Volatile Flavor Components of Dried Bonito (Katsuobushi) II. From Neutral Fraction

The aqueous extract of dried bonito (Katsuobushi) was distilled under reduced pressure. The resulting distillate with diethyl ether and the extract was separated into acidic, phenolic, basic and neutral fractions. The neutral fraction was further fractionated into ten sub-fractions by silica gel column chromatography. All these sub-fractions were analyzed by gas chromatography and gas chromatography-mass spectrometry.

One hundred and sixty-five compounds were identified and 12 compounds were tentatively identified from the neutral fraction. Among them, 111 compounds were newly identified as flavor components of Katsuobushi.     

Reaction between Nitrite and Low Salt-soluble Diffusible Fraction of Meat. Some Compounds Influencing Nitrite Depletion and Producing Unidentified-N Compounds

A low salt-soluble, diffusible fraction of meat was separated into basic, neutral and acidic fractions. Investigations were then conducted on the reaction of each fraction with nitrite and on the recovery of the added nitrite.

The basic fraction shared the lowest ability to decompose nitrite and had 88% recovery of added nitrite-N. Hypoxanthine was one of the main components that affected the recovery in the basic fraction. In the neutral fraction, 42% of nitrite was decomposed, and the recovery of the nitrite was 107%. In the acidic fraction, more than 80% of nitrite was decomposed and 30% was converted to unidentified-N compounds. Among endogenous acidic substances tested, cysteic acid showed the highest ability to decompose nitrite, accompanying the production of unidentified-N compounds.     

Mutagenicity spectra in bacterial strains of airborne and engine exhaust particulate extracts.

The mutagenicity spectra of the organic extracts of both airborne particulate matter and diesel and gasoline soot particles were determined using a battery of 9 bacterial strains of different genetic specificity. The assays with crude extracts and with fractionated acidic, neutral and basic components revealed striking differences in the patterns of mutagenic responses produced by each of the complex mixtures investigated. The mutagenicity of air particulate matter was shown to depend mainly on direct-acting acidic and neutral compounds, with a lesser contribution of basic promutagens which required exogenous metabolic activation by liver S9. The assays with a diesel soot extract indicated the prevailing contribution of direct-acting acidic and neutral compounds, and suggested an important role also for nitro derivatives other than nitropyrenes. The gasoline exhaust was characterized by powerful promutagenic compounds, belonging to either the acidic, neutral or basic fractions. The implications of these results are discussed with respect to the contribution of engine exhausts to air pollution, and the possible use of mutagenicity spectra in the analysis of environmental complex mixtures.     

Volatile Flavor Compounds of Boiled Buckwheat Flour

Volatile flavor compounds of boiled buckwheat flour were collected using a Likens and Nickerson apparatus type and were fractionated into acidic, weak acidic, basic and neutral fractions. The neutral fraction was further fractionated by silica gel column chromatography. All fractions were analyzed by gas chromatography and gas chromatography-mass spectrometry.

Two hundred and nine compounds were thus identified. Among them, 168 were newly identified as volatile flavor compounds of buckwheat flour; special note is made of 2-(1′-ethoxyethyl)pyrazine and 2-(1′-ethoxyethyl)-5-methylpyrazine being found which are compounds previously unreported in any literature.     

Possible role of hydrogen cyanide in chemical evolution investigation of the proposed direct synthesis of peptides from hydrogen cyanide

The compounds formed upon oligomerization of cyanide in aqueous solution have been separated into acidic, basic, amphoteric and neutral fractions. Urea is the major constituent of the neutral fraction and oxalic acid is present in the acidic fraction. The oligomerization mixtures isolated in the acid, basic and amphoteric fractions consist of low molecular weight substances which yield amino acids on acid hydrolysis. Citrulline has been identified as a major amino acid released on acid hydrolysis of the product mixture. No amino acids are released from the oligomerization mixture by pronase or carboxypeptidase A. This catalyzed hydrolysis demonstrates that this system does not contain compounds with peptide links. The oligomerization products are susceptible to oxidizing agents but are affected little by reducing agents.     

Volatile Components of Roasted Shrimp

The components of the strong and favorable aroma obtained from roasted shrimp were investigated. The aroma concentrate of roasted shrimp was isolated by combining the dichloromethane extract and carbon dioxide distillation methods. The concentrate was fractionated into acidic, basic and neutral fractions. Each fraction was analyzed by combined gas chromatography-mass spectrometry. Seventy-seven compounds were identified, among which isovaleric acid, alkyl pyrazines, isovaleramide, ketones and some sulfur-containing compounds were the main and characteristic constituents of the roasted shrimp aroma.

These constituents were compared with those of the volatiles of boiled shrimp, which was prepared by using simultaneous distillation and extraction methods in a modified Likens and Nickerson’s apparatus.     

Fractionation and Identification of Volatile Compounds in Patis,a Philippine Fish Sauce

Commercial Philippine fish sauce (Patis) was steam-distilled and the distillate was fractionated into four fractions, neutral, basic, acidic and phenolic and each fraction was analyzed using gas chromatography and gas chromatography-mass spectrometry.

As a result of this study, a total of 66 compounds were identified in Patis, 14 of which were only a tentative identification. Out of these 66 identified compounds, 40 identified compounds have not been reported in previous studies on fish sauces. These identified compounds include 19 acids, 14 alcohols, 12 nitrogen containing compounds, 5 esters, 3 sulfur containing compounds, 1 phenol, 3 carbonyls, 7 hydrocarbons and 2 others. In the acidic fraction, 5 acids were considered major constituents which accounted for about 98% of the total acids. n-Butanoic acid was found to be the most abundant accounting for about 50% of the total acids.     

Volatile Flavor Components of Dried Bonito (Katsuobushi). I. On Basic,Acidic and Weak Acidic Fractions

The aqueous extract of dried bonito (Katsuobushi) was distilled by using a thin film evaporator. The resulting distillate was extracted with diethyl ether, and the extract was separated into basic, acidic, weak acidic, and neutral fractions.

The basic, acidic, and weak acidic fractions were analyzed by gas chromatography and gas chromatography-mass spectrometry.

Seventy-four compounds, including 24 acids, 24 phenols, 8 pyridines, 12 pyrazines, 3 thiazoles, and 3 other compounds were identified. Thirty-six of these compounds were newly identified as volatile flavor compounds of Katsuobushi.     

Evidence for the presence of N2-(1,3-dicarboxypropyl)-L-amino acids in crown-gall tumors induced by Agrobacterium tumefaciens strains 181 and EU6

Using crown-gall tumors induced by Agrobacterium tumefaciens strains 181 and EU6 several unusual compounds have been isolated in a single fraction. Physicochemical analysis of the compounds in this fraction showed that they are N²-(1,3-dicarboxypropyl)-L-amino acids derived from neutral and acidic amino acids. Asparagine and glutamine derivatives (asparaginopine and glutaminopine) are the main components. Synthesis of asparaginopine from asparagine, α-ketoglutarate and NADPH, and degradation of asparaginopine to asparagine have also been demonstrated using enzyme prepared from Pinto bean strain 181 tumor.     

Chemical composition of acid–base fractions separated from biooil derived by fast pyrolysis of chicken manure

Our earlier investigations on the chemical composition of biooils derived by the fast pyrolysis of chicken manure revealed the presence of more than 500 compounds. In order to simplify this heterogeneous and complex chemical system, we produced four biooil fractions namely strongly acidic fraction A, weakly acidic fraction B, basic fraction C and neutral fraction D on the basis of their solubilities in aqueous solutions at different pHs. The yield (wt/wt.%) for fraction A was 3%, for fraction B 21.3%, for fraction C 2.4% and for fraction D 32.4%, respectively. The four fractions were analyzed by elemental analyses, Fourier Transform infrared spectrophotometry (FTIR), ¹H and ¹³C nuclear magnetic spectroscopy (NMR), and electrospray ionization mass spectrometry (ESI-MS). The major components of the four fractions were saturated and unsaturated fatty acids, N-heterocyclics, phenols, sterols, diols and alkylbenzenes. The pH separation system produced fractions of enhanced chemical homogeneity.     

Fractionation of Cigarette Smoke Components and Some Low Boiling Point Nitrogenous Compounds (I)

The chemical components of cigarette smoke produced in constant volume continuous smoking by an artificial smoking device were systematically fractionated into basic, acidic, neutral, phenolic, carbonyl and mercuric chloride-precipitable compounds. From the low boiling basic fractions ammonia, methylamine and ethylamine were qualitatively identified by paper chromatography, and pyridine and nicotine were isolated and identified by elementary analyses, mixed examinations and infrared spectra. An unidentified substance having elementary analysis values of C, 29.40; H, 1.93; N, 22.40 as picrate (m.p. 250°C, dec.) was isolated.     

Genotoxicity and PAC analysis of particulate and vapour phases of environmental tobacco smoke

Samples of indoor air were collected from an office room (88 m3) both before smoking and during experimental smoking of 96 cigarettes by 10 persons within 6 h. The particulates were collected on glass-fibre filters and the vapour-phase compounds on XAD-2 resin. The samples were extracted with acetone and analysed quantitatively for polycyclic aromatic compounds and qualitatively with GC-MS. The extracts of filters and XAD-2 resins were fractionated into neutral/acidic and 2 basic (strong and weak bases) fractions; all these fractions were tested with the sister-chromatid exchange (SCE) assay in Chinese hamster ovary (CHO) cells and with the Salmonella/microsome test (strain TA98). Total concentrations of PAC were 205 ng/m3 in the background sample and 1207 ng/m3 after contamination by cigarette smoking. The total PAC concentrations were 4-6 times higher in the vapour phase than in the particulate phase. The fractions of the particulate samples collected before smoking showed mainly marginal genotoxic activity, whereas after smoking their genotoxicity increased dramatically. The fractions of the vapour phase samples were not genotoxic before smoking, but after smoking the neutral/acidic and strong basic fractions induced responses in both assays. The SCE assay was more sensitive towards the vapour-phase mutagens of environmental tobacco smoke (ETS). The relative responses of the two basic fractions, whereas the fraction containing neutral and acidic compounds was the most potent in the SCE assay. In the Salmonella test, the mutagenic activity was mainly detected with metabolic activation, while the induction of SCE in CHO cells was also seen without an exogenous metabolic activation system.     

Bioassay-directed chemical analysis and detection of mutagenicity in ambient air of the coke oven

In the present study, we summarize the results of studies on the mutagenic potential of the main fractions and subfractions of extractable organic material (EOM) in the ambient air at the workplaces of the coke oven. The objective of our experiments was to apply the Bioassay-Directed Chemical Analysis (with the use of the Ames test) for the identification of the differences in the mutagenicity of these fractions, in relationship to the complex mixture of EOM in occupational air. From the evaluation of results, it is possible to deduce the following conclusions: (1) The comparison of the mutagenicity in the main fractions (basic, acidic, neutral) demonstrates the existence of differences in mutagenic potential. Of the total mutagenicity, 20.4% is in the basic fraction, 25.4% in the acidic fraction and 54.2% in the neutral fraction. (2) In general, 90.1% of the mutagenicity found in the basic, acidic and neutral fractions together was associated with the requirement of metabolic activation in vitro (+S9). In the case of the neutral fraction, it was 51.8%. (3) These results also suggest that frameshift mutations are the major component (53.8%) of the total mutagenicity of the main fractions. (4) With regards to the mutagenicity of organic compounds in the neutral fraction it appeared that genotoxicants of its subfractions (slightly and moderately polar and aromatic) play the main role. Carcinogenic aromatic hydrocarbons (PAH) and genotoxic nitrocompounds play an important role as determinants of the mutagenic potential of complex mixtures of harmful compounds in ambient air. This is confirmed first by the results of short-term bacterial tests.     

A strategy for discovering biologically active compounds with high probability in traditional Chinese herb remedies: an application of saiboku-to in bronchial asthma.

A novel strategy for discovering biologically active components in traditional Chinese herb remedies was performed from a pharmacokinetic view. The hypothesis was that the active compounds should appear in blood and urine with appropriate blood concentrations and urinary excretion rates after the administration of herbal-extract mixtures. In this research, we applied our procedures to Saiboku-To, one of the most popular Chinese herbal medicines in Japan. Consisting of 10 different plant extracts, it is used for the treatment of bronchial asthma. The analytical method adopted was a rapid-flow fractionation (RFF) for extraction-fractionation of lipophilic components in urine followed by silica-gel high-performance liquid chromatography (HPLC) equipped with a multichannel ultraviolet (uv) absorption detector. beta-D-Glucuronidase-treated urine samples collected before and after the administration of Saiboku-To to healthy and asthmatic subjects were treated with the RFF apparatus to afford three pH-dependent fractions: strongly acidic (S), weakly acidic (W), and neutral (N). HPLC of these fractions, monitored by the multichannel uv detector, showed three new peaks in the postadministrative urine: one in the N fraction, two in the W fraction, and none in the S fraction. A compound in the N fraction was identified with authentic magnolol, a major component in Magnolia officinalis. Two compounds in the W fraction were identified by comparison with authentic samples as 8,9-dihydroxydihydromagnolol and liquiritigenin, metabolites previously isolated from M. officinalis and Glycyrrhiza glabra, respectively.     

Metabolism of DL-Tryptophan-3-14C by the Fruitlets of Citrus aurantifolia

Developing lime fruit [Citrus aurantifolia (Christm.) Swingle] was supplied with dl-tryptophan-3-¹⁴C in a special medium. An incubation period of six hours was sufficient for the radioactivity to reach an equilibrium between the fruit tissue and the incubation medium. Analyses of the fruit tissue and the medium for acidic and neutral metabolites of tryptophan indicated the existence of indolic products. The auxin indole-3-acetic acid (IAA) was identified among the products by dry column chromatography and biological assay. Other acidic metabolites included indolepyruvic acid and an unidentified material. Neutral metabolites included indolealdehyde, indoleacetaldehyde, and two unidentified compounds. Biological activity in the Avena curvature test was obtained from extracted compounds which corresponded to IAA and indolepyruvic acid in the acidic fraction and indoleacetaldehyde in the neutral fraction. Radioactive tryptophan was also found in both the acidic and the neutral fractions due to its amphoteric nature. The experiment demonstrated the conversion of tryptophan to its indolic metabolites, including indole-3-acetic acid, in this Citrus tissue.     

Identification of an Acidic Dipeptide, β-Aspartylglycine, in the CNS of Aplysia

Abstract: A novel dipeptide, (β-aspartylglycine (β-DG), has been isolated from tissues of the marine gastropod mollusc Aplysia californica. This compound was detected only in Aplysia and not in other molluscs, such as Helix or Mercenaria , or in lobster or frog. Among the Aplysia tissues, the highest levels of β-DG were in nervous tissue and in the reproductive tract. β-DG was assayed by HPLC as the o -phthaldialdehyde derivative and found to be present in all individual, identified neurons at a concentration of approximately 40 pmol/μg protein. The peptide was identified as β-DG by gas chromatography-mass spectrometry (GCMS) using trimethylsilyl derivatives prepared before and after acid hydrolysis. It was further characterized as the β-isomer by TLC, including R_f, atypical blue-gray color with ninhydrin, and a violet color with Cu²⁺-ninhydrin. A fractionation scheme is described whereby acid-soluble tissue constituents can be divided into acidic, neutral, and basic components using mini ion-exchange columns. This partial purification prior to TLC analysis was necessary to remove compounds that interfered with the isolation of β-DG.     

Collection and Identification of Allelopathic Compounds from the Undisturbed Root System of Bigalta Limpograss (Hemarthria altissima)

Collection of allelopathic chemicals from the undisturbed plant root system is difficult because of their low concentrations and the high level of contaminants in growth media such as soil. A new approach for the continuous trapping of quantities of extracellular chemicals from donor plants is described. Bigalta limpograss (Hemarthria altissima), a tropical forage with allelopathic activities, was established in sand culture. Nutrient solution was circulated continuously through the root system and a column containing XAD-4 resin. Extracellular hydrophobic metabolites were selectively adsorbed by the resin, while inorganic nutrients were recycled to sustain plant growth. Columns were eluted with methanol and the eluate separated into neutral, acidic, and basic fractions. Bioassays of trapped root exudates using lettuce seed combined with paper and thin layer chromatography showed that the inhibitors were mainly phenolic compounds. The active neutral fraction was methylated and analyzed by gas chromatography-mass spectrometry. Twelve compounds were identified, with two additional compounds tentatively identified. 3-Hydroxyhydrocinnamic, benzoic, phenylacetic, and hydrocinnamic acids were the major rhizospheric compounds with known growth regulatory activities.     

Urinary metabolites of polyamines in rats

Urinary metabolites of polyamines in rats were studied systematically by the intraperitoneal injection of radioactive polyamines. Urinary metabolites were fractionated into 4 fractions containing non-polar and acidic compounds, acidic and neutral ampholytes, basic ampholytes and polyamines. A large amount of radioactivity was detected in the fractions containing non-polar and acidic compounds and polyamines of urine of rats injected with radioactive putrescine, while in the case of the injection of radioactive spermidine or spermine a relatively large amount of radioactivity was found in the basic ampholyte fraction as well as in the polyamine fraction. Analysis of these fractions indicated that gamma-aminobutyric acid, N-monoacetylputrescine, 2(3)-hydroxyputrescine, putreanine, N-(3-aminopropyl)-4-aminobutyric acid (isoputreanine), spermic acid, N-(3-aminopropyl), N'-(2-carboxyethyl)-1,4-diaminobutane, and N-monoacetylspermidine A and B were excreted as urinary metabolites of the polyamines in addition to putrescine, spermidine and spermine.     

Shoyu (Soy Sauce) Flavor Components: Neutral Fraction

From the vacuum distilled volatiles of shoyu, a neutral fraction was obtained. Shoyu was also directly extracted with dichloromethane and then the extract was separated into ten (A~J) fractions. The J fraction was a neutral one. The J fraction was further separated into twelve fractions by liquid column chromatography. All fractions obtained were analyzed by gas chromatography and combined gas chromatography-mass spectrometry. Consequently, 142 components were identified, 82 of which have not been reported previously as volatile constituents of shoyu. The identified compounds were 37 hydrocarbons, 22 alcohols, 22 carbonyls, 22 esters, 12 furans, 6 sulfurous compounds, 1 pyrone, 5 phenols, 1 furanone, 1 acid, 1 lactone and 12 other compounds. From the results of quantitative analysis and organoleptic evaluation, phenylacetal-dehyde is considered to be most important in the neutral fraction.     

Lipids of Pinus halepensis pollen

The total lipids of Pinus halepensis pollen were separated into individual classes of neutral and polar lipids and the components of each class were identified and determined quantitatively. Free fatty acids, waxes and triacylglycerols were found as the main constituents of neutral lipids and phosphatidylcholine and phosphatidylethanolamine of polar lipids. Glycerylether derivatives were detected in neutral and polar lipid fractions. Free and esterified volatile fatty acids were also found in pollen and its neutral lipid fraction.