Ascorbic acid, carotenoids, and visual quality of baby spinach as affected by shade netting and postharvest storage

Baby spinach ( Spinacia oleracea L.) was grown under three types of shade netting (high transmittance, spectrum-altering, and low transmittance) to study the effect on the concentrations of vitamin C (ascorbic acid and dehydroascorbic acid), carotenoids, and chlorophyll and on the visual quality of the leaves. The spinach was sown in April and August and harvested at two growth stages. After harvest, leaves were stored in polypropylene bags at 2 and 10 degrees C. Shading significantly decreased the ascorbic acid concentration of April-sown spinach by 12-33%, but in the August-sown spinach, the response was inconsistent. Concentrations of total carotenoids and total chlorophylls were significantly higher under the nettings in many cases, especially under the spectrum-altering and low-transmittance nettings. Postharvest visual quality and postharvest persistence of the compounds analyzed were not greatly affected by shading. We conclude that these shade nettings are acceptable to use in baby spinach production when it comes to the studied aspects of internal and external quality of the produce.     

Determination of theaflavins including methylated theaflavins in black tea leaves by solid-phase extraction and HPLC analysis

A quantitative method for four theaflavins and two methylated theaflavin derivatives in black tea leaves was developed by solid-phase extraction and a high-performance liquid chromatographic method with photodiode array detection. The theaflavins in black tea leaves were extracted three times with 40 vol 50% aqueous ethanol (mg dry tea powder/mL) containing 2% ascorbic acid. The ethanol extracts were diluted 4-fold with distilled water. All diluted extracts were directly applied to the solid-phase C18 cartridge column without concentration. The fraction of theaflavins was obtained by 40% ethanol extraction after rinsing with water followed with 15% ethanol extraction. An aliquot of theaflavins after concentration was injected onto an ODS C18 reversed-phase column, and four theaflavins and two methylated theaflavins were sufficiently separated by a linear gradient system using distilled water and acetonitrile with 0.5% acetic acid. This analytical method is sensitive for the determination of a small amount of methylated theaflavins, since various interfering substances produced during the fermentation process were eliminated in advance by solid-phase extraction. Using this analytical method, we also demonstrated that methylated theaflavins were easily produced during the manufacture of black tea.     

Phylogeny of Fusarium oxysporum f. sp. lactucae Inferred from Mitochondrial Small Subunit, Elongation Factor 1-alpha, and Nuclear Ribosomal Intergenic Spacer Sequence Data

ABSTRACT Fusarium oxysporum f. sp. lactucae, causal agent of Fusarium wilt of lettuce, is a serious pathogen recently reported in Arizona. Sequence analysis of the mitochondrial small subunit (mtSSU), translation elongation factor 1-alpha (EF-1alpha) gene, and the nuclear ribosomal DNA intergenic spacer (IGS) region was conducted to resolve relationships among f. sp. lactucae isolates, F. oxysporum isolates from other hosts, and local non-pathogenic isolates. Analysis of mtSSU sequences provided limited phylogenetic resolution and did not differentiate the lactucae isolates from 13 other F. oxysporum isolates. Analysis of EF-1alpha sequences resulted in moderate resolution, grouping seven formae speciales with the lactucae isolates. Analysis of the IGS region revealed numerous sequence polymorphisms among F. oxysporum formae speciales consisting of insertions, deletions, and single nucleotide transitions and substitutions. Repeat sequence analysis revealed several duplicated subrepeat units that were distributed across much of the region. Based on analysis of the IGS sequence data, lactucae race 1 isolates resolved as a monophyletic group with three other formae speciales of F. oxysporum. In all analyses, lactucae race 2 isolates composed a separate lineage that was phylo-genetically distinct and distantly related to the lactucae race 1 isolates.     

Molecular Characterization and Diagnosis of QoI Resistance in Cucumber and Eggplant Fungal Pathogens

ABSTRACT The molecular mechanism of QoI fungicide resistance was studied using isolates of cucumber Corynespora leaf spot fungus (Corynespora cassiicola) and the eggplant leaf mold (Mycovellosiella nattrassii). In both pathogens, a mutation at position 143 from glycine to alanine (G143A) was detected in the cytochrome b gene that encodes for the fungicide-targeted protein. Moreover, the nucleotide sequence at amino acid position 143 was converted from GGT or GGA in sensitive (wild-type) to GCT or GCA in resistant (mutant-type) isolates. The methods of polymerase chain reaction restriction fragment length polymorphism commonly used for QoI resistance monitoring were employed successfully, leading to the amplified gene fragment from resistant isolates being cut with the restriction enzyme ItaI. However, heteroplasmy (the coexistence of wild-type and mutated alleles) was found when the resistant isolates of C. cassiicola, M. nattrassii, and Colletotrichum gloeosporioides (strawberry anthracnose fungus) were subcultured in the presence or absence of QoI fungicides. QoI resistance of cucumber powdery and downy mildew isolates persisted for a few years following the removal of the selection pressure imposed by the fungicide under both laboratory and commercial greenhouse conditions. The proportion of mutated sequences in cytochrome b gene decreased over time in the pathogen population. The protective efficacy of the full dose of azoxystrobin decreased when the populations of powdery and downy mildews contained resistant isolates at 10%. Using FMBIO, a fluorescence bio-imaging analyzer, the mutant allele from the QoI-resistant isolates could be detected at the level of 1%, whereas the detection sensitivity of ethidium-bromide-stained gels was approximately 10 times lower.     

The bioactivity of gonadotropin releasing hormones and its regulation in the gilthead seabream,Sparus aurata: in vivo andin vitro studies

Thein vivo andin vitro potency of native and modified forms of gonadotropin releasing hormone (GnRH) to release gonadotropin (GtH) was studied inSparus aurata and correlated with their relative susceptibility to degradation by cytosolic-bound enzymes of the pituitary, kidney, and liver. Salmon (s) GnRH and luteinizing hormone-releasing hormone (LHRH) are equipotent whereas analogs of these peptides ((D-Arg⁶-Pro⁹NET)-sGnRH, (D-Ala⁶-Pro⁹NET)-LHRH, (D-Trp⁶)-LHRH) are superactive in inducingin vivo GtH release (at 10 μg/kg body weight). In anin vitro superfusion system of pituitary fragments all analogs are equipotent to the native peptides (at 10⁻¹⁰ to 2.5 × 10⁻⁷M). sGnRH and LHRH are rapidly degraded by cytosolic peptidases of the pituitary, liver, and kidney. The preferred site of cleavage is the Tyr⁵-Gly⁶ bond. Substitution of the position 6 glycine by D-amino acids renders the 5–6 bond resistant to degradation and shifts the main site of cleavage to the Pro⁹-Gly¹⁰NH₂ bond. Substitution at position 6 (as above) and at position 10 with Pro⁹NET results in analogs that are resistant to degradation. We propose that enzymatic cleavage terminates GnRH bioactivityin vivo and thus increased resistance to degradation is a major determinant of GnRH analog superactivity.     

Hormonal changes during meiotic maturation and ovulation in the brook trout (Salvelinus fontinalis)

The plasma levels of estradiol-17 (E2), 17, 20-dihydroxy-4-pregnen-3-one (17,20-P) and gonadotropin (GTH) were measured in brook trout (Salvelinus fontinalis) during the period from the end of vitellogenesis to postovulation. Blood samples were taken according to specific stages of maturation, including germinal vesicle breakdown (GVBD) and ovulation. E2 levels were quite high (45 ng/ml) at the end of vitellogenesis (and prior to GVBD) and dropped precipitously by GVBD (2 ng/ml). They remained low through ovulation and postovulation. 17,20-P levels were low prior to GVBD (0.7 ng/ml) and increased dramatically at GVBD (148 ng/ml). The levels of 17,20-P remained high at ovulation (142 ng/ml) and then dropped significantly within 24 h to approximately half of the ovulatory values. They decreased even further by 7 days postovulation. GTH levels rose gradually through GVBD and ovulation from a postvitellogenic level of approximately 3 ng/ml to a 7 day postovulatory value of approximately 10 ng/ml. The overall results; 1) decrease in estradiol prior to GVBD, 2) increase in 17,20-P at GVBD and 3) gradual GTH rise through GVBD and ovulation, are similar to those reported for other salmonids.     

Advanced broodstock diets for the mangrove red snapper and a potential importance of arachidonic acid in eggs and fry

Mangrove red snapper fed advanced broodstock diets containing squid meal and squid oil exhibited higher hatching rates, cumulative survival and survival activity index than those fed a basal diet or a basal diet supplemented with mixture of antioxidants. On the other hand, fatty acid analyses of ovaries and fry of wild fish and eggs and larvae of broodstock fed raw fish revealed high arachidonic acid (ARA) and docosahexaenoic acid (DHA) levels and relatively lower eicosapentaenoic acid (EPA) levels consequently showing high ARA/EPA and DHA/EPA ratios compared to cold water species. This suggests that ARA may be nutritionally more important for egg and larval development and survival in tropical marine fish and its supplementation in broodstock diets may enhance reproductive performance of mangrove red snapper.     

Molecular cloning of the cDNA encoding the β subunit of thyrotropin and regulation of its gene expression by thyroid hormones in the goldfish, Carassius auratus

A cDNA encoding the subunit of thyrotropin (TSH) was isolated from a goldfish (Carassius auratus) pituitary gland cDNA library. By comparing the sequence with other teleost TSHs, a signal peptide of 19 amino acids and a mature hormone of 131 amino acids were predicted for goldfish TSH subunits. The resulting putative mature hormone of 131 amino acids had well-conserved cysteine positions and a putative N-linked glycosylation site; homology was 51–67% with TSHs from other teleosts, 38–43% with tetrapod TSHs, but only 27 and 29% with goldfish GTH-I and -II, respectively. We also examined the effects of thyroid hormones (TH) and thiourea (TU, an inhibitor of TH production) treatments on TSH and GTH subunit gene expressions in the goldfish pituitary gland. After thyroxine (T₄) treatment, circulating T₄ concentration increased and TSH mRNA level decreased. Supressing the amount of circulating T₄ and triiodothyronine (T₃) by TU treatment increased the TSH mRNA level. Moreover, T₄ replacement therapy (simultaneous treatment of both TU and T₄) caused a high level of circulating T₄ and a low level of circulating T₃, and a decrease in the TSH mRNA level. Thus, changing levels of circulating TH exert a negative feedback on the level of TSH subunit mRNA in goldfish in vivo. On the other hand, GTH subunit mRNA levels were not affected by changes in the levels of circulating TH.