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The conversion of coconut oil and fusel alcohols, the sustainable and low-cost natural materials, to valuable products through Lipozyme TL IM-catalysed transesterification was investigated in a solvent-free system. Flavour esters, especially the octanoic acid esters (ethyl-, butyl-, isobutyl-, propyl- and (iso)amyl octanoate), were formed during the transesterification reactions. The reaction parameters were optimised as follows: molar ratio of 3.0:1 (alcohol to oil), enzyme loading of 15% wt/wt, reaction temperature of 23 °C, shaking speed of 130 rpm, and reaction time of 20 h. Further, the operational stability of Lipozyme TL IM was improved through washing with solvents, after which the enzyme could be continuously used for at least 100 h within 5 batches’ reactions without significant loss of activity. The results indicate that coconut oil and fusel alcohols can be effectively converted to valuable flavour esters through the Lipozyme TL IM-catalysed transesterification.

The viability of probiotic Lactobacillus rhamnosus HN001 was examined in fermented milk with and without Saccharomyces cerevisiae var. bayanus EC-1118. The yeast enhanced the viability of L. rhamnosus at 30C but not at 12C. High temperature (40C) was detrimental to yeast vitality and diminished its viability-enhancing effect. The yeast inoculated at 103 cfu/g multiplied to 106 cfu/g and enhanced the viability of L. rhamnosus. Viable yeast exerted maximum beneficial effects, although the cell-free yeast supernatant also enhanced the survival of L. rhamnosus. The addition of yeast after fermentation decreased its viability-enhancing effect. Removal of oxygen achieved the viability of L. rhamnosus that was comparable with that achieved in the presence of viable yeast. The results suggest that the improved viability of L. rhamnosus might involve yeast viability, oxygen removal and yeast metabolite(s). The findings would help develop novel microbial starter technology for ambient-stable fermented milks with live probiotics. Practical Applications Maintaining cell viability of probiotics is a scientific challenge, especially under ambient conditions. The findings of this study would lay the foundation for developing novel microbial starter technology for ambient-stable fermented foods with high cell counts of live probiotics, which entails stringent process and engineering control to ensure hygienic conditions and product safety. Benefits of such products would include: reducing distribution and storage costs by saving capital cost involved in chilled-chain distribution, storage and retailing; market penetration and growth where it was not possible due to a lack of or poor refrigeration; extending shelf life and increasing consumer convenience.

Lipase (Lipozyme TL IM)-mediated biocatalysis in a solvent-free system was effectively employed to convert coconut oil and fusel oil to flavor-active octanoic acid esters (mainly ethyl-, butyl-, isobutyl- and (iso)amyl octanoate). Response surface methodology based on five-level-three-factor central composite design was applied to investigate the effects of single and multiple interactive factors on the conversion. Results indicated that the studied independent parameters including reactant molar ratio, enzyme loading and shaking speed were all significant factors that affected the formation of octanoic acid esters during transesterification reaction. According to the developed polynomial model (R 2 = 0.947), the optimum transesterification conditions were determined as follows: molar ratio of 3.25:1 (alcohol/oil), enzyme loading of 16.7 % (wt%) and shaking speed of 160 rpm. Under the optimized reaction conditions an average conversion of 62.0 % was achieved which was in reasonable agreement with the predicted value of 64.1 % and about 9.4 mg of ethyl octanoate, 14.4 mg of isobutyl octanoate, 0.3 mg of butyl octanoate and 49.2 mg of (iso)amyl octanoate were produced per gram of coconut oil.

The lipase-catalyzed transesterification of coconut oil with fusel oil was investigated in this study. Various aroma-active octanoic acid esters, such as ethyl-, butyl-, propyl-, isobutyl- and (iso)amyl octanoate, were synthesized in addition to other esters. To determine the amount of flavor esters, a simple yet effective sample preparation method through refrigeration was developed to separate valuable fatty acid esters from monoglycerides, diglycerides and triglycerides. With this sample preparation method, gas chromatography coupled with mass spectrometry and flame ionization detector was applied to determine flavor esters in transesterified coconut oil with a total run time of 51 min. The levels of coefficient of variation were within 2% and the recovery rates were in the range of 89.5–100.0%, indicating good accuracy and high precision. The sample preparation and quantification method established appears to be useful for the determination of a wide range of fatty acid esters in oil samples. PRACTICAL APPLICATIONS Fatty acid esters, especially short-chain fatty acid esters, are important flavoring agents applied in the food industry. Using lipase as the biocatalyst to synthesize desirable flavor fatty acid esters from lipids and alcohols is gaining the interest of researchers, since the esters formed by this method are deemed as natural. The reaction equilibrium was achieved at around 20 h. For industrial production of flavor fatty acid esters, it is essential to monitor the amount of these target compounds in the reaction solutions. In this study we developed a simple and effective quantification method to determine the esters in transesterified coconut oil which may also be applicable for the analysis of esters in other oil samples such as lipid-derived biofuels.

Durian is a seasonal tropical fruit widely grown and highly prized in Southeast Asia. This is a first study to assess the transformation of durian constituents during fermentation with mono- and mixed-cultures of Saccharomyces cerevisiae and Williopsis saturnus with a view to developing a novel alcoholic beverage. A diversity of volatiles was produced especially alcohols and esters, while those volatiles initially present in durian, particularly the character-impact sulphur-containing odorants, were transformed. Most of the sulphur-containing compounds decreased with fermentation. 3-(Ethylthio)-1-propanol was produced initially then remained relatively stable, whereas ethyl thioacetate was formed initially then decreased in all culture types. The kinetic changes and concentrations of volatile compounds were similar in both the mixed-culture and the S. cerevisiae monoculture, except for higher amounts of alcohols in the mixed-culture. The W. saturnus monoculture, on the other hand, was the main producer of acetate esters. Sensory evaluation revealed a strong sulphury note retained in all fermentations but less unfavourable metallic and yeasty notes in the mixed-culture. The study suggests that biotransformation by yeasts can be effective in modifying the constituents and modulating the organoleptic properties of durian.

Summary The aim of the research was to study the volatile composition of mango wine fermented with two Williopsis yeast strains: Williopsis saturnus var. mrakii NCYC500 and W. staurnus var. suaveolens NCYC2586. Thirty terpenoids, twenty-six esters, ten alcohols, nine acids, seven aldehydes and ketones, two ethers, two phenols and one sulphur compound were identified in the mango wine fermented with strain NCYC500, while twenty-seven terpenoids, thirty esters, eleven alcohols, eight acids, eight aldehydes and ketones, three ethers, two phenols, one sulphur compound and one furan were detected in the mango wine fermented with the strain NCYC2586. The kinetic changes, final concentrations and odour activity values of major volatiles were compared between the two Williopsis yeast strains and also with other yeast reported in the literature. The results showed that Williopsis yeast strains NCYC500 and NCYC2586 were high producers of acetic acid and acetate esters, but low producers of medium- to long-chain fatty acids and their corresponding ethyl esters. Unlike mango wine fermented with Saccharomyces cerevisiae, most terpenoids derived from mango juice were retained in the resultant mango wine fermented with the two Williopsis yeast strains, suggesting the mango wine could retain the aromatic hints of fresh mango.

The effects of sequential inoculation of yeasts Williopsis saturnus var. mrakii NCYC2251 and Saccharomyces cerevisiae var. bayanus R2 on the volatile profiles of papaya wine were investigated at an inoculum ratio of 1000 ( W. saturnus ) to 1 ( S. cerevisiae ). Inoculation of S. cerevisiae after seven days' fermentation with W. saturnus produced papaya wine with more acetate esters and fruitiness than the control (simultaneous inoculation). However, inoculation of W. saturnus after two days' fermentation with S. cerevisiae resulted in most of the volatile composition being comparable to the control, except for the enhanced amount of ethyl esters. The first inoculated yeast dominated the fermentation. The study suggests that sequential inoculation of non- Saccharomyces and Saccharomyces yeasts at a certain inoculum ratio may be a valuable tool to manipulate yeast succession and to modulate the volatile profiles and organoleptic properties of papaya wine.

A simple and fast headspace-solid-phase microextraction (HS-SPME) method coupled with gas chromatography–mass spectrometry (GC–MS) was developed for the analysis of volatile compounds in transesterified coconut oil by applying solvent dilution. Solvent dilution conditions (solvent type and solvent amount) and HS-SPME sampling parameters (adsorption temperature and time) were optimised through monitoring the adsorption result of the selected volatiles (octanoic acid esters) in transesterified coconut oil samples. The incubation of methanol (800 μl)-diluted oil (200 μl) sample at 60 °C for 30 min led to the best result. The method was further validated by determining the calibration linear range, correlation coefficient (R2), accuracy, precision, limit of detection and limit of quantification through spiking standards into the blank matrix consisting of coconut oil and methanol. This method may also be applicable for detection and determination of volatile compounds in other transesterified oil samples.

Malaysian pink and white pomelo (Citrusgrandis (L.) Osbeck) peels were extracted with dichloromethane (DCM). Using GC-FID/MS and gas chromatography-olfactometry (GC-O), 50 and 47 aroma-active compounds were identified in pink and white pomelo peel extracts, respectively. The potency of each odorant in both pomelo peel extracts was determined by aroma extraction dilution analysis (AEDA). On the basis of flavor dilution (FD) factor, odor activity value (OAV) and relative flavor activity (RFA), main odorants in pink pomelo peel extract were trans-linalool oxide, cis,trans-2,6-nonadienal, octanal, citronellal, nonanal, trans-nerolidol, neryl acetate, indole, 6-methyl-5-hepten-2-one, terpinolene, trans-2-heptenal, perilla alcohol and nootkatone, while the most intense aromas in white pomelo peel extract were terpinolene, α-copaene, 4-terpinenol, trans-nerolidol, trans-linalool oxide, nootkatone, cis,trans-2,6- nonadienal, citronellol, elemol, carvyl acetate, 6-methyl-5-hepten-2-one, perilla aldehyde a...

A high throughput screening assay was developed based on the yeast chronological aging model and applied in evaluating several factors that mediate lifespan, including inoculum size, cellular state in nutrient-rich medium, and calorie level. Using our assay, we confirmed the previously reported genetic mimics of calorie restriction, including deletion of TOR1, SCH9 or RAS2. In contrast, deletion of SIR2 had longevity effect but seemed to produce only small beneficial effect on the response to calorie restriction. Overall, this new high throughput screening assay may facilitate identification of calorie restriction mimetics with a rapid and simple protocol, uncomplicated data analysis, and high sensitivity. In addition, the assay also provides quantifiable data including lag-time, growth rate, doubling time, and survival percentage.

The formation of volatile sulfur-containing flavor compounds from L-methionine catabolism by Saccharomyces cerevisiae EC-1118 was investigated in soymilk supplemented with this sulfur-containing amino acid with a focus on methionol (3-methylthio-1-propanol). Methionol produced from L-methionine metabolism of yeast in soymilk was extracted by solid-phase microextraction (SPME) and analyzed by gas chromatography-mass spectrometry (GC-MS). The effects of time (0 – 72 h), L-methionine concentration (0.05–0.20%), yeast extract concentration (0.00–0.30%), initial medium pH (4.5–6.9), and temperature (25–34°C) on the production of methionol were investigated. One-liter scale-up fermentation of soymilk by Saccharomyces cerevisiae EC-1118 was conducted for 48 h under static conditions with 0.15% of L-methionine, 0.10% of yeast extract, initial medium pH of 5.5, and temperature of 25°C, yielding 190 ppm of methionol. GC-olfactometry dilution analysis was conducted on extracts of the fermented soymilk, and no single...

Longan (Dimocarpus longan Lour.) is a type of tropical fruit. This study investigated the impact of L-leucine and L-phenylalanine on the volatile profiles of longan wine fermented with a non-Saccharomyces yeast Williopsis saturnus ssp. saturnus CBS254 with a view to enhancing longan wine aroma. The results revealed the ability of this yeast to enhance isoamyl alcohol and its ester isoamyl acetate (banana-like aroma), and 2-phenylethanol and its ester 2-phenylethyl acetate (rose-like aroma) with the addition of L-leucine and L-phenylalanine, respectively. The increased production of the targeted acetate esters appeared to be at the expense of other acetate esters except for methyl acetate, whereas the effects on the biotransformation of other volatiles were minimal. Therefore, these findings suggest that the combination(s) of an amino acid and yeast can be employed as a tool to manipulate the level of the targeted aroma compounds so as to impart a specific aroma or to add flavor complexity to longan wine. PRACTICAL APPLICATIONS Tropical fruits such as longan are often in oversupply during the peak season and some are wasted because of lack of adequate processing technologies. Thus, there is a need to add value to longan fruit via processing. Although drying as a form of processing is commonly applied to longan fruit, fermentation is not widely practiced to process and preserve this fruit because of lack of research and information. Longan wine fermentation using the conventional grape wine fermentation technology results in longan wine with inferior flavor. The aim of this study is to investigate the fermentation of longan juice using a non-Saccharomyces yeast Williopsis saturnus var. saturnus with addition of L-leucine and L-phenylalanine so as to modulate the formation of aroma compounds.

Three yeasts from the genus Williopsis (W. saturnus var. mrakii NCYC500, W. saturnus var. saturnus CBS254 and W. californica NCYC2590) were examined for their ability to ferment longan juice and to enhance formation of longan wine aroma compounds. The three yeasts showed similar growth kinetics and pH changes during fermentation. W. californica was the least sugar consumer. Many of the naturally occurring volatiles (ethyl esters, fatty acids, aldehydes, and terpenes) in the juice were partially or completely degraded. The three yeasts varied with their ability to produce and utilise volatiles. Esters were the major volatiles produced with some esters being catabolised while other esters remained stable. The amount of most alcohols increased while of aldehydes decreased. W. saturnus CBS254 was the best producer of ethyl acetate, isobutyl acetate, isoamyl acetate and 2-phenethyl acetate, whereas W. californica NCYC2590 was the highest producer of butyl acetate. Ethanol was produced in similar amounts by W. ...

The impact of fusel oil addition on volatile compounds formation in papaya wine fermented with yeast Williopsis saturnus var. mrakii NCYC2251 was studied with a view to enhancing papaya wine aroma production. Time-course papaya juice fermentations were carried out using W. saturnus var. mrakii NCYC2251 with fusel oil added (0, 0.1 and 0.5% v/v). Fermentation characteristics in terms of yeast growth, Brix and pH changes were similar for all fermentations except for those added with 0.5% (v/v) fusel oil. The addition of 0.5% (v/v) fusel oil inhibited yeast growth. A wide range of volatile compounds were produced during fermentation including acids, alcohols, esters and aldehydes with esters being the most abundant volatile compounds produced. The addition of 0.1% (v/v) fusel oil reduced the production of undesirable volatiles such as ethyl acetate and acetic acid, while increasing the desirable volatiles production such as ethanol and acetate esters. This study suggests that papaya juice fermentation with W. saturnus var mrakii NCYC 2251 together with a low concentration of added fusel oil can be another means of modulating papaya wine aroma compound formation.

The knowledge of the chemical compositions and sensory profiles of their peels and blossoms could lead to better understanding of two cultivars of Malaysian pomelo. This work was to determine the volatile compositions of Malaysian pomelo blossoms and peels through HS-SPME-GC/MS analysis and principal component analysis (PCA) approach as well as to identify their key aroma profiles by sensory evaluation. Pink and white pomelo blossoms contained similar volatiles, which mainly consisted of limonene, cis-β-ocimene, α-terpinene, linalool, methyl anthranilate, and indole. Their primary difference was in the concentration ratio between limonene and linalool. Major volatiles in pomelo peels were terpene hydrocarbons. Pink pomelo peel contained higher levels of aldehydes (e.g. octanal, decanal and citral), while some trace-level important compounds (e.g. β-sinensal, α-sinensal and nootkatone) were found only in white pomelo peel. Through sensory evaluation, both intact pomelo blossoms had similar aroma p...

The aim of this research was to study the feasibility of biosynthesising natural aroma-active 2-phenylethyl esters in coconut cream via biocatalysis. Five commercial fungal lipases (Palatase 20 000L, Lipase AYS “Amano”, Lipase A “Amano” 12, Piccantase A and Piccantase AN) were screened for their ability to produce 2-phenylethyl hexanoate and 2-phenylethyl octanoate from coconut cream supplemented with natural 2-phenylethanol. Palatase 20 000L and Lipase AYS “Amano” showed the greatest potential for the synthesis of 2-phenylethyl esters. The biosynthetic reaction by the lipase Palatase 20 000L was further studied under different temperature, pH, enzyme and substrate concentrations, with maximum ester synthesis obtained at 9% v/v 2-phenylethanol and 2.15–8.60 mg enzyme protein/100 ml of reaction mixture. A temperature range of 30–53 °C and a pH range of 5.2–8.5 had little effect on the lipase Palatase activity, respectively. The ester-synthesising reaction appeared to proceed by both alcoholysis and esterification.

Summary Cofermentation of longan juice by mixed cultures of Saccharomyces cerevisiae var. bayanus EC-1118 and Williopsis saturnus var. saturnus CBS254 at two inoculation ratios (EC-1118:CBS254 = 1:100 and 1:1000 cfu mL−1) was performed to ascertain their impact on longan wine aroma compound formation. The results showed improved aroma compound profiles in the longan wine fermented with mixed yeasts in comparison with the longan wines fermented with single yeasts in terms of increased production of acetate esters, fatty acid ethyl esters, alcohols and carboxylic acids. The impact of cofermentation on longan wine aroma formation was affected by the ratio of S. cerevisiae EC-1118 to W. saturnus CBS254 with 1:100 cfu mL−1 being more effective. This research suggests that the inoculation ratio of mixed yeasts may be used as an effective means of manipulating longan wine aroma.

PurposeThe purpose of this paper is to assess the potential of papaya juice fermentation and to evaluate the kinetic changes of volatile organic compounds (VOCs) produced during fermentation by three commercial wine yeasts.Design/methodology/approachLaboratory‐scale fermentations were carried out in papaya juice using three commercial wine yeasts Saccharomyces cerevisiae var. bayanus strains EC‐1118 and R2, and S. cerevisiae MERIT.ferm. Brix, pH, optical density and yeast cell count were determined during fermentation. VOCs were analysed by headspace‐solid phase microextraction with gas chromatography‐mass spectrometry‐flame ionization detector (HS‐SPME‐GC‐MS/FID).FindingsDuring fermentation, the three wine yeasts grew actively and showed similar growth patterns. Changes in Brix and pH were similar among the three yeasts. A range of VOCs were produced during fermentation including fatty acids, alcohols, acetaldehyde, esters and acetoin. Esters were the most abundant VOCs produced. Some VOCs indigenous to the papaya juice such as benzaldehyde, β‐damascenone and benzyl isothiocyanate were consumed during fermentation. Some VOCs increased initially, and then decreased during fermentation. Overall, the profiles of VOC changes during fermentation were similar among the three yeasts with some differences observed.Originality/valueThe paper suggests that papaya fermentation with S. cerevisiae yeasts are likely to result in papaya wine with similar flavours. New approaches are required to produce papaya wine with distinct flavours and to enable differentiation of papaya wines by exploiting non‐Saccharomyces yeasts in conjunction with Saccharomyces yeasts and/or by adding selected nitrogen sources.

This study investigated the formation and utilization of volatile compounds during papaya juice fermentation by a mixed culture of Saccharomyces cerevisiae and Williopsis saturnus. Time-course papaya juice fermentations were carried out using pure cultures of S. cerevisiae var. bayanus R2 and W. saturnus var. mrakii NCYC2251 and a mixed culture of the two yeasts at a ratio of 1:1000 (R2:NCYC2251). Changes in S. cerevisiae cell population, Brix, sugar consumption and pH were similar in the mixed culture and in the S. cerevisiae monoculture. There was an early growth arrest of W. saturnus in the mixed culture fermentation. A range of volatile compounds were produced during fermentation including fatty acids, alcohols, aldehydes and esters and some volatile compounds including those initially present in the juice were utilized. The mixed culture fermentation of S. cerevisiae and W. saturnus benefited from the presence of both yeasts, with more esters being produced than the S. cerevisiae monoculture and more alcohols being formed than the W. saturnus monoculture. The study suggests that papaya juice fermentation with a mixed culture of S. cerevisiae and W. saturnus may be able to result in the formation of more complex aroma compounds and higher ethanol level than those using single yeasts.

Durian fruit is rich in volatile sulfur compounds (VSCs), especially thiols and disulfides, which contribute to its onion-like odor. After fermentation, these VSCs were reduced to trace or undetectable levels in durian wine. The possible reduction mechanism of these VSCs (especially diethyl disulfide and ethanethiol) was investigated in a modified buffer in the presence of sulfite at different pH. An interconversion between diethyl disulfide and ethanethiol was found to be dependent on the pH: the higher the pH, the higher production of ethanethiol. It is suggested that, during durian wine fermentation, disulfides endogenous to durian pulp might be firstly converted into their corresponding thiols in the presence of reductant sulfite formed by yeast. The produced thiols as well as the thiols endogenous to the durian pulp were then removed by the mannoproteins of yeast lees.

PurposeSpoilage due to yeast and mould growth is a major issue for yoghurt quality and shelf‐life. There is a need to develop natural alternatives to chemical preservation. The purpose of this paper is to determine the effectiveness of mycocinogenic yeast Williopsis saturnus var. saturnus as a biocontrol agent against spoilage yeasts and moulds in plain yoghurt.Design/methodology/approachYoghurts were prepared from reconstituted skim milk and were challenged with spoilage yeasts and moulds. The treatment contained the added mycocinogenic yeast and the control without. All yoghurts were incubated at 30○C. Yeast and mould growth were determined by observing gas formation and mould colony occurrence at regular intervals.FindingsW. saturnus var. saturnus inhibited growth of lactose‐fermenting and galactose‐fermenting yeasts (Candida kefir and Kluvyveromyces marxianus), and lactose non‐fermenting but galactose fermenting yeasts (strains of Saccharomyces cerevisiae and Saccharomyces bayanus). The yeast also inhibited growth of dairy moulds including Byssochlamys, Eurotium and Penicillium.Research limitations/implicationsThe inhibition of this mycocinogenic yeast against yeasts and moulds was dependent upon the concentration of the latter. Thus, hygiene and good manufacturing practice are essential in order to keep the contaminant load down and to ensure the effectiveness of the mycocinogenic yeast.Originality/valueThe use of mycocinogenic yeast to control spoilage yeasts and moulds in yoghurt is a novel approach with a potential to minimise yoghurt spoilage and extend the shelf‐life of yoghurt.

Production of sulphur flavor concentrate by yeast fermentation was investigated using dairy media (milk and cream) spiked with L-methionine. The yeasts studied included so-called dairy yeasts Candida kefyr, Kluyveromyces marxianus, Debaryomyces hansenii, Geotrichum candidum, and Yarrowia lipolytica as well as wine yeasts Saccharomyces cerevisiae and Saccharomyces bayanus. Methionol was produced as the predominant volatile sulphur flavor compound. Other volatile sulphur flavor compounds produced were dimethyl disulphide, S-methyl thioacetate, 3-methylthio-1-propanoic acid, methional, 3-methylthio-1-propene, and ethyl 3-methylthio-1-propanoate. In addition, numerous other volatile flavor compounds were produced, including ethanol, branched-chain alcohols, aromatic alcohol, ethyl esters, branched-chain esters, branched-chain fatty acids, and branched-chain aldehydes. The sensory attribute of a methionol sulphur flavor concentrate in cream is concentration-dependent, ranging from a savory-slight potato note t...

PurposeThe purpose of this paper is to investigate in situ production of aroma‐active esters in dairy foods so as to improve flavour and to produce fruity flavour concentrate.Design/methodology/approachLipase, ethanol or bacterial cultures are added to dairy media (milk, cream or cheese) and incubated for a period of time (from hours to months). Samples are then taken and analysed for aroma‐active esters using gas chromatography (GC) or gas chromatography‐mass spectrometry (GC‐MS).FindingsAnalyses of samples show that significant levels of ethyl esters of fatty acids are produced in milk, cream, enzyme‐modified cheese and natural cheese. All the dairy foods possess an intense pleasant fruity aroma.Originality/valueThis is a natural way to generate fruity flavours in dairy foods to enhance flavour and thus, consumer acceptance. The fruity flavour concentrate can also be used as a flavouring ingredient in dairy and non‐dairy food applications. Natural pure esters may also be extracted, separated and concentrated for wider flavour and fragrance applications. This approach may provide a cost‐effective solution to the increasing surplus of milk fat.

Some strains of the yeast Debaryomyces hansenii are known to be antagonistic toward moulds. In this study, we describe the inhibitory effects of a dairy strain of D. hansenii as a biocontrol agent against a number of dairy moulds in plain yoghurt and cheese under non-refrigerated conditions. This antagonistic yeast showed inhibition of growth of the following dairy moulds: Aspergillus sp., Byssochlamys fulva, Byssochlamys nivea, Cladosporium sp., Eurotium chevalieri, Penicillium candidum and Penicillium roqueforti. However, the inhibitory effect of this antagonistic yeast against dairy moulds is dependent upon the concentration of the moulds: the lower the concentration, the more effective the yeast. The findings of this study have implications for both cheese maturation and dairy biopreservation. Good manufacturing practice and hygiene to keep the contaminant load down is essential for the prevention of dairy spoilage.

This study represented for the first time the impact of malolactic fermentation (MLF) induced by Oenococcus oeni and its inoculation strategies (simultaneous vs. sequential) on the fermentation performance as well as aroma compound profile of durian wine. There was no negative impact of simultaneous inoculation of O. oeni and Saccharomyces cerevisiae on the growth and fermentation kinetics of S. cerevisiae as compared to sequential fermentation. Simultaneous MLF did not lead to an excessive increase in volatile acidity as compared to sequential MLF. The kinetic changes of organic acids (i.e. malic, lactic, succinic, acetic and α-ketoglutaric acids) varied with simultaneous and sequential MLF relative to yeast alone. MLF, regardless of inoculation mode, resulted in higher production of fermentation-derived volatiles as compared to control (alcoholic fermentation only), including esters, volatile fatty acids, and terpenes, except for higher alcohols. Most indigenous volatile sulphur compounds in durian were decreased to trace levels with little differences among the control, simultaneous and sequential MLF. Among the different wines, the wine with simultaneous MLF had higher concentrations of terpenes and acetate esters while sequential MLF had increased concentrations of medium- and long-chain ethyl esters. Relative to alcoholic fermentation only, both simultaneous and sequential MLF reduced acetaldehyde substantially with sequential MLF being more effective. These findings illustrate that MLF is an effective and novel way of modulating the volatile and aroma compound profile of durian wine.

Modulation of coffee aroma via the biotransformation/fermentation of different coffee matrices during post-harvest remains sparingly explored despite some studies showing their positive impacts on coffee aroma. Therefore, this is an unprecedented study aimed at modulating coffee aroma via the fermentation of green coffee beans with a food-grade fungus Rhizopus oligosporus. The objective of part I of this two-part study was to characterize the volatile and non-volatile profiles of green coffee beans after fermentation. Proteolysis during fermentation resulted in 1.5-fold increase in the concentrations of proline and aspartic acid which exhibited high Maillard reactivity. Extensive degradation of ferulic and caffeic acids led to 2-fold increase in the total concentrations of volatile phenolic derivatives. 36% of the total volatiles detected in fermented green coffee beans were generated during fermentation. Hence, the work presented demonstrated that R. oligosporus fermentation of green coffee beans could induce modification of the aroma precursors of green coffees.

Chemical compositions of durian wines fermented with Metschnikowia pulcherrima Flavia, Torulaspora delbrueckii Biodiva, Pichia kluyveri FrootZen and Kluyveromyces thermotolerans Concerto were investigated.Sucrose was not utilized by M. pulcherrima and P. kluyveri, resulting in little formation of ethanol (0.3-0.5%, v/v), while about 7% ethanol was produced by the other two yeasts. Volatiles such as esters and sulfur-containing compounds were synthesized or catabolized and distinctive differences existed among yeasts. Larger amounts of higher alcohols and ethyl esters were detected in wines fermented by T. delbrueckii and K. thermotolerans, whereas M. pulcherrima and P. kluyveri produced more acetate esters such as ethyl acetate (1034.43 and 131.05 mg L(-1) respectively) and isoamyl acetate (0.56 and 27.68 mg L(-1) respectively). Most endogenous sulfur volatiles such as disulfides declined to trace levels, but new ones such as thioesters were formed. Sulfur volatiles in wines fermented by T. delbrueckii accounted for 0.20% relative peak area (RPA), followed by K. thermotolerans (0.23% RPA), P. kluyveri (1.43% RPA) and M. pulcherrima (4.16% RPA).The findings showed that a more complex flavor could result from fermentation with different non-Saccharomyces yeasts and the typical durian odor would still remain.

The present study attempted to partially characterize and elucidate the viability-enhancing effect of a yeast strain Saccharomyces cerevisiae EC-1118 on a probiotic strain Lactobacillus rhamnosus HN001 under acidic conditions using a model system (non-growing cells). The yeast was found to significantly enhance (P < 0.05) the viability of the probiotic strain under acidic conditions (pH 2.5 to 4.0) by 2 to 4 log cycles, and the viability-enhancing effects were observed to be influenced by pH, and probiotic and yeast concentrations. Microscopic observation and co-aggregation assay revealed that the viability-enhancing effect of the yeast could be attributed to direct cell-cell contact co-aggregation mediated by yeast cell surface and/or cell wall components or metabolites. Furthermore, non-viable yeast cells killed by thermal means were observed to enhance the viability of the probiotic strain as well, suggesting that the surface and/or cell wall component(s) of the yeast contributing to co-aggregation was heat-stable. Cell-free yeast supernatant was also found to enhance the viability of the probiotic strain, indicating the presence of protective yeast metabolite(s) in the supernatant. These findings laid the foundation for further understanding of the mechanism(s) involved and for developing novel microbial starter cultures possibly without the use of live yeast for ambient-stable high-moisture probiotic foods.

Free fatty acids (FFAs) and esters derived from FFAs are important flavour compounds in cheese. Evidence is provided that esterases of lactic acid bacteria (LAB) catalyse not only hydrolysis of milk fat glycerides to release FFAs, but also synthesis of esters from glycerides and alcohols via a transferase reaction. The esterases of LAB prefer di- and monoglycerides for both hydrolysis and ester synthesis and are, in fact, alcohol acyltransferases that use both water (hydrolysis) and alcohol (alcoholysis) as acyl acceptors. Therefore, esterases of LAB can impact on both the lipolytic and ester flavours of cheese. The impact of esterases of LAB on cheese flavour can be controlled by manipulating the amount of the esterase, by regulating alcohol availability and/or by increasing the mono- and diglyceride composition of milk fat. In addition, esterification may play a role in ester synthesis in hard cheeses (e.g. Italian type) where water activity is low.

Esters of short-chain fatty acids are aroma-impact compounds found in fermented dairy products. These esters are responsible for fruity flavours that can be regarded either as a defect or as an attribute by the consumer. An understanding of the mechanisms of ester biosynthesis will enable control of the development of fruity flavours in fermented dairy products. The biosynthesis of flavour-active esters in dairy systems proceeds through two enzymatic mechanisms—esterification and alcoholysis. Esterification is the formation of esters from alcohols and carboxylic acids, whereas alcoholysis is the production of esters from alcohols and acylglycerols or from alcohols and fatty acyl-CoAs derived from metabolism of fatty acids, amino acids and/or carbohydrates. Alcoholysis is essentially a transferase reaction in which fatty acyl groups from acylglycerols and acyl-CoA derivatives are directly transferred to alcohols and is the major mechanism of ester biosynthesis by dairy lactic acid bacteria and yeasts.

Evidence was obtained that esterases purified from Streptococcus thermophilus ST1 and Lactococcus lactis subsp. cremoris B1079 are able to catalyse the synthesis of an ester (ethyl butanoate) in an aqueous environment via a transferase reaction (alcoholysis) in which fatty acyl (butyryl) groups from the glyceride (tributyrin) are transferred to the alcohol (ethanol). The esterases displayed transferase activity in which the acyl acceptor may be either water (hydrolysis) or alcohol (alcoholysis). The same enzymes were responsible for both activities. We propose that the esterases of St. thermophilus ST1 and Lc. lactis subsp. cremoris B1079 are alcohol acyltransferases that catalyse both hydrolysis and alcoholysis in an aqueous environment. This is the first report demonstrating ester synthesis by the esterases of these two lactic acid bacteria (LAB) via alcoholysis in an aqueous environment. Alcoholysis is the major mechanism of ester synthesis by the esterases of these two LAB in an aqueous environment.

The metabolism of amino acids by 22 starter and 49 non-starter lactic acid bacteria (LAB) was studied in a system consisting of amino acids and non-growing cells without added amino acceptors such as alpha-ketoglutarate. There were significant inter- and intra-species differences in the metabolism of amino acids. Some amino acids such as alanine, arginine, aspartate, serine and branched-chain amino acids (leucine, isoleucine and valine) were utilised, whereas other amino acids such as glycine, ornithine and citrulline were produced. Alanine and aspartate were utilised by some LAB and accumulated during the incubation of other LAB. Arginine was degraded not only by Lactococcus lactis subsp. lactis (the lactococcal subspecies known to catabolise arginine), but also by pediococci, heterofermentative lactobacilli (Lactobacillus brevis and Lb. fermentum) and some unidentified homofermentative lactobacilli. Serine was utilised predominantly by homofermentative Lb. paracasei subsp. paracasei, Lb. rhamnosus and Lb. plantarum. Of the LAB studied, Lb. brevis and Lb. fermentum were the most metabolically active, utilising alanine, arginine, aspartate, glutamate and branched-chain amino acids. Leuconostocs were the least metabolically active, showing little potential to metabolise amino acids. The formation of ammonia and acetate from amino acid metabolism varied both between species and between strains within species. These findings suggest that the potential of LAB for amino acid metabolism via non-transaminating reactions and endogenous transamination will impact both on the physiology of LAB and on cheese ripening, especially when transamination is rate-limiting in the absence of an exogenous amino acceptor such as alpha-ketoglutarate.

A fruity flavour note is traditionally regarded as a defect in cheese varieties such as Cheddar (Bills et al. 1965; McGugan et al. 1975; Horwood et al. 1987). However, fruitiness is an attribute of other cheese varieties such as Parmesan and Parmigiano Reggiano (Dumont et al. 1974; Meinhart & Schreier, 1986). It is well accepted that esters such as ethyl butanoate and ethyl hexanoate cause the fruity flavour described as apple-like or pineapple-like in raw milk and cheeses (Bills et al. 1965; Engels et al. 1997; Friedrich & Acree, 1998). The development of fruity flavour is often attributed to the esterification of free fatty acids and ethanol by esterases from lactic acid bacteria and psychrotrophic pseudomonads (Hosono et al. 1974; Morgan, 1976).

The relationship between coffee fermentation and coffee aroma is intricate and delicate at which the coffee aroma profile is easily impacted by the fermentation process during coffee processing. However, as the fermentation process in coffee processing is conducted mainly for mucilage removal, its impacts on coffee aroma profile are usually neglected. Therefore, this review serves to summarize the available literature on the impacts of fermentation in coffee processing on coffee aroma as well as other unconventional avenues where fermentation is employed for coffee aroma modulation. Studies have noted that proper control over the fermentation process imparts desirable attributes and prevents undesirable fermentation which generates off-flavors. Other unconventional avenues in which fermentation is employed for aroma modulation include digestive bioprocessing and the fermentation of coffee extracts and green coffee beans. The latter is an area that should be explored further with appropriate microorganisms given its potential for coffee aroma modulation.

Two of the three intracellular esterases identified in Streptococcus thermophilus were purified to homogeneity using ammonium sulphate fractionation and three chromatographic steps: anion exchange, hydrophobic interaction and gel filtration. The subunit molecular masses of esterases I and II were ∼34 and ∼60 kDa, respectively. The holoenzyme molecular masses of esterases I and II were ∼50 and ∼60 kDa, respectively, indicating that esterase I could be a dimer and that esterase II was a monomer. Phenylmethylsulphonyl fluoride inhibited the activity of both esterases, but to different degrees. Dithiothreitol, N -ethylmaleimide and EDTA strongly inhibited the activity of esterase I but significantly enhanced the activity of esterase II. Esterase I was active on p -nitrophenyl esters of the short-chain fatty acids from C 2 to C 10 and esterase II was active on p -nitrophenyl esters of the C 2 –C 6 fatty acids. For both enzymes, maximum activity was obtained with p -nitrophenyl butyrate (C 4 ). The K m values of esterase I on p -nitrophenyl esters of C 2 –C 8 fatty acids ranged from 6.7 to 0.004 m m and the corresponding V max values ranged from 8.12 to 1.12 μmol min −1 mg −1 protein. The N-terminal amino acid sequences of the two esterases also differed. The major esterase (I), accounting for ∼95% of the total esterase activity, was further characterized. Esterase I was also active against tributyrin (C 4 ), dicaproin (C 6 ) and monoglycerides of up to C 14 with maximum activity on monocaprylin (C 8 ). Decreasing pH (from 8.0 to 5.5), temperature (from 37° to 25°C) or water activity (from 0.99 to 0.80) considerably reduced the activity of esterase I, whereas increasing NaCl concentration up to 7.5% (w/v) markedly enhanced the activity of this enzyme. Esterase I may play a role in the development of cheese flavour with respect to lipolysis.

The excretion of citrulline, a precursor of carcinogenic ethyl carbamate, formed from arginine degradation by malolactic bacteria in wine is of toxicological concern. The arginine metabolism of resting cells of Lactobacillus buchneri CUC-3 and Oenococcus oeni Loll 1 was examined. The citrulline excretion rate was found to be linearly correlated to the arginine degradation rate. It was possible to calculate an arginine to citrulline conversion ratio which could be used to predict the amount of citrulline expected after the degradation of a known quantity of arginine. The conversion ratios determined in this study were similar to data calculated from other authors for fermentations in wine and ranged between 4.0% and 7.7%. Ribose, fructose and glucose inhibited the degradation of arginine in Lact. buchneri CUC-3, and inhibition of arginine degradation by glucose correlated with higher arginine to citrulline conversion ratios. The work presents new results of arginine metabolism in malolactic bacteria and gives starting points for investigations in wine.

Major commercially available strains for induction of malolactic fermentation in wine were examined for arginine metabolism in a resting cell system at wine pH with the aim of evaluating their ability to excrete and utilize citrulline, a precursor of carcinogenic ethyl carbamate (urethane). All strains tested excreted citrulline from arginine degradation. Citrulline was stored intracellularly during growth in arginine rich medium and was released upon lysis of the cells. All strains were found to degrade citrulline as a sole amino acid and some of them were able to reutilize previously excreted citrulline.

The formation of ethyl butanoate by non-growing cells of 22 starter and 49 non-starter dairy lactic acid bacteria (LAB) varied widely (0.4–310 units 100 mg-1 dry weight cells) and was both species and strain dependent. Strains of the thermophilic starter Streptococcus salivarius subsp. thermophilus produced the highest levels of ethyl butanoate (an average of 156 units 100 mg-1 dry weight cells), while strains of the mesophilic starters Lactococcus lactis subsp. cremoris and Lactococcus lactis subsp. lactis gave a more moderate production of the ester (averages of 39 and 27 units 100 mg-1 dry weight cells, respectively). Non-starter LAB (lactobacilli, pediococci and leuconostocs) and propionibacteria varied widely in their ability to produce ethyl butanoate (0.4–160 units 100 mg-1 dry weight cells). The effect of physicochemical factors (pH, NaCl, water activity (aw) and temperature) on ethyl butanoate production was also investigated using Lactococcus lactis subsp. cremoris 2272 and Lactobacillus rhamnosus 2615. For both strains, ester formation increased by up to 175% when the pH was lowered from 5.8 to 4.0, but was drastically inhibited by up to 99% in the presence of NaCl and at reduced aw. Lowering the temperature from 30 to 13°C decreased ester production by 25% for both strains. The findings suggest that the normal cheese pH (∼5.0) and ripening temperature (∼13°C) are not critical factors in ester formation, but NaCl concentration and aw level are pivotal in determining ester formation.

The influence of reduced water activity (a w ) on lactose metabolism by Lactococcus lactis subsp. cremoris 2254 and 2272 was studied at different pH values. In control incubations (a w , 0.99) with nongrowing cells in pH-controlled phosphate buffer, the levels of carbon recovered as l -(+)-lactate were 92% at pH 6.1 and 5.3 and 78% at pH 4.5. However, the levels of recovery decreased to ∼50% at all pH values tested when the a w was 0.88 (with glycerol as the humectant). When growing cells in broth controlled at pH 6.3 were used, a reduction in the a w from 0.99 to 0.96 resulted in a decrease in the level of lactose carbon recovered as l -(+)-lactate from 100 to 71%. Low levels of l -(+)-lactate carbon recovery (w values of 0.99 and 0.87 and in young cheese curds. The missing lactose carbon could not be accounted for by acetate, ethanol, formate, acetaldehyde, or pyruvate. Attempts were made to determine where the missing lactose carbon was diverted to under the stress conditions used. Some of the missing lactose carbon was recovered as galactose (0.1 to 2.5 mM) in culture supernatants. Decreasing either the a w or the pH resulted in increased galactose accumulation by nongrowing cells; adjusting both environmental factors together potentiated the effect. The sensitivities of the two lactococcal strains tested were different; strain 2272 was more prone to accumulate galactose under stress conditions. A methyl pentose(s) and additional galactose were found in acid-hydrolyzed supernatants from cultures containing both growing and nongrowing cells, indicating that a saccharide(s) rich in these components was formed by lactococci under low-a w and low-pH stress conditions.

This article begins with an introduction to malolactic fermentation in wine, followed by a review of the occurrence of arginine degradation in wine lactic acid bacteria and the pathway of arginine catabolism, the distribution of enzymes responsible, and the formation of products. The bioenergetics of wine lactic acid bacteria and arginine degradation are then reviewed. This is followed by a review of the possible mechanisms of arginine transport, and regulation of arginine metabolism and synthesis of the enzymes for arginine catabolism. Finally, the practical significance of arginine metabolism in wine lactic acid bacteria is reviewed with respect to taxonomic utility, biological significance and oenological implications.

We report a simple enzymatic approach to synthesise phenethyl esters as natural flavouring materials. Chemical and lipase-catalysed esterification reactions between fatty acids of C4–C18 and 2-phenethyl alcohol were studied. Both methods were compared qualitatively and quantitatively by GC–MS/FID. The acid and thermal stabilities of 2-phenethyl esters were excellent and can meet the requirements of food matrices under most processing conditions. Sensory evaluation showed that each 2-phenethyl ester with a different carbon-chain-length fatty acid had unique sensory notes. Moreover, through Lipozyme TL IM-mediated transesterification, valuable 2-phenethyl alcohol-derived esters were synthesised from butter oil and 2-phenethyl alcohol. The influence of several physicochemical parameters (temperature, substrate molar ratio, enzyme loading, shaking speed and time) on the transesterification reaction was investigated to give optimal reaction conditions, leading to a high yield of 80.0%.

Summary This study determined the influence of pulp maceration and β-glucosidase on mango wine physico-chemical properties and volatile profiles. The distinction in pH, sugars and organic acids among different treatments was not statistically significant. All wine samples reached around 8% (v/v) ethanol from about 16% (w/v) of sugars. The wine with pulp contact contained about ten times higher α-terpinolene and up to three times higher acetate esters than the wine without pulp contact, but mitigated the production of medium-chain fatty acids and relative ethyl esters by up to six times. β-glucosidase enhanced terpenols by up to ten times and acetate esters by up to three times. Furthermore, enzyme treatment mitigated, by up to five times, the formation of medium-chain fatty acids and ethyl esters to moderate levels. Sensory evaluation showed pulp contact, and β-glucosidase not only improved the intensity and complexity of wine aroma but balanced odour attributes.

Methionol-derived fatty acid esters were synthesised by both chemical and lipase catalysed esterification between fatty acids and methionol. Beneficial effects of both methods were compared qualitatively and quantitatively by GC-MS/GC-FID results. And the high acid and heat stability of our designed methionyl esters meet the requirement of the food industry. Most importantly, the sensory test showed that fatty acid carbon-chain length had an important effect on the flavour attributes of methionyl esters. Moreover, through Lipozyme TL IM-mediated transesterification, valuable methionol-derived esters were synthesised from the readily available natural material butter oil as the fatty acid source. The conversion of methionol and yield of each methionyl ester were also elucidated by GC-MS-FID.

In this work, we reported a novel application of pressurised liquid extraction (PLE) on coffee bean. The condition of PLE was carefully optimised with the aid of response surface methodology (RSM) including adjustment of experimental parameters (solvent type and sample to hydromatrix ratio) and other operating parameters (i.e. temperature (50–100 °C), pressure (1000–2000 psi) and static extraction time (5–15 min)). The coffee extracts obtained under three different extraction conditions were evaluated through descriptive sensory analysis. Then, the results showed that those targeted compounds obtained from PLE were nearly three times higher (1473 ppm) than conventional solvent extraction (571 ppm). Thus, PLE demonstrated the feasibility of producing a series of coffee extracts under controllable extraction conditions in correlation with desirable sensory attributes. This approach has not previously reported to characterise the aroma of coffee bean.

The impact of the addition of fusel oil or amino acids on the volatile compounds in papaya wine fermented with a mixed culture of Saccharomyces cerevisiae var. bayanus R2 and Williopsis saturnus var. mrakii NCYC 2251 at a ratio of 1:1000 was studied. Fusel oil addition increased the fraction of alcohols and promoted the production of isoamyl octanoate, isoamyl decanoate and isobutyl decanoate, while decreased the fraction of ethyl acetate and 2-phenylethyl acetate. The addition of amino acids enhanced the formation of total volatile fatty acids, 2-phenylethanol and some ethyl esters. The papaya wine with added amino acids possessed more acidic and buttery notes than the control, while that with added fusel oil had an overall aroma profile comparable to that of the control. This study suggests that papaya juice fermentation with mixed yeasts in conjunction with the added fusel oil or selected amino acids may be another method of modulating the flavour of papaya wine., U ovom je radu ispitan utjecaj dodatka patočnog ulja ili aminokiselina na hlapljive sastojke vina dobivenog fermentacijom soka papaje pomoću mješovite kulture kvasaca Saccharomyces cerevisiae var. bayanus R2 i Williopsis saturnus var. mrakii NCYC 2251 u omjeru 1:1000. Dodatkom patočnog ulja povećan je udjel alkohola, te je poboljšana proizvodnja izoamilnog oktanoata, izoamilnog dekanoata i izobutilnog dekanoata, dok je smanjen udjel etilnog acetata i 2-feniletilnog acetata. Nastanak hlapljivih masnih kiselina, 2-feniletanola i nekih etilnih estera pospješen je dodatkom aminokiselina. U usporedbi s kontrolnim uzorkom, vino proizvedeno fermentacijom papaje uz dodatak aminokiselina imalo je izraženiju kiselu i putrastu notu, dok se aroma vina kojem je dodano patočno ulje nije razlikovala. Ovo je istraživanje pokazalo da se fermentacijom soka papaje pomoću mješovite kulture kvasaca uz dodatak patočnog ulja ili odabranih aminokiselina može promijeniti aroma vina.