Ralph J. Stevenson

Application of headspace solid-phase microextraction to volatile flavour profile development during storage and ripening of kiwifruit

Kiwifruit volatile flavour compounds were evaluated with headspace solid-phase microextraction (SPME) as a sample concentration technique. Gas chromatography–mass spectrometry (GC–MS) was used for qualitative and semi-quantitative analysis after SPME. Components such as heptanal, ethyl hex-3-enoate, 6-methylhept-5-en-2-one, acetic acid, c/t-2-nonenal (mixture) and c/t-2-decenal (mixture), which were previously found only in kiwifruit juice, were detected by headspace SPME–GC–MS. Other compounds (pent-4-enal, t,t-nona-2,4-dienal, 2-nonanone, ethyl octanoate, butyrolactone and 2-propenyl butanoate), which had not been found previously, were identified. Flavour volatiles of kiwifruit were very sensitive to storage time and state of ripeness.

Headspace sampling of whey protein concentrate solutions using solid-phase microextraction

Abstract Whey protein concentrate (WPC) represents a potentially substantial source of nutritious, flinctional protein ingredient for many traditional and novel food products. Characterisation of WPC’s volatile profile plays an important role in the manipulation of its flavour perception for various applications. The combination of solid-phase microextraction (SPME) as a headspace sampling technique and gas chromatography/mass spectrometry for compound separation/identification have proven to be convenient and sensitive. An optimisation exercise of the headspace SME (HS-SPME) technique with WPC solutions is described: the effects of SPME fibre polarity, sampling time, temperature and equilibration period have been investigated.

Synthesis and Evaluation of Stable Bidentate Transition Metal Complexes of 1-(Chloromethyl)-5-hydroxy-3-(5,6,7-trimethoxyindol-2-ylcarbonyl)-2,3-dihydro-1H-pyrrolo[3,2-f]quinoline (seco-6-azaCBI-TMI) as Hypoxia Selective Cytotoxins

A series of metal complexes were prepared as potential prodrugs of the extremely toxic DNA minor groove alkylator 1-(chloromethyl)-5-hydroxy-3-[(5,6,7-trimethoxyindol-2-yl)carbonyl]-2,3-dihydro-1H-pyrrolo[3,2-f]quinoline (seco-6-azaCBI-TMI) and close analogues. The pyrrolo[3,2-f]quinoline cytotoxins were prepared from 2-methoxy-4-nitroaniline in a nine-step synthesis involving a Skraup construction of a quinoline intermediate, its appropriate functionalization, and a final radical cyclization. The metal complexes were prepared from these and the labile metal complex synthons [Co(cyclen)(OTf)(2)](+), [Cr(acac)(2)(H(2)O)(2)](+), and [Co(2)(Me(2)dtc)(5)](+). The cobalt complexes were considerably more stable than the free effectors and showed significant attenuation of the cytotoxicity of the latter, with IC(50) ratios (complex/effector) of 50- to 150-fold, and substantial hypoxic cell selectivity, with IC(50) ratios (oxic/hypoxic cells) of 20- to 40-fold. The cobalt complexes were also efficiently activated by ionizing radiation, with G values for loss of the compound close to the theoretical value for one-electron reduction of 0.68 micromol/J. This work extends earlier observations that cobalt cyclen complexes are suitable for both the bioreductive and radiolytic release of potent pyrrolo[3,2-f]quinoline effectors.

Modern analyses and binding studies of flavour volatiles with particular reference to dairy protein products

Various techniques are used to separate and isolate mixtures of volatile flavour compounds from sample matrices. These include headspace sampling (static and dynamic), distillation followed by liquid-liquid extraction, simultaneous distillation-extraction, solid-phase extraction and new methods of extraction such as solid-phase microextraction and membrane-based systems. After clean-up and concentration, most mixtures of volatiles are separated and analysed in gas chromatographs using open-tubular columns. Gas chromatographic injection techniques, columns and detectors are discussed. Mass spectrometry coupled with gas chromatography is a major method used to identify volatile flavour compounds. Establishment of the real flavour profiles perceived by humans is discussed with reference to some appropriate experiments. Volatiles can be lost from foodstuffs by oxidation, polymerisation, reactions with other components in the foodstuff and evaporation. Effective binding of flavour constituents is important during storage and transportation of foodstuffs. On the other hand, the release of off-flavours from a foodstuff is desirable before consumption or formulation with other foodstuffs. Binding experiments discussed include those in liquid systems, desiccation experiments with dry foods and microencapsulation.

Characterization of the bound volatile extract from baby kiwi (Actinidia arguta).

The glycosidically bound volatile fraction of baby kiwi ( Actinidia arguta ) was studied. Glycosidic precursors were isolated from juice by adsorption onto an Amberlite XAD-2 column. After enzymatic hydrolysis with Rapidase AR2000, the released aglycones were analyzed by GC-MS. Alcohols, terpenoids, and benzenoids were the most abundant compound classes. Aromatic compounds and norisoprenoids showed the highest concentrations. Major compounds were 2,5-dimethyl-4-hydroxy-3(2H)-furanone (Furaneol), benzyl alcohol, 3-hydroxy-β-damascone, hexanal, and (Z)-3-hexen-1-ol. Precursors of aroma compounds including benzoic acid, cinnamic acid, and coniferyl alcohol were also found. Eugenol, raspberry ketone, and 4-vinylguaiacol were identified for the first time in the fruit of an Actinidia species. The high concentration of 2,5-dimethyl-4-hydroxy-3(2H)-furanone in bound form (95.36 μg/kg) is particularly interesting and justifies further investigation.

Hypoxia-Activated Prodrugs: Substituent Effects on the Properties of Nitro seco-1,2,9,9a-Tetrahydrocyclopropa[c]benz[e]indol-4-one (nitroCBI) Prodrugs of DNA Minor Groove Alkylating Agents

Nitrochloromethylbenzindolines (nitroCBIs) are a new class of hypoxia-activated prodrugs for antitumor therapy. The recently reported prototypes undergo hypoxia-selective metabolism to form potent DNA minor groove alkylating agents and are selectively toxic to some but not all hypoxic tumor cell lines. Here we report a series of 31 analogues that bear an extra electron-withdrawing substituent that serves to raise the one-electron reduction potential of the nitroCBI. We identify a subset of compounds, those with a basic side chain and sulfonamide or carboxamide substituent, that have consistently high hypoxic selectivity. The best of these, with a 7-sulfonamide substituent, displays hypoxic cytotoxicity ratios of 275 and 330 in Skov3 and HT29 human tumor cell lines, respectively. This compound (28) is efficiently and selectively metabolized to the corresponding aminoCBI, is selectively cytotoxic under hypoxia in all 11 cell lines examined, and demonstrates activity against hypoxic tumor cells in a human tumor xenograft in vivo.

A study of volatile “trapping” in spray dried whey protein concentrate by “crushing” and/or vacuuming, and detection by solid-phase microextraction/gas chromatography/mass spectrometry

Abstract Flavour is an important quality aspect of spray dried whey protein concentrate (WPC). A blander flavour of WPC would find greater acceptance in a wide range of other combined foods. The objective of this study was to investigate the extent of physical “trapping” or “binding” of volatiles during powder formation in spray drying. Two simple procedures were used: (1) “crushing” and (2) subjecting WPC powder to vacuuming. Scanning electron microscopy images were used to observe the structure of the “original” WPC and treated WPC particles. Solid-phase microextraction followed by gas chromatography/mass spectrometry, and sensory evaluations of both powder and liquid samples of WPC, were used to demonstrate that although vacuuming and/or “crushing” WPC powder certainly liberate some aroma/odour-impact constituents, indicating a certain degree of volatile trapping by the physical structure, others may require the powder to be “crushed” more effectively, or dissolved in water, before a large percentage of volatiles are liberated. Thus, it appears that flavour volatiles are entrapped (thus supporting the selective diffusion theory) or bound in different ways to WPC, and that some of them are weakly bound, possibly hydrophobically.

A facile synthesis of aryldihydropyrans using a Sonogashira-selenoetherification strategy

A high yielding and convenient synthesis of 2-aryl-2,5-dihydro-2H-pyrans is reported. Sonogashira coupling of 4-pentyn-1-ol and 5-hexyn-2-ol with a range of phenyl and naphthyl bromides affords the appropriate aryl acetylenic alcohols which then undergo ready reduction to the corresponding (E)-1-aryl-5-hydroxyalkenes. Subsequent selenoetherification of the (E)-5-hydroxyalkenes proceeds via a 6-endo-trig pathway cleanly affording the trans-2-aryl-3-phenylselenyltetrahydropyrans. Finally oxidative elimination of the selenides afforded the 2-aryl-2,5-dihydro-2H-pyrans which are convenient intermediates for the synthesis of C-aryl glycosides in that they contain a double bond which is available for further oxygenation.

Nitro seco Analogues of the Duocarmycins Containing Sulfonate Leaving Groups as Hypoxia-Activated Prodrugs for Cancer Therapy

The synthesis of 19 (5-nitro-2,3-dihydro-1H-benzo[e]indol-1-yl)methyl sulfonate prodrugs containing sulfonate leaving groups and 7-substituted electron-withdrawing groups is reported. These were designed to undergo hypoxia-selective metabolism to form potent DNA minor groove-alkylating agents. Analogues 17 and 24, containing the benzyl sulfonate leaving group and a neutral DNA minor groove-binding side chain, displayed hypoxic cytotoxicity ratios (HCRs) of >1000 in HT29 human cancer cells in vitro in an antiproliferative assay. Four analogues maintained large HCRs across a panel of eight human cancer cell lines. In a clonogenic assay, 19 showed an HCR of 4090 in HT29 cells. Ten soluble phosphate preprodrugs were also prepared and evaluated in vivo, alone and in combination with radiation in SiHa human tumor xenografts at a nontoxic dose. Compounds 34 and 39 displayed hypoxic log(10) cell kills (LCKs) of 1.78 and 2.71, respectively, equivalent or superior activity to previously reported chloride or bromide analogues, thus showing outstanding promise as hypoxia-activated prodrugs.

N-alkylated cyclen cobalt(III) complexes of 1-(chloromethyl)-3-(5,6,7-trimethoxyindol-2-ylcarbonyl)-2,3-dihydro-1H-pyrrolo[3,2-f]quinolin-5-ol DNA alkylating agent as hypoxia-activated prodrugs

A series of cobalt complexes of the potent DNA minor groove alkylator 1-(chloromethyl)-3-(5,6,7-trimethoxyindol-2-ylcarbonyl)-2,3-dihydro-1H-pyrrolo[3,2-f]quinolin-5-ol (seco-6-azaCBI-TMI) were prepared from a series of N-substituted cyclen ligands. The final N-substituted complexes carried formal overall charges ranging from +2 to -2 and showed limited improvements in solubility. They showed similar stabilities to that of the complex with the unsubstituted cyclen ligand, and large but variable attenuation of the cytotoxicity of the free alkylator (2-30-fold), compared to 150-fold for the unsubstituted ligand. However, they had oxic/hypoxic ratios (2-22-fold) comparable to that of the unsubstituted cyclen complex (5).