Use of nisin-coated plastic films to control Listeria monocytogenes on vacuum-packaged cold-smoked salmon

Cold-smoked (Salmo salar) salmon samples were surface-inoculated with a cocktail of three nisin-resistant strains of L. monocytogenes (PSU1, PSU2 and PSU21) to a level of approximately 5 x 10(2) or 5 x 10(5) CFU/cm2 of salmon surface. The inoculated smoked salmon samples were vacuum-packaged with control film (no nisin) or nisin-coated plastic films and stored at either 4 or 10 degrees C. When the inoculated smoked salmon samples were packaged with film coated with 2000 IU/cm2 of nisin, a reduction of 3.9 log CFU/cm2 (compared with control) was achieved at either temperature for samples inoculated with 5 x 10(2) CFU/cm(2 of L. monocytogenes after 56 (4 degrees C) and 49 (10 degrees C) days of storage while reductions of 2.4 and 0.7 log CFU/cm2 were achieved for samples inoculated with a high level of L. monocytogenes (5 x 10(5) CFU/cm2) after 58 (4 degrees C) and 43 (10 degrees C) days, respectively. For samples packaged in film coated with 500 IU/cm2 of nisin, reductions of 0.5 and 1.7 log CFU/cm2 were achieved for samples inoculated with a low level of L. monocytogenes (5 x 10(2) CFU/cm2) after 56 (4 degrees C) and 49 (10 degrees C) days of storage while reductions of 1.8 and 0.8 log CFU/cm2 were achieved for samples inoculated with high level of L. monocytogenes after 58(4 degrees C) and 43 (10 degrees C) days, respectively. In addition, nisin inhibited the proliferation of background microbiota on smoked salmon in a concentration-dependent manner at both storage temperatures although the bacteriostatic effect was more pronounced at refrigeration temperature. This work highlights the potential for incorporating nisin into plastic films for enhancing the microbial safety of smoked salmon as well as controlling its microbial spoilage.     

Formation of corn fiber gum-milk protein conjugates and their molecular characterization

Corn fiber arabinoxylan is hemicellulose B isolated from the fibrous portions (pericarp, tip cap, and endosperm cell wall fractions) of corn kernels and is commonly referred to as corn fiber gum (CFG). Our previous studies showed that CFG isolated from corn bran (a byproduct of corn dry milling) contains very little protein and is an inferior emulsifier for oil-in-water emulsion systems as compared to corn fiber gum isolated from corn fiber derived from the corn wet-milling process. The protein deficient CFG isolated from corn bran was covalently conjugated with byproducts of cheese processing, β-lactoglobulin (β-LG) and whey protein isolate (WPI) using an economical food-grade Maillard-type heating reaction for the purpose of increasing their commercialization potential as a food emulsifier and soluble nutritional additive in beverages. The formation of the CFG-protein conjugates has been confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). It has also been demonstrated that CFG-protein conjugates are capable of producing fine emulsions with better stability than either CFG or protein alone. The molecular characterization of CFG-protein conjugates was performed by high-performance size-exclusion chromatography (HPSEC) coupled to on-line multi-angle laser light scattering and viscometric detection. The analysis by HPSEC revealed that CFG-protein conjugates had higher weight-average molar mass (M_w, 340-359 kDa) and polydispersity (M_w/M_n, 1.74) than the corresponding unconjugated CFG (M_w, 290 kDa and M_w/M_n, 1.35). The z-average root-mean-square radius of gyration (R_gz) of CFG-protein conjugates was slightly higher (30.5-33.5 nm) in comparison to CFG (29.5 nm) but their weight-average-intrinsic viscosity (η) remained unchanged (about 1.32 dL g⁻¹). The Mark-Houwink exponent “a” of conjugates (0.40) was lower than the unconjugated CFG (0.53) indicating the formation of a more compact structure after conjugation with protein. These findings should benefit the beverage industry, which can use this information to produce a high quality emulsifier from the low-value byproducts of corn dry milling and cheese processing.     

O6-alkylguanine-DNA alkyltransferase inactivation by ester prodrugs of O6-benzylguanine derivatives and their rate of hydrolysis by cellular esterases

To modulate the bioavailability and perhaps improve the tumor cell selectivity of O6-alkylguanine-DNA alkyltransferase (AGT) inactivators, pivaloyloxymethyl ester derivatives of O6-benzylguanine (BG) were synthesized and tested as AGT inactivators and as substrates for cellular esterases. The potential prodrugs examined were the 7- and 9-pivaloyloxymethyl derivatives of O6-benzylguanine (7- and 9-esterBG), and of 8-aza-O6-benzylguanine (8-aza-7-esterBG and 8-aza-9-esterBG) and the 9-pivaloyloxymethyl derivative of 8-bromo-O6-benzylguanine (8-bromo-9-esterBG). The benzylated purines were all potent inactivators of the pure AGT and of the AGT activity in HT29 cells and cell extracts. Each ester was at least 75 times less potent than the corresponding benzylated purine against the pure human AGT. In contrast, the activities of esters and their respective benzylated purine were similar in crude cell extracts and in intact cells. The increase in potency of esters in cellular extracts could be explained by a conversion of the respective prodrug to the more potent benzylated purine in the presence of cellular esterases. The apparent catalytic activity (Vmax/Km) of liver microsomal esterase for 8-azaBG ester prodrugs was 70-130 times greater than for BG prodrugs and 10-20 times greater than for 8-bromo-9-esterBG. Tumor cell hydrolysis of the esters varied considerably as a function of cell type and prodrug structure. These data suggest that these or related prodrugs may be advantageous for selective AGT inactivation in certain tumor types.     

The human serum paraoxonase/arylesterase gene (PON1) is one member of a multigene family

A physiological role for paraoxonase (PON1) is still uncertain, but it catalyzes the hydrolysis of toxic organophosphates. Evidence that the human genome contains two PON1-like genes, designated PON2 and PON3, is presented here. Human PON1 and PON2 each have nine exons, and the exon/intron junctions occur at equivalent positions. PON1 and PON2 genes are both on chromosome 7 in human and on chromosome 6 in the mouse. Turkey and chicken, like most birds, lack paraoxonase activity and are very susceptible to organophosphates. However, they have a PON-like gene with approximately 70% identity with human PON1, PON2, and PON3. Another unexpected finding is that the deduced amino acid sequences of PON2 in human, mouse, dog, turkey, and chicken and of human PON3 are all missing the amino acid residue 105, which is lysine in human PON1. The expanded number of PON genes will have important implications for future experiments designed to discover the individual functions, catalytic properties, and physiological roles of the paraoxonases.     

Thapsigargin and receptor-mediated activation of Drosophila TRPL channels stably expressed in a Drosophila S2 cell line

The Drosophila melanogaster genes, transient receptor potential (trp) and transient receptor potential-like (trpl) encode putative plasma membrane cation channels TRP and TRPL, respectively. We have stably co-expressed Drosophila TRPL with a Drosophila muscarinic acetylcholine receptor (DM1) in a Drosophila cell line (S2 cells). Basal Ca2+ levels measured using Fura-2/AM in unstimulated S2-DM1-TRPL cells were low and indistinguishable from untransfected cells, indicating that the TRPL channels were not constitutively active in this expression system. Activation of DM1 receptor in S2-DM1-TRPL cells by 100 microM carbamylcholine induced Ca2+ release from an intracellular Ca2+ pool followed by a Gd(3+)-insensitive Ca2+ influx. Pretreatment of S2-DM1-TRPL cells with 10 microM atropine abolished Gd(3+)-insensitive Ca2+ influx triggered by carbamylcholine, but the response was not blocked by prior incubation with pertussis toxin. TRPL channels could also be reliably activated by bath application of 1 microM thapsigargin for 10 min or 100 nM thapsigargin for 60 min in Ca(2+)-free solution. In some cells, TRPL channels activated by thapsigargin could further be activated by carbamylcholine. The findings suggest that, when stably expressed in the S2 cell line, TRPL may be regulated by two distinct mechanisms: (i) store depletion; and (ii) stimulation of DM1 receptor via pertussis-toxin insensitive G-protein (or the subsequent activation of PLC), but without further requirement for Ca2+ release.     

Structure of tetrameric human phenylalanine hydroxylase and its implications for phenylketonuria

Phenylalanine hydroxylase (PheOH) catalyzes the conversion of L-phenylalanine to L-tyrosine, the rate-limiting step in the oxidative degradation of phenylalanine. Mutations in the human PheOH gene cause phenylketonuria, a common autosomal recessive metabolic disorder that in untreated patients often results in varying degrees of mental retardation. We have determined the crystal structure of human PheOH (residues 118-452). The enzyme crystallizes as a tetramer with each monomer consisting of a catalytic and a tetramerization domain. The tetramerization domain is characterized by the presence of a domain swapping arm that interacts with the other monomers forming an antiparallel coiled-coil. The structure is the first report of a tetrameric PheOH and displays an overall architecture similar to that of the functionally related tyrosine hydroxylase. In contrast to the tyrosine hydroxylase tetramer structure, a very pronounced asymmetry is observed in the phenylalanine hydroxylase, caused by the occurrence of two alternate conformations in the hinge region that leads to the coiled-coil helix. Examination of the mutations causing PKU shows that some of the most frequent mutations are located at the interface of the catalytic and tetramerization domains. Their effects on the structural and cellular stability of the enzyme are discussed.     

Intrathyroidal parathyroid glands can be a cause of failed cervical exploration for hyperparathyroidism

BACKGROUND: The incidence of intrathyroidal parathyroid glands remains controversial. The purpose of this study was to determine the incidence in a series of patients with hyperparathyroidism. METHODS: Three hundred nine patients underwent parathyroidectomy. Patients were divided into two groups: uniglandular disease versus hyperplasia. RESULTS: Eighteen of 309 patients (6%) had abnormal intrathyroidal parathyroid glands. The incidence was 3% (7 of 222) in patients with uniglandular disease versus 15% (11 of 73) in those with hyperplasia. With a mean follow-up of 54 months, 12 patients are eucalcemic, 5 have persistent hypocalcemia, and 1 has recurrent hypercalcemia. There were no recurrent laryngeal nerve injuries. CONCLUSIONS: These data suggest that an intrathyroidal adenoma is an uncommon cause of failure, whereas abnormal intrathyroidal parathyroid tissue may be a more common cause of failure in patients with hyperplasia.     

Free amino acid supplementation to steers: effects on ruminal fermentation and performance

Three studies were conducted to evaluate amino acid utilization by cattle. In Exp. 1, five steers (580 kg) were fed 86% rolled corn diets with mixtures of amino acids containing up to 6 g/d DL-Met, 24 g/d L-Lys, 6 g/d L-Thr, and 3 g/d L-Trp. Treatments had little effect on ruminal fermentation, diet digestibility, N flow to the duodenum, or microbial efficiency. Ruminal concentrations of Met and Lys increased linearly (P < .05) with amino acid supplementation, whereas Thr responded quadratically, and Trp was not altered. In Exp. 2, four steers (414 kg) were used to measure effects of dietary monensin or laidlomycin propionate in high-grain diets supplemented with amino acids. Ionophores had no significant effect on ruminal fermentation or outflows of amino acids from the rumen. In Exp. 3, 100 steers (287 kg initial BW) were fed diets containing 1% of a nonprotein N source. Treatments were 1) no supplemental N (UREA), 2) UREA plus soybean meal (SBM), 3) UREA plus 2 g/d DL-Met, 8 g/d L-Lys, 2 g/d L-Thr, and 1 g/d L-Trp, or 4) UREA plus 4 g/d DL-Met, 16 g/d L-Lys, 4 g/d L-Thr, and 2 g/d L-Trp. During the growing period (diets based on whole-plant milo silage), gains were higher for SBM-supplemented steers than for UREA steers and intermediate for steers supplemented with amino acids. Few significant differences in performance were observed among treatments during the finishing phase (diets based on dry-rolled corn) or for the entire experiment, but cattle fed SBM or amino acids tended to be fatter and have better marbling scores and quality grades. Amino acids did not greatly alter ruminal fermentation or cattle performance.