Spontaneous Malolactic Fermentation Research Articles

HPLC/HILIC determination of biogenic amines in wines produced by different winemaking technologies

AbstractThe present study evaluated the effect of winemaking technologies on the concentration of different biogenic amines in Chardonnay wines. Wines produced from sedimented, inoculated must with active dry yeast without malolactic fermentation were compared with wine produced from nonsedimented must spontaneously fermented with malolactic fermentation. Histamine and putrescine concentrations were not significantly different in either variant. The highest concentration of histamine was 0.055 mg L−1, and the highest concentration of putrescine was 1.6 mg L−1 in both variants. Statistically significantly higher values of cadaverine (from 0.06 to 0.07 mg L−1), spermidine (from 0.8 to 1.4 mg L−1), spermine (from 0.15 to 0.25 mg L−1), and isoamylamine (from 0.40 to 0.46 mg L−1) were found in the variant made from nonsedimented must, in which spontaneous malolactic fermentation was performed. The higher concentration of biogenic amines in this variant may be due to the different composition of lactic bacteria during the spontaneous malolactic fermentation. A simplified, unpublished HILIC method of chromatographic separation of biogenic amines without prior deprivation with MS-MS detection was used to determine individual biogenic amines.

Malolactic fermentation of lactic acid bacteria isolated from southern Brazilian red wine.

The objective was to isolate lactic acid bacteria (LAB) from southern Brazil's wines and investigate their potential as starter cultures for malolactic fermentation (MLF) in Merlot (ME) and Cabernet Sauvignon (CS) wines through the fermentative capacity. The LAB were isolated from CS, ME, and Pinot Noir (PN) wines in the 2016 and 2017 harvests and evaluated for morphological (color and shape of the colonies), genetic, fermentative (increase in pH, acidity reduction, preservation of anthocyanins, decarboxylation of L-malic acid, yield of L-lactic acid, and content of reduced sugars), and sensory characteristics. Four strains were identified as Oenococcus oeni [CS(16)3B1, ME(16)1A1, ME(17)26, and PN(17)65], one as Lactiplantibacillus plantarum [PN(17)75], and one as Paucilactobacillus suebicus [CS(17)5]. Isolates were evaluated in the MLF and compared to a commercial strain (O. oeni), as well as a control (without inoculation and spontaneous MLF), and standard (without MLF). CS(16)3B1 and ME(17)26 isolates finished the MLF for CS and ME wines, respectively, after 35 days, similar to the commercial strain, and CS(17)5 and ME(16)1A1 isolates ended the MLF in 45 days. In the sensory analysis, ME wines with isolated strains received better scores for flavor and overall quality than the control. Compared to the commercial strain, CS(16)3B1 isolate obtained the highest scores for buttery flavor and taste persistence. CS(17)5 isolate received the higher scores for a fruity flavor and overall quality and the lowest for a buttery flavor. The native LAB displayed MLF potential, regardless of the year and grape species from which they were isolated.

Acetaldehyde accumulation during wine micro oxygenation: The influence of microbial metabolism

Acetaldehyde is a key oxidation product during red wine micro oxygenation (Mox) and it appears to accumulate by the metabolism of residual yeast Saccharomyces cerevisiae (S. cerevisiae), which may be difficult to detect by plating due to low numbers. The aim of this study was to investigate the roles and interaction of S. cerevisiae and lactic acid bacterial (LAB) with regard to acetaldehyde accumulation during red wine Mox treatment. Immediately after alcohol fermentation using three different yeast S. cerevisiae strains, some wine was sterile filtered, and of that, some was inoculated with the same three yeasts. All were treated with Mox; acetaldehyde and malolactic fermentation (MLF) were monitored. Besides, the capacity of LAB to metabolize acetaldehyde was also evaluated in model wine. The results showed that Mox appeared to maintain S. cerevisiae populations, and the survival of S. cerevisiae postponed spontaneous MLF. S. cerevisiae did produce acetaldehyde during Mox, while levels varied by strains, production rates declined during treatment, and levels dropped after treatment ended. The appearance of LAB coincided with an increase in production, but levels later dropped more quickly when LAB were present. Residual sugar and amino acid levels did not change significantly during microbial metabolism of Mox wines. The acetaldehyde generated from chemical oxidation continued to accumulate during Mox. In the experiment of model wine, LAB was shown to produce acetaldehyde in the inception phase and a certain amount of acetaldehyde did not influence the start of MLF. The results confirmed the variation character of acetaldehyde with microbial action and can serve as a theoretical underpinning for better control of Mox applications in enology.

Preliminary study of novel autochthonous starter culture for red wine production with reduced biogenic amine content

Most Chilean red wine is produced by spontaneous malolactic fermentation (MLF), which generates prolonged fermentations resulting in uncertain results, increase production cost, and lead to biogenic amine (BA) formation. The objective of this work was to carry out a preliminary study of an autochthonous MLF starter culture to perform shorter and controlled MLF to elaborated red wines with low BA content. The autochthonous Oenococcus oeni strain (OoB6) isolated from spontaneous MLF was assayed in bioreactors using filtered and unfiltered wines. OoB6 strain efficiently carried out MLF samples showing a similar performance to commercial starter culture. Both accomplished malic acid concentrations (<0.12 g/L) significantly lower (p < .05) than the initial content (1.21 ± 0.11 g/L). OoB6 strain produced a 50% reduction in MLF time and showed very low BA formation (ca. 5 mg/L) without detecting the presence toxicological important BA histamine and tyramine. Novelty impact statement To the best of our knowledge, this work report for the first time a preliminary study of potential autochthonous MLF starter culture to elaborate Chilean red wines. Autochthonous MLF starter culture produced a 50% reduction in MLF time compared with traditional spontaneous MLF. Autochthonous MLF starter culture showed very low BA formation (ca. 5 mg/L) without detecting the presence toxicological important BA histamine and tyramine.

Phage-host interactions as a driver of population dynamics during wine fermentation: Betting on underdogs

Winemaking is a complex process in which numerous microorganisms, mainly yeasts and lactic acid bacteria (LAB), play important roles. After alcoholic fermentation (AF), most wines undergo malolactic fermentation (MLF) to improve their organoleptic properties and microbiological stability. Oenococcus oeni is mainly responsible for this crucial process where L-malic acid (MA) in wine converts to softer L-lactic acid. The bacterium is better adapted to the limiting conditions imposed by the wine matrix and performs MLF under regular winemaking conditions, especially in wines with a pH below 3.5. Traditionally, this process has been conducted by the natural microbiota present within the winery. However, the start, duration and qualitative impact of spontaneous MLF are unpredictable, which prompts winemakers to use pure starter cultures of selected bacteria to promote a more reliable, simple, fast and efficient fermentation. Yet, their use does not always ensure a problem-free fermentation. Spontaneous initiation of the process may prove very difficult or does not occur at all. Such difficulties arise from a combination of factors found in some wines upon the completion of AF (high ethanol concentration, low temperature and pH, low nutrient concentrations, presence of free and bound SO2). Alongside these well documented facts, research has also provided evidence that negative interactions between O. oeni and other biological entities such as yeasts may also impact MLF. Another insufficiently described, but highly significant factor inhibiting bacterial growth is connected to the presence of bacteriophages of O. oeni which are frequently associated to musts and wines. The purpose of this review is to summarize the current knowledge about the phage life cycles and possible impacts on the trajectory of the microbiota during winemaking.

Protein content of the Oenococcus oeni extracellular vesicles-enriched fraction

Malolactic fermentation is essential for the quality of red wines and some other wine styles. Spontaneous malolactic fermentation is often driven by Oenococcus oeni, and commercial starters for this purpose are also often of this species. The increasing number of microbial species and inoculation strategies in winemaking has prompted a growing interest in microbial interactions during wine fermentation. Among other interaction mechanisms, extracellular vesicles have been hypothesized to play a role in this context. Extracellular vesicles have already been described and analysed for several wine yeast species. In this work, the production of extracellular vesicles by O. oeni is reported for the first time. The protein content of these extracellular vesicles is also characterised. It shows differences and similarities with the recently described protein content of Lactiplantibacillus plantarum, a bacterial species also capable of performing malolactic fermentation of wine (and used sometimes as an alternative starter). This work further contributes to the development of the field of extracellular vesicles in food biotechnology.

New Malolactic Bacteria Strains Isolated from Wine Microbiota: Characterization and Technological Properties

Malolactic fermentation (MLF) or biological decrease of wine acidity is defined as the enzymatic bioconversion of malic acid in lactic acid, a process performed by lactic acid bacteria (LAB). The procedures for the isolation of new indigenous LAB strains from the red wines produced in Copou Iasi wine center (NE of Romania) undergoing spontaneous malolactic fermentation, resulted in the obtaining of 67 catalase-negative and Gram-positive LAB strains. After testing in the malolactic fermentative process, application of specific screening procedures and identification (API 50 CH), two bacterial strains belonging to the species Oenococcus oeni (strain 13-7) and Lactobacillus plantarum (strain R1-1) with high yield of malolactic bioconversion, non-producing biogenic amines, and with active extracellular enzymes related to wine aroma, were retained and characterized. Tested in synthetic medium (MRS-TJ) for 10 days, the new isolated LAB strains metabolized over 98% of the malic acid at ethanol concentrations between 10 and 14 % (v/v), low pH (&gt;3.0), total SO2 doses up to 70 mg/L and temperatures between 15 and 35 °C, showing high potential for future use in the winemaking process as bacterial starter cultures, in order to obtain high quality wines with increased typicity.

Assessment of the lysogenic status in the lactic acid bacterium O. oeni during the spontaneous malolactic fermentation of red wines

After alcoholic fermentation, most wines undergo malolactic fermentation (MLF), driven by the lactic acid bacterium Oenococcus oeni, which improves their organoleptic properties and microbiological stability. Prophages were recently shown to be notably diverse and widely disseminated in O. oeni genomes. Such in silico predictions confirmed previous cultivation-based approaches which showed frequent lysis of strains upon treatment with the inducing agent mitomycin C. Both strategies used to assess lysogeny in the species were so far applied to a number of strains collected from distinct countries, wineries, cepages and fermentation processes. Results may not therefore be representative of the lysogenic population in natural communities driving the MLF during winemaking. Here we report the prevalence of lysogeny during winemaking in three wineries in the Bordeaux area. The dominant LAB population was collected in 11 red wines upon completion of MLF. Using VNTR and prophage typing analyses, our data confirm the presence of lysogens in the population driving the spontaneous MLF in all tested wines, although lysogeny rates varied across wineries. Higher prevalence of lysogeny was associated to a reduced diversity in VNTR profiles, the dominance of a few prophage-types and presence of some bacterial genetic backgrounds that were particularly prone to lysogenization.

The Impact of the Fermentation Strategy on the Flavour Formation of Ilzer Rose (Malus domestica Borkh.) Apple Wine.

The flavour and the volatilome of apple wines made from the Austrian heritage variety Ilzer Rose was in the scope of this study. The apple wines were produced by adopting oenological practises that are not commonly used in fruit wine production. Different fermentation strategies including the addition of enzymes with β-glucosidase activity, addition of a fining agent, maceration of the mash along with mash fermentation were applied. The volatile compounds of the juices as intermediates and the resulting apple wines were analysed using headspace-SPME GC-MS. CATA technique with a well-trained panel was applied for sensory evaluation. The results show that the flavour of single-variety apple wine can be significantly altered by taking oenological measures. High correlations were found between the results of the analytical investigation and the sensory evaluation. Maceration of the mash leads to an increase in the fruity character of the products, also reflected by significantly higher fruit ester quantities in the wine. During mash fermentation, spontaneous malolactic fermentation was induced leading to a product with new, but thoroughly interesting sensory properties of the apple wine. The results of this study demonstrate that the integration of oenological measures may open a wide field to the development of a high diversity in apple wine flavour.

The content of biogenic amines in Rondo and Zweigelt wines and correlations between selected wine parameters

The purpose of this study was to evaluate the content of biogenic amines (BAs) in wines using dispersive liquid–liquid microextraction–gas chromatography–mass spectrometry (DLLME-GC–MS). An additional objective was to assess the correlations between selected parameters characterizing the samples such as the content of BAs, sugars, and organic acids, pH, and total acidity. Wines produced from the same grape variety in which alcoholic fermentation (AF) was carried out by different yeast strains and in which malolactic fermentation (MLF) was spontaneous, differed in the content of biogenic amines. The concentrations of putrescine, cadaverine and tryptamine were higher in the Rondo wines (237–405, 34.04–61.11, <LOD-12.456 µg/L, respectively) and Zweigelt wines (416–489, 72.67–88.43, <LOD-13.083 µg/L, respectively) subjected to spontaneous MLF than in the wines subjected to induced MLF. Chemometric analysis allowed us to determine correlations between selected wine parameters. The wine samples are well separated into two patterns depending on the grape variety. Despite the fact that information on BAs is not included in databases of wine composition, information on their concentration as well as knowledge of existing correlations between BAs and other wine parameters is crucial and may be useful for the food industry, health professionals and consumers.

Utjecaj različitih načina provođenja jabučno-mliječne fermentacije na promjene kemijskih svojstava i hlapivih komponenti crnog vina Teran (Vitis vinifera L.)

Malolactic fermentation (MLF) is a complex biochemical process playing an important role in the production of red wines. The main reasons for its implementation are the reduction of titratable acidity and the consequent increase of pH, microbiological stability of wine, and changes in aromatic and sensory properties of wine. The aim of this study was to determine the compatibility of yeast and bacteria used by different malolactic fermentation techniques and their influence on the fermentation duration, concentration of individual organic acids, aroma compounds, and on chemical and sensory properties of Teran wines. The experiment included control treatment (without MLF), spontaneous MLF, induced MLF at the beginning of alcoholic fermentation with simultaneous inoculation of yeast and bacteria (co-inoculation) and induced MLF after alcoholic fermentation (sequential MLF). In the co-inoculation treatment MLF had no negative effect on the alcoholic fermentation kinetic. Alcoholic fermentation was complete in all treatments. Co-inoculation resulted in a significantly shorter duration of the MLF process. In all MLF treatments, significant reduction of titratable acidity and the increase of pH values was noted. Wines of the spontaneous MLF treatment obtained the highest concentration of volatile acidity and ethyl acetate. In all MLF treatments a complete consumption of malic acid and a decrease in concentration of citric acid, total amount of higher alcohols, and acetaldehyde were observed. Furthermore, significantly higher concentrations of ethyl esters, diacetyl, acetoin, and 2,3-butanediol were present in wines from all MLF treatments.

Genome Sequence of Oenococcus oeni OE37, an Autochthonous Strain Isolated from an Italian White Wine.

Oenococcus oeni OE37 is an autochthonous strain that was isolated from a Chardonnay wine from Piedmont (Italy) during spontaneous malolactic fermentation. Here, the OE37 genome sequence is presented, and a brief description of the main genes is reported.

Effect of mixed fermentation by Torulaspora delbrueckii, Saccharomyces cerevisiae, and Lactobacillus plantarum on the sensory quality of black raspberry wines

Till now, very few studies are focused on the interactive effect of wine-associated microbes including non-Saccharomyces, Saccharomyces, and lactic acid bacteria on modulating the organoleptic quality of black raspberry wines. In our study, Torulaspora delbrueckii, a widely studied non-Saccharomyces yeast, and Lactobacillus plantarum, a promising malolactic fermentation starter, were used in combination with S. cerevisiae for the production of black raspberry wine by three inoculation modes, i.e., sequential inoculation of three species (SE), co-culture of S. cerevisiae and L. plantarum (CO) and T. delbrueckii/S. cerevisiae sequential inoculation plus a spontaneous malolactic fermentation (SP), on the quality of black raspberry wine. A total of 34 aromatic compounds were identified by HS–GC–IMS technology. CO was found to boost the production of many fruity ester compounds, and scored the highest for ‘fruity’, ‘sweet’, and ‘total aroma’ attributes during sensory evaluation. SP was characterized by the lowest production of most volatile components and obtained the lowest rating. Principal component analysis showed that three samples were quite different from each other. Overall, CO had greater potential to be used in the production of black raspberry wine.

Effect of Saccharomyces cerevisiae, Torulaspora delbrueckii and malolactic fermentation on fermentation kinetics and sensory property of black raspberry wines

Alcoholic fermentation (AF) and malolactic fermentation (MLF) both have significant influence on the production of black raspberry wine. In this study, three microbes associated with AF and MLF including S. cerevisiae, T. delbrueckii and O. oeni were used to investigate their combined effect on basic compositional, volatile and sensory property of black raspberry wine, and four fermentation trials including single S. cerevisiae inoculation plus spontaneous MLF (BSU) and controlled MLF with O. oeni (BSO), sequential culture of T. delbrueckii and S. cerevisiae plus spontaneous MLF (BTSU) and controlled MLF (BTSO) were tested and compared. Fermentation results showed MLF in BSU, BSO and BTSO were successful, with respective period of 40, 25 and 23 days, whereas a stuck MLF occurred in BTSU. Volatile compounds were determined by HS-GC-IMS method, with a total of 45 aromas identified. BTSO was distinguished by a significant higher signal intensity of many fruity esters and a lower production of several alcohols and terpenes, which was in agreement with its perception result of strong ‘fruity’ and slight note of ‘solvent’ and ‘herbaceous’ during quantitative descriptive analysis. On the contrary, BSU was found to reinforce the synthesis of most detected volatiles, resulting in the enhancement of both beneficial and off-flavour compounds, therefore scoring lower in the ‘global aroma’ descriptor. Principal component analysis showed BSU and BSO were similar in the volatile composition, whereas BTSO was quite different. Overall, BTSO had greater potential to be used in the production of black raspberry wine.

Lactobacillus plantarum, a New Biological Tool to Control Malolactic Fermentation: A Review and an Outlook

Malolactic fermentation (MLF) in wine is an important step in the vinification of most red and some white wines, as stands for the biological conversion of l-malic acid into l-lactic acid and carbon dioxide, resulting in a decrease in wine acidity. MLF not only results in a biological deacidification, it can exert a significant impact on the organoleptic qualities of wine. This paper reviews the biodiversity of lactic acid bacteria (LAB) in wine, their origin, and the limiting conditions encountered in wine, which allow only the most adapted species and strains to survive and induce malolactic fermentation. Of all the species of wine LAB, Oenococcus oeni is probably the best adapted to overcome the harsh environmental wine conditions and therefore represents the majority of commercial MLF starter cultures. Wine pH is most challenging, but, as a result of global warming, Lactobacillus sp. is more often reported to predominate and be responsible for spontaneous malolactic fermentation. Some Lactobacillus plantarum strains can tolerate the high alcohol and SO2 levels normally encountered in wine. This paper shows the potential within this species for the application as a starter culture for induction of MLF in juice or wine. Due to its complex metabolism, a range of compositional changes can be induced, which may positively affect the quality of the final product. An example of a recent isolate has shown most interesting results, not only for its capacity to induce MLF after direct inoculation, but also for its positive contribution to the wine quality. Degrading hexose sugars by the homo-fermentative pathway, which poses no risk of acetic acid production from the sugars, is an interesting alternative to control MLF in high pH wines. Within this species, we can expect more strains with interesting enological properties.

Co-Existence of Inoculated Yeast and Lactic Acid Bacteria and Their Impact on the Aroma Profile and Sensory Traits of Tempranillo Red Wine

This study investigates the effects of simultaneous inoculation of a selected Saccharomyces cerevisiae yeast strain with two different commercial strains of wine bacteria Oenococcus oeni at the beginning of the alcoholic fermentation on the kinetics of malolactic fermentation (MLF), wine chemical composition, and organoleptic characteristics in comparison with spontaneous MLF in Tempranillo grape must from Castilla-La Mancha (Spain). Evolution of MLF was assessed by the periodic analysis of L-malic acid through the enzymatic method, and most common physiochemical parameters and sensory traits were evaluated using a standardized sensory analysis. The samples were analyzed by GC/MS in SCAN mode using a Trace GC gas chromatograph and a DSQII quadrupole mass analyzer. Co-inoculation reduced the overall fermentation time by up to 2 weeks leading to a lower increase in volatile acidity. The fermentation-derived wine volatiles profile was distinct between the co-inoculated wines and spontaneous MLF and was influenced by the selected wine bacteria used in co-inoculation. Co-inoculation allows MLF to develop under reductive conditions and results in wines with very few lactic and buttery flavors, which is related to the impact of specific compounds like 2,3-butanedione. This compound has been also confirmed as being dependent on the wine bacteria used.

A Metagenomic-Based Approach for the Characterization of Bacterial Diversity Associated with Spontaneous Malolactic Fermentations in Wine.

This study reports the first application of a next generation sequencing (NGS) analysis. The analysis was designed to monitor the effect of the management of microbial resources associated with alcoholic fermentation on spontaneous malolactic consortium. Together with the analysis of 16S rRNA genes from the metagenome, we monitored the principal parameters linked to MLF (e.g., malic and lactic acid concentration, pH). We encompass seven dissimilar concrete practices to manage microorganisms associated with alcoholic fermentation: Un-inoculated must (UM), pied-de-cuve (PdC), Saccharomyces cerevisiae (SC), S. cerevisiae and Torulaspora delbrueckii co-inoculated and sequentially inoculated, as well as S. cerevisiae and Metschnikowia pulcherrima co-inoculated and sequentially inoculated. Surprisingly, each experimental modes led to different taxonomic composition of the bacterial communities of the malolactic consortia, in terms of prokaryotic phyla and genera. Our findings indicated that, uncontrolled AF (UM, PdC) led to heterogeneous consortia associated with MLF (with a relevant presence of the genera Acetobacter and Gluconobacter), when compared with controlled AF (SC) (showing a clear dominance of the genus Oenococcus). Effectively, the SC trial malic acid was completely degraded in about two weeks after the end of AF, while, on the contrary, malic acid decarboxylation remained uncomplete after 7 weeks in the case of UM and PdC. In addition, for the first time, we demonstrated that both (i) the inoculation of different non-Saccharomyces (T. delbrueckii and M. pulcherrima) and, (ii) the inoculation time of the non-Saccharomyces with respect to S. cerevisiae resources (co-inoculated and sequentially inoculated) influence the composition of the connected MLF consortia, modulating MLF performance. Finally, we demonstrated the first findings of delayed and inhibited MLF when M. pulcherrima, and T. delbrueckii were inoculated, respectively. In addition, as a further control test, we also assessed the effect of the inoculation with Oenococcus oeni and Lactobacillus plantarum at the end of alcoholic fermentation, as MLF starter cultures. Our study suggests the potential interest in the application of NGS analysis, to monitor the effect of alcoholic fermentation on the spontaneous malolactic consortium, in relation to wine.

Survival and implantation of indigenous psychrotrophic Oenococcus oeni strains during malolactic fermentation in a Patagonian Pinot noir wine

Spontaneous malolactic fermentation (MLF) in the North Patagonian region of Argentina may have to bear low environmental temperatures, an important stress factor for the lactic acid bacteria (LAB) that conduct this process. Thus, inducing MLF by inoculation of a native starter culture able to lead this biological process at low temperatures is encouraged because it would allow preserving the regional character of the wine. The goal of this work was to obtain autochthonous Oenococcus oeni strains able to consume l-malic acid when MLF takes place at temperatures lower than 15 °C. Among the psychrotrophic isolates obtained from a Pinot noir wine, sixteen O. oeni strains were identified by sequencing the 16S rRNA gene and typing by RAPD PCR, allowing the differentiation of five genotypic groups. After the evaluation of the viability and metabolic performance of the strains in sterile wine at low temperatures, strains UNQOe4 and UNQOe19 were selected to evaluate their behavior in a non-sterile wine at 4 and 10 °C. The strains selected showed good implantation capacity and were able to successfully conduct MLF, suggesting that their use as starter cultures is suitable to improve this biological process in a wine region where environmental temperatures can be low.

Identification of the potential inhibitors of malolactic fermentation in wines

Abstract This exploratory work aims to identify the potential inhibitors of lactic bacterial growth and to propose enological practices to guarantee the occurrence of spontaneous malolactic fermentation (MLF) in wines from traditional and double-pruning management harvests in southeast Brazil. One white wine from a summer harvest and one red wine from a winter harvest that failed to complete MLF were utilized as comparative models to identify inhibitor compounds to lactic bacteria. Wine composition, alcoholic-fermentation temperature and bacterial strain contribute to the success or failure of MLF. Temperatures below 12 °C during alcoholic fermentation decrease lactic bacterial metabolism and may impair the bacteria’s growth after yeast cells lysis. A must pH below 3.2 in a summer harvest impairs bacterial growth, and the association of low pH with a free-SO2 concentration above 10 mg L-1 may inhibit MLF. For grapes with a high sugar content, harvested in the winter cycle, enologists should keep the alcohol content below 15% and control the alcoholic-fermentation temperature.

Impact of malolactic fermentation on the volatile composition of Turkish Kalecik karası red wines

The effects of spontaneous malolactic fermentation (MLF) and a malolactic starter culture (Oenococcus oeni) on the volatile compounds of Kalecik karası red wines were investigated. The volatile compounds were extracted using liquid–liquid extraction and quantified by GC–MS–flame ionisation detection. Starter cultures of O. oeni had a positive effect in shortening MLF and producing lower volatile acidity. After MLF, the total volatile compounds increased in both the inoculated wines and the spontaneously fermented wine. Ethyl lactate, diethyl succinate and γ-butyrolactone increased upon inoculation with cultures of O. oeni strains PN4 and VP41. MLF increased the diacetyl concentration in all Kalecik karası wines. Strain dependent differences were found in the level of diacetyl, with O. oeni PN4 producing more diacetyl than O. oeni VP41. After MLF, the concentrations of acetoin, 4-ethylguaicol and 4-ethylphenol increased in spontaneously fermented wine compared with wines inoculated with O. oeni. Accordingly, the use O. oeni PN4 is recommended for a controlled MLF to produce better quality Kalecik karası wines. © 2018 The Institute of Brewing & Distilling