ester. The derivatized sample is cleaned up on a deactivated silica gel column. The methylated dicamba is separated on an SE-30 capillary column and quantitated by electron-capture or mass spectrometric detection. Average recoveries (X +/- SD) for ground water samples fortified with 0.40 microgram/L of dicamba are 86 +/- 5% by GC-EC and 97 +/- 7% by GC-MS detections. The EDL (estimated detection limit) for this method is 0.1 microgram dicamba/L water (ppb).     

Capillary column gas chromatographic determination of ethyl carbamate in alcoholic beverages with confirmation by gas chromatography/mass spectrometry

A method is described for determining ethyl carbamate at low microgram/kg levels in several types of alcoholic beverages by capillary column gas chromatography with Hall electrolytic conductivity detection and confirmation by mass spectrometry. Samples are diluted to obtain a uniform concentration of ethanol (ca 10%) then saturated with NaCl and extracted with methylene chloride. Extracts are evaporated to a small volume and injected in ethyl acetate solution for chromatographic analysis. The method was evaluated by 5 laboratories, 4 employing the Hall detector and one using mass spectrometric detection. Overall between-laboratory mean percent recoveries were: wine, 85.3 +/- 21.0% coefficient of variation (CV) (spiking level 20-45 micrograms/kg); sherry, 83.8 +/- 16.1% CV (spiking level, 81-142 micrograms/kg); whiskey, 79.5 +/- 13.9% CV (spiking level 127-190 micrograms/kg); and brandy, 85.0 +/- 12.5% CV (spiking level 297-446 micrograms/kg). Mass spectrometric results agreed well with the Hall results for all commodities. Detection limits were about 5 micrograms/kg for the Hall detector and about 0.5 microgram/kg for mass spectrometric detection.     

Capillary column gas chromatographic determination of trace residues of the herbicide chlorsulfuron in agricultural runoff water

A capillary column gas chromatographic method is described for determining parts-per-trillion (ppt) levels of chlorsulfuron in agricultural runoff water. The water sample is acidified with acetic acid and extracted with methylene chloride. The chlorsulfuron in the extracts derivatized to its monomethyl derivative. After Florisil column cleanup, the methylated chlorsulfuron is determined by electron-capture gas chromatography. Recovery of chlorsulfuron from fortified water samples is greater than 80%. Detection limit of the method is 25 ng chlorsulfuron/L water (25 ppt). There are 2 reaction sites on the chlorsulfuron molecule, both of which are susceptible to methylation leading to monomethyl chlorsulfuron and dimethyl chlorsulfuron. A procedure is described to methylate selectively the sulfonamide nitrogen of chlorsulfuron.     

Capillary gas chromatographic determination of cyclohexanone and 2-ethyl-1-hexanol leached from solution administration sets

A capillary gas chromatographic method is described for the determination of cyclohexanone and 2-ethyl-1-hexanol leached from solution administration sets. A preliminary study was made of compounds leached from solution administration sets by 5% sodium bicarbonate solution (pH 8.1), 0.9% sodium chloride solution (pH 6.8), and water. Water was selected as the leaching solvent because similar quantities of the compounds were leached into water and into both types of parenteral solutions. The correlation coefficients were 0.99977 for cyclohexanone and 0.99974 for 2-ethyl-1-hexanol, and recoveries were good (93-94%). Five administration sets from each of 2 manufacturers were analyzed by this method. The amounts of cyclohexanone that were leached from the individual sets varied considerably; however, similar quantities were leached from sets of both manufacturers. 2-Ethyl-1-hexanol was also found in extracts from each of the sets analyzed.     

Partition chromatographic separation of pesticide residues from fats.

A simple, rapid, and efficient partitioning column consisting of acetonitrile on Florisil has been developed for the separation of pesticides from fish, beef, and butter fat. The efficiency of the cleanup column is between 97 and 100%. Nine pesticides having partition coefficients between n-hexane and acetonitrile of less than or equal to 0.05 were satisfactorily separated from fat with good recoveries. When the column was used to clean up temephos in a fish extract, 99.91% of the fat was eluted with 20 ml n-hexane with no loss of the pesticide.     

Thin layer chromatographic detection and indirect gas chromatographic determination of three carbamate pesticides.

Carbamate pesticide residues are extracted from vegetables and fruits with methylene chloride. The extracts are spotted on silica gel plates and the pesticides are detected by an enzymatic inhibition technique. For quantitative determination, aliquots of the methylene chloride extracts are evaporated to dryness in a rotary evaporator. After the residues are dissolved in ethanol, 0.5N NaOH is added in the hydrolysis step. To remove a number of possible interferences the hydrolyzed phenols are steam-distilled and treated with 1-fluoro-2,4-dinitrobenzene and/or 4-chloro-alpha,alpha,alpha-trifluoro-3,5-dinitrotoluene to form the ether derivatives. Efficiency in the conversion of the phenolic moieties to the phenyl ethers is about 100%. The resulting electron-capturing derivatives enable the carbamate pesticides to be detected in vegetables and fruits at the 0.05 ppm level. Recoveries of 90-94% were obtained from vegetables and fruits fortified with 0.5-2.0 ppm carbaryl, Mesurol, and propoxur.     

Capillary gas chromatographic detection of invert sugar in heated, adulterated, and adulterated and heated apple juice concentrates employing the equilibrium method.

The equilibrium method is introduced for the detection of invert sugar addition to apple juice. The method consists of a pre-equilibration of the sample with dry pyridine at 50 degrees C for 20 min followed by the addition of trimethylsilylimidazole and heating at 75 degrees C for 40 min. The resulting derivatized carbohydrates are then analyzed by capillary gas chromatography. This method was successfully used by independent laboratories to distinguish heated pure, intentionally adulterated (with invert sugar), and intentionally adulterated and then heated apple juice concentrates. The equilibrium method was shown to give significantly lower coefficients of variation for this sample set when compared to the original capillary gas chromatographic method. In addition, these results indicate that it may also be an effective method for the detection of medium invert sugar, depending on the level of the fingerprint oligosaccharides in this sweetener.     

Liquid chromatographic determination of propranolol hydrochloride in various dosage forms

A simple and rapid liquid chromatographic method is described for the determination of propranolol hydrochloride in pharmaceutical preparations. The separation was achieved on a reverse-phase octylsilane (C8) column by using a mobile phase composed of a mixture of 0.5 g dodecyl sodium sulfate in 18 mL (0.15 M) H3PO4 plus 90 mL methanol, 90 mL acetonitrile, and 52 mL water. Detector response was linear for 0.03-3.1 mg/mL of propranolol. Recoveries from synthetic mixtures ranged from 99.6 to 101.7%. The results obtained by the proposed method were similar to those obtained by the USP XXI method.     

Electron capture gas chromatographic determination of thiabendazole in yams

A method for the determination of thiabendazole in yams is reported. The method consists of extracting the chopped crops with ethyl acetate followed by distilled water wash and liquid liquid extraction of the chemicals into dilute hydrochloric acid. The sample is further cleaned up by making the aqueous acidic fraction alkaline and repartitioning into ethyl acetate. The determination step includes evaporation of the cleaned up extract to dryness, derivatization with pentafluorobenzoyl chloride, and determination of the derivatized thiabendazole in acetone by gas chromatography with electron capture detection.     

Isolation and gas chromatographic determination of chlorsulfuron in milk

A method for the isolation and gas chromatographic determination of chlorsulfuron in milk is presented. Blank or chlorsulfuron-spiked milk samples were blended into C-18 (octadecylsilyl derivatized silica, ODS) packing material. A column made from the C-18/milk matrix was washed with hexane after which chlorsulfuron was eluted with dichloromethane (DCM). The DCM eluate contained chlorsulfuron which was free from interfering co-extractants when analyzed by gas chromatography utilizing a nitrogen/phosphorus detector. Chlorsulfuron was found to undergo a thermally induced decomposition to give 2-amino-4-methoxy-6-methyl-1,3,5-triazine, which was detected and quantitated by this method. Standard curves for these analyses were linear (r = 0.992 +/- 0.004, n = 5), with an average percentage recovery of 91.6 +/- 10.8%, over the concentration range examined (62.5-2000 ng/mL). The inter- and intra-assay variabilities were 11.6 +/- 7.5% and 6.2%, respectively.     

小叶女贞果实花青素组分鉴定及色谱纯化技术

为提高小叶女贞果实的食用、药用价值,该文系统研究了果实中花青素种类构成及提取物的制备技术。试验采用紫外可见光谱法、高效液相色谱-质谱串联法、酸水解制备苷元等技术对小叶女贞果实花青素含量、单体种类进行了测定,并借助提取、萃取、柱层析等技术研究了花青素提取物的分离纯化过程。研究结果如下:测得每100 g小叶女贞果实中含花青素总量为(499±18.42)mg,从中鉴定出2种花青素单体,分别为矢车菊素-3-O-葡萄糖苷和牵牛花色素-3-O-葡萄糖苷,并以后者为主要存在形式;获得了纯天然、简单易行的花青素提取物制备技术,主要包括酸化乙醇提取、乙酸乙脂萃取、Amberlite XAD-7HP大孔树脂层析分离步骤,最终制得的花青素提取物纯度为35%、得率为0.6%。该研究为后期制备高纯度牵牛花素-3-O-葡萄糖苷单体提供了良好原料基础,为深入研究小叶女贞果实花青素功能活性及其在食品、药品领域潜在应用提供了参考。