Prevalence and comparison of Streptococcus infantarius subsp. infantarius and Streptococcus gallolyticus subsp. macedonicus in raw and fermented dairy products from East and West Africa

Streptococcus infantarius subsp. infantarius (Sii) and Streptococcus gallolyticus subsp. macedonicus are members of the Streptococcus bovis/Streptococcus equinus complex (SBSEC) associated with human infections. SBSEC-related endocarditis was furthermore associated with rural residency in Southern Europe. SBSEC members are increasingly isolated as predominant species from fermented dairy products in Europe, Asia and Africa. African variants of Sii displayed dairy adaptations to lactose metabolism paralleling those of Streptococcus thermophilus including genome decay. In this study, the aim was to assess the prevalence of Sii and possibly other SBSEC members in dairy products of East and West Africa in order to identify their habitat, estimate their importance in dairy fermentation processes and determine geographic areas affected by this potential health risk. Presumptive SBSEC members were isolated on semi-selective M17 and SM agar media. Subsequent genotypic identification of isolates was based on rep-PCR fingerprinting and SBSEC-specific16S rRNA gene PCR assay. Detailed identification was achieved through application of novel primers enhancing the binding stringency in partial groES/groEL gene amplification and subsequent DNA sequencing. The presence of S. thermophilus-like lacS and lacZ genes in the SBSEC isolates was determined to elucidate the prevalence of this dairy adaptation. Isolates (n = 754) were obtained from 72 raw and 95 fermented milk samples from Côte d'Ivoire and Kenya on semi-selective agar media. Colonies of Sii were not detected from raw milk despite high microbial titers of approximately 10⁶ CFU/mL on M17 agar medium. However, after spontaneous milk fermentation Sii was genotypically identified in 94.1% of Kenyan samples and 60.8% of Kenyan isolates. Sii prevalence in Côte d'Ivoire displayed seasonal variations in samples from 32.3% (June) to 40.0% (Dec/Jan) and isolates from 20.5% (June) to 27.7% (Dec/Jan) present at titers of 10⁶–10⁸ CFU/mL. lacS and lacZ genes were detected in all Kenyan and 25.8% (June) to 65.4% (Dec/Jan) of Ivorian Sii isolates. Regional differences in prevalence of Sii and dairy adaptations were observed, but no clear effect of dairy animal, fermentation procedure and climate was revealed. Conclusively, the high prevalence of Sii in Kenya, Côte d'Ivoire in addition to Somalia, Sudan and Mali strongly indicates a pivotal role of Sii in traditional African dairy fermentations potentially paralleling that of typical western dairy species S. thermophilus. Putative health risks associated with the consumption of high amounts of live Sii and potential different degrees of evolutionary adaptation or ecological colonization require further epidemiologic and genomic investigations, particularly in Africa.     

Application of Kluyveromyces marxianus, Lactobacillus delbrueckii ssp. bulgaricus and L. helveticus for sourdough bread making

The application of Kluyveromyces marxianus (IFO 288), Lactobacillus delbrueckii ssp. bulgaricus (ATCC 11842) and Lactobacillus helveticus (ATCC 15009) as starter cultures for sourdough bread making was examined. Production of lactic and acetic acids, bread rising, volatile composition, shelf-life and organoleptic quality of the sourdough breads were evaluated. The amount of starter culture added to the flour, the dough fermentation temperature and the amount of sourdough used were examined in order to optimise the bread making process. The use of mixed cultures led to higher total titratable acidities and lactic acid concentrations compared to traditionally made breads. Highest acidity (3.41 g lactic acid/kg of bread) and highest resistance to mould spoilage were observed when bread was made using 50% sourdough containing 1% K. marxianus and 4% L. delbrueckii ssp. bulgaricus. The use of these cultures also improved the aroma of sourdough breads, as shown by sensory evaluations and as revealed by GC–MS analysis.     

Essential oils of the aerial parts of three Salvia species from Jordan: Salvia lanigera, S. spinosa and S. syriaca

The compositions of the essential oils of three Jordanian Salvia species, S. lanigera and S. spinosa from a desert climate, and S. syriaca from a Mediterranean habitat, were studied. All three oils are rich in monoterpene derivatives (68–73%).     

Essential oils of Zingiber officinale var. rubrum Theilade and their antibacterial activities

The essential oils obtained by hydrodistilation of the leaves and rhizomes of Zingiber officinale var. rubrum Theilade were analysed by capillary GC and GC–MS. Forty-six constituents were identified in the leaf oil, while 54 were identified in the oil from the rhizomes. The leaf oil was clearly dominated by β-caryophyllene (31.7%), while the oil from the rhizomes was predominantly monoterpenoid, with camphene (14.5%), geranial (14.3%), and geranyl acetate (13.7%) the three most abundant constituents. The evaluation of antibacterial activities using the micro-dilution technique revealed that both the leaf and rhizome oils were moderately active against the Gram-positive bacteria Bacilluslicheniformis, Bacillus spizizenii and Staphylococcus aureus, and the Gram-negative bacteria Escherichia coli, Klebsiella pneumoniae and Pseudomonas stutzeri.     

Determination of antioxidant activities of various extracts and essential oil compositions of Thymus praecox subsp. skorpilii var. skorpilii

The aim of this work is to examine the chemical composition and antioxidant activity of separated essential oils and different solvent extracts of Thymus praecox subsp. skorpilii var. skorpilii (TPS). The ethanol, acetone, methanol, hexane, aqueous extracts and separated essential oils of TPS were assessed for their antioxidant activities. Antioxidant activities were evaluated by reduction of Mo(VI) to Mo(V), reducing power, superoxide scavenging activity, free radical-scavenging activity, metal chelating activity, linoleic acid peroxidation, hydrogen peroxide scavenging activity and peroxide scavenging activity. Essential oils were characterized in total to be 41 components, whereas 9 components were isolated by column chromatography for antioxidant activity. TPS essential oil was found to contain thymol (40.31%) and o-cymene (13.66%) as the major components. The ethanol, methanol and water extracts exerted significant free radical-scavenging activity. The methanol and water extracts displayed highest superoxide scavenging activity. The water extract has the highest total phenolics (6.211 mg gallic acid (GAE)/g DW) and flavonoids (0.809 mg quercetin/g DW).     

Composition of the essential oil of Daucus gingidium L. ssp. gingidium

The essential oils obtained from leaves and fruits of Daucus gingidium ssp. gingidium have been studied. The main constituents of the essential oil from the leaves were sabinene (26.8%), α-pinene (10.8%), germacrene D (6.9%) and limonene (5.7%). Sabinene (60.6%) was the main compound identified in the essential oil of the fruits, followed by α-pinene (12.2%) and 4-terpineol (5.4%). Furthermore, qualitative and quantitative considerations about differences with literature data on Daucus carota have been made in order to confirm the species status of D. gingidium.     

Simultaneous analysis of purine alkaloids and catechins in Camellia sinensis, Camellia ptilophylla and Camellia assamica var. kucha by HPLC

Simultaneous analysis of purine alkaloids and catechins in tea from dry leaves of Camellia sinensis, Camellia ptilophylla and Camellia assamica var. kucha by a reversed-phase high-performance liquid chromatography (RP-HPLC) method. This HPLC method had been proved to be appropriate for the identification and quantification of purine alkaloids and catechins, and exhibited good correlation coefficients, detection levels and recovery rates. Caffeine, theobromine, theacrine, (+)-catechin, (−)-epicatechin, (−)-epigallocatechin, (−)-epigallocatechin gallate, (−)-epicatechin gallate, (−)-gallocatechin gallate and (−)-gallocatechin were identified and quantified in the three species of genus Camellia Sect. Thea. There was 2.72% caffeine and 0.26% theobromine in C. sinensis, 4.85% theobromine in C. ptilophylla, and 1.58% theacrine, 0.94% caffeine and 0.45% theobromine in C. assamica var. kucha. Theacrine in C. sinensis and C. ptilophylla, and caffeine in C. ptilophylla were not detected. These data highlight differences in the relative proportions of purine alkaloids in the three species of Camellia Sect. Thea. In addition, different catechins were identified and quantified. The highest content of catechin in dry leaves was (−)-epigallocatechin gallate (EGCG) 3.51%, (−)-gallocatechin gallate (GCG) 9.88% and (−)-epigallocatechin gallate (EGCG) 6.78% in C. sinensis, C. ptilophylla and C. assamica var. kucha, respectively.     

Chemical and biological characteristics of Cuminum cyminum and Rosmarinus officinalis essential oils

Essential oils extracted by hydrodistillation from Cuminum cyminum and Rosmarinus officinalis were characterized by means of GC and GC–MS. C. cyminum and R. officinalis contained α-pinene (29.1%, 14.9%), 1,8-cineole (17.9%, 7.43%) and linalool (10.4%, 14.9%), respectively, as the major compounds. C. cyminum oil exhibited stronger antimicrobial activity than did R. officinalis oil against E. coli, S. aureus and L. monocytogenes. Complete death time on exposure to Cuminum cyminum L. and Rosmarinus officinalis L. oils were 20 and 25 min 180 and 240 min and 90 and 120 min for E. coli, S. aureus and L. monocytogenes, respectively. Radical-scavenging and antioxidant properties were tested by means of 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay and the β-carotene bleaching test. These properties were compared to those of Thymus x-porlock essential oil, used as a reference ingredient. The radical scavenging performance of the rosemary oil was better than that of C. cyminum. Results from the antioxidant test were better than those provided by the radical-scavenging activity. C. cyminum and R. officinalis essential oils may be considered as potent agents in food preservation.     

Sucrose hydrolysis by gelatin-immobilized inulinase from Kluyveromyces marxianus var. bulgaricus

The crude cell-free medium from a culture of Kluyveromyces marxianus var. bulgaricus was immobilized in a gelatin-water support, with an immobilization yield of 82.60% for inulinase activity. The optimum pH for both free and immobilized inulinase was the same (3.5) and the optimum temperatures were 55 °C for the free and 60 °C for the immobilized enzyme. The Arrhenius plots were linear and activation energies were 56.20 (free enzyme) and 20.27 kJ/mol K (immobilized enzyme). The kinetic parameters were calculated by Lineweaver–Burk plots and the V_max and K_m were 37.60 IU/mg protein and 61.83 mM for the free inulinase and 31.45 IU/mg protein and 149.28 mM for the immobilized enzyme, respectively. The operational stability of the immobilized inulinase was studied in a continuous fixed-bed column reactor for 33 days, at the end of which the sucrose conversion was 58.12%.     

Diversity of culturable microorganisms and occurrence of Listeria monocytogenes and Salmonella spp. in Tuber aestivum and Tuber melanosporum ascocarps

The aim of this study was to investigate the total mesophilic microorganisms, Pseudomonas genus, Enterobacteriaceae family, mold and yeast counts and the presence of Listeria monocytogenes and Salmonella spp on Tuber aestivum and Tuber melanosporum ascocarps. The results confirmed that the major percentage of the microorganisms, approximately 9.0 log ufc/g, were present in the peridium, the glebas of healthy truffles being practically free of microorganisms. The predominant microbial group was the Pseudomonas averaging 8.3 and 8.4 log cfu/g on T. aestivum and T. melanosporum whole ascocarps, respectively. The Enterobacteriaceae also achieved high populations, especially in T. aestivum truffles, with 6.3 log cfu/g. Molds and yeasts never exceeded 5.0 log cfu/g. The characterization of the isolates revealed that the fluorescens pseudomonads were the most prevalent. Raoultella terrigena and Enterobacter intermedius were the dominant Enterobacteriaceae. The identification of the yeast isolates revealed five species: Debaryomyces hansenii, Issatchenkia scutulata, Rhodotorula aurantiaca, Saccharomyces dairensis and Trichosporon beigelii subspecies A and B. The mold genera detected in both species of truffles were Aspergillus, Cladosporium, Penicillium and Fusarium, Trichoderma being present only in T. aestivum. L. monocytogenes was found in 10% of the samples of T. aestivum analysed but Salmonella spp. was not detected. Knowledge of the microbial population in terms of possible food borne and pathogen microorganisms is very useful for establishing successful disinfection and storage methods to prolong the shelf-life of ascocarps of T. aestivum and T. melanosporum.     

Isolation of some active compounds from Origanum vulgare L. ssp. vulgare and determination of their genotoxic potentials

Origanum vulgare L. (oregano) is a flavoring herb widely used around the world. In the present study, crude extract of O. vulgare L. (oregano) and its petroleum ether, chloroform, ethyl acetate, n-butanol, water fractions were prepared in order to isolate and investigate antimutagenic compounds from the active extract through the bacterial genotoxicity assay guided fractionation procedures. The methanol extract and its n-butanol fraction showed significant antimutagenic activity. In the end of sub-fractionation process of the n-butanol extract, luteolin 7-O-glucuronide and luteolin 7-O-xyloside were isolated. These compounds showed significant antimutagenic activity against 9-Aminoacridine and N-Methyl-N′-nitro-N-nitrosoguanidine mutagenicity. The inhibition rates ranged from 22.1% (luteolin 7-O-xyloside: Salmonella typhimurium TA1537 – 0.4 mM/plate) to 67.8% (luteolin 7-O-glucuronide: S. typhimurium TA1537 – 0.8 μM/plate). In conclusion, the results revealed that luteolin 7-O-glucuronide and luteolin 7-O-xyloside are two of the antimutagenic compounds of O. vulgare L. ssp. vulgare.     

Chemical composition and biological activities of Algerian Thymus oils

The compositions of essential oils isolated from nine samples of three Thymus species (Thymus algeriensis, Thymus pallescens and Thymus dréatensis) were analysed by GC and GC–MS, and a total of 114 components were identified. T. pallescens collected from various regions showed a great similarity in their compositions and were characterised by carvacrol (44.4–57.7%), p-cymene (10.3–17.3%) and γ-terpinene (10.8–14.2%) as the major components for four samples; only one sample was thymol-rich (49.3%) with a small amount of carvacrol (9.0%). On the other hand, T. algeriensis showed a chemical polymorphism, even for samples from the same location, and two new chemotypes for this species were proposed. Oxygen-containing monoterpenes were the predominant class (76.3%) in T. dreatensis oil, with linalool (30.4%), thymol (20.2%) and geraniol (19.6%) as the principal constituents. The oils were screened for their possible antioxidant activities by four complementary assays, namely DPPH free radical scavenging, hydroxyl radical scavenging, inhibition of lipid peroxidation and reducing power. The two new chemotypes of T. algeriensis exhibited strong hydroxyl radical scavenging (IC₅₀ = 2.2–3.3 μg/ml), but were not or only slightly active against the other radicals and exhibited a weak reducing power. Despite their chemical similarity, T. pallescens oils sometimes produced significant differences in their antioxidant activities. The essential oils were also screened for their antimicrobial activity against five bacteria (three Gram-positive and two Gram-negative) and one yeast (Candida albicans). The tested essential oils showed antimicrobial activity against the microorganisms used, in particular against two important pathogens, C. albicans and Helicobacter pylori.     

In vitro antimicrobial activity of essential oil from endemic Origanum minutiflorum on ciprofloxacin-resistant Campylobacter spp.

This study evaluated the antimicrobial activities of an essential oil of Origanum minutiflorum (O. Schwarz and P.H. Davis) against ciprofloxacin-resistant Campylobacter spp., by broth microdilution and agar well-diffusion methods. Moreover, O. minutiflorum oil was analyzed by gas chromatography/mass spectrometry (GC/MS). Twenty-nine components were identified, representing 98.7 of the oil. The oil yield from the plants was 4.0–4.4% v/w. The major components of O. minutiflorum oil were carvacrol (73.9%) and p-cymene (7.20%). The oil has lower contents of carvacrol methyl ether (0.05%), heptadecanol (0.06%) and carvacryl acetate (0.06%). Twenty-one Campylobacter spp. (12 C. jejuni, 5 C. lari and 4 C. coli) strains using in this study were selected among 300 isolates according to their resistance to ciprofloxacin. The minimum inhibitory concentration (MIC) values for bacterial strains, which were sensitive to the essential oil of O. minutiflorum, were in the range of 7.8–800 μg/ml. The essential oil obtained showed strong antimicrobial activity against all of the tested ciprofloxacin-resistance Campylobacter spp. These results suggest that the essential of O. minutiflorum may be used as a natural preservative in food against food-born disease, such as Campylobacteriosis.     

Growth response and modelling of the effects of Zataria multiflora Boiss. essential oil, pH and temperature on Salmonella Typhimurium and Staphylococcus aureus

The effects of different concentrations of Zataria multiflora Boiss. essential oil (EO: 0.0, 0.015, 0.03 and 0.06%), pH values (7.3 and 6.0) and storage temperatures (T: 15, 25 and 35 °C) on some growth kinetics, time-to-detection (TTD, time to the nearest visible detection) and log₁₀ probability percentage (log P%) of growth initiation of Salmonella Typhimurium and Staphylococcus aureus in Brain Heart Infusion broth were evaluated in a factorial design study. For the log P%, the effects of another variable, storage time (D, up to 43 days) was also assessed as an additional factor. The TTD and log P% of both organisms were significantly affected (P<0.01) by the values of EO, pH, T and D (just for log P%). The combinations of pH=7.3, and EO?0.015 could not obviously affect the growth kinetics of the organisms in this study. Whereas, strong inhibitory action was observed by increasing of EO concentration to 0.06% at the considered pH (7.3) and T (?35 °C). The growth of both organisms was completely inhibited at the combinations of EO=0.06%, pH=6.0 and up to 43 days of storage. Regression equations were derived relating TTDs to EO, pH and T and log P% of both organisms to EO, pH, T and D. From these models the values of predicted TTD and log P% of S. Typhimurium and St. aureus can be calculated by any combinations of EO, pH, T and D (for log P%) within the limits studied. However, between the two TTD and log P% models obtained for both organisms in this study, the TTD models of S. Typhimurium and St. aureus showed better prediction (R²=0.951 and 0.978, respectively) than the log P% models of them (R²=0.852 and 0.804, respectively).     

Characterisation of esterolytic activity from two wild mushroom species, Amanita vaginata var. vaginata and Tricholoma terreum

Characterisation of esterase activities from the edible mushroom species, Amanita vaginata var. vaginata and Tricholoma terreum, were investigated. Native electrophoresis of the crude extracts prepared from both mushroom samples showed the presence of esterolytic activities. The extracts had the greatest activity in the presence of p-nitrophenyl butyrate (pNPB) as a substrate. pH and temperature optima were found to be 8.0 and 30 °C for both enzymes, respectively. V_max and K_m values were determined as 14.2 U/l and 71 μM for A. vaginata var. vaginata and 34.6 U/l and 9.6 μM for T. terreum, respectively. The pH-stability profile showed a stationary line between 3.0 and 10.0 for both enzymes. The esterolytic activities from the extracts were maintained between 10 and 40 °C for 4 h and started to decrease at 50 °C. The effects of EDTA, NaN₃, DTT and PMSF on the enzyme activity were also investigated.     

Kinetic modeling of spice extraction from S. aromaticum and C. cassia

A mathematical model was developed for the extracts obtained from Syzygium aromaticum and Cinnamomum cassia with different particle size, solvent–solid ratios on extraction yield. Different particle sizes in the range of 2.8 mm to ?0.5 mm were employed and maximum extraction efficiency was achieved with particles of size ?0.5 mm. Among the solvent–solid ratios (20:1, 30:1, 40:1 and 50:1) ratio of 50:1 showed higher extraction yield. In the extraction kinetics, higher effective diffusivity value of 36.01 × 10⁻¹⁰ m²/s for S. aromaticum and 26.78 × 10⁻¹⁰ m²/s for C. cassia were achieved. Antioxidant values were determined and extracts prepared from ethanol showed higher scavenging activities for S. aromaticum and C. cassia as 78% and 85% respectively. Maximum phenolic content of 1.6 and 12.4 mg GAE/g of sample were achieved for S. aromaticum and C. cassia by hexane and water respectively.     

Characteristics of native and enzymatically hydrolyzed Zea mays L., Fritillaria ussuriensis Maxim. and Dioscorea opposita Thunb. starches

Comparative studies on glucoamylase hydrolysis of A-type Zea mays L., B-type F. ussuriensis Maxim., and C-type Dioscorea opposita Thunb. were carried out by scanning electron microscope (SEM), X-ray diffractometer (XRD), Fourier transform infrared (FT-IR) spectra and differential scanning calorimetry (DSC). Maize, Fritillaria, Dioscorea starches were hydrolyzed with glucoamylase for 2, 4, 8, 12 and 24 h, respectively. The SEM and XRD results revealed that A-type, B-type starch and C-type starch displayed different hydrolysis mechanisms. A-type starch was digested with enzyme penetrating into starch granules through natural pores on the surface and disrupted the interior of the starch granules. The glucoamylase worked by attacking the surface of B-type starch and forming cracks. When endo-corrosion occurred, the internal part of the granule was corroded through small cracks. However, the glucoamylase primarily attacked the interior of the C-type starch granules and then the exterior of starch granules. FT-IR confirmed that the amorphous regions in the starch granules are firstly hydrolyzed and could be hydrolyzed completely as long as the hydrolysis time is sufficient. The transition temperatures and enthalpy of gelatinization (ΔH_gel) were determined using DSC. According to the gelatinization parameters, it could be further proved amorphous and crystalline structures were hydrolyzed.     

Comparative chemical composition,antioxidant and hypoglycaemic activities of Juniperus oxycedrus ssp. oxycedrus L. berry and wood oils from Lebanon

Juniper (Juniperus oxycedrus) is used in European cuisine for its distinguishing flavour. J. oxycedrus ssp. oxycedrus berry and wood essential oils were tentatively identified by GC and GC/MS. Fifty compounds were identified in the berry oil and 23 compounds were identified in the wood oil. The J. oxycedrus ssp. oxycedrus berry oil was characterised by high contents of α-pinene (27.4%) and β-myrcene (18.9%). Other important compounds were α-phellandrene (7.1%), limonene (6.7%), epi-bicyclosesquiphellandrene (2.3%) and δ-cadinene (2.2%) while, in the wood oil, δ-cadinene (14.5%) is a major main component, together with cis-thujopsene (9.2%) and α-muurolene (4.9%). In vitro evaluation of antioxidant activity by the DPPH method showed a significant activity for both oils with IC₅₀ values of 1.45 μl/ml for wood and 7.42 μl/ml for berries. Hypoglycaemic activity was investigated through the inhibition of α-amylase. The results revealed that oil obtained by hydrodistillation from J. oxycedrus ssp. oxycedrus wood exhibits an interesting activity with IC₅₀ of 3.49 μl/ml.     

Optimization of the production of Momordica charantia L. Var. abbreviata Ser. protein hydrolysates with hypoglycemic effect using Alcalase

Momordica charantia L. Var. abbreviata Ser. protein was hydrolyzed using six different proteases. The results showed Alcalase 2.4L to have the best hydrolyzing capacity, followed by Pancreatin. In addition, Alcalase hydrolysate had stronger hypoglycemic effect than that of Pancreatin hydrolysate at the same dose. Alcalase was chosen to produce M. charantia L. Var. abbreviata Ser. protein hydrolysates (MCPHs) with hypoglycemic effect. Response surface methodology (RSM) was applied to optimize the hydrolysis conditions using Alcalase. Model equation was proposed with regard to the effect of enzyme/substrate ratio, pH and temperature. The optimum values for enzyme/substrate ratio, pH and temperature were found to be 2.37%, 9.2 and 57 °C respectively.     

Effect on lamb meat quality of including thyme (Thymus zygis ssp. gracilis) leaves in ewes’ diet

The aim of this study was to investigate the effect of including thyme leaves (TL) in the diet of pregnant sheep on the sensorial characteristics, bacterial spoilage and oxidative stability of lamb meat stored in modified atmosphere (70% O₂:30% CO₂). For this, thirty-six sheep were randomly assigned to three groups: control (basal diet), T₁ (3.7% thyme leaves), T₂ (7.5% thyme leaves). Meat spoilage (TV, PSY, MY, ENT, and LA), TBARS, CIELAB coordinates, metmyoglobin and the sensory characteristics of fresh lamb meat were analyzed on days 0, 7, 14 and 21. The presence of antioxidant compounds in the diet containing TL delayed (P < 0.05) colour deterioration, lipid oxidation and bacterial counts, while at the same time imparting a better appearance to the fresh lamb meat. In general, this effect was more pronounced at the higher level of TL (7.5%). High Pearson’s correlation coefficients were found between the sensory attributes, CIELAB coordinates and TBARS.