Lactoperoxidase-mediated degradation of single-walled carbon nanotubes in the presence of pulmonary surfactant

Carbon nanotubes (CNTs) may elicit inflammatory responses following pulmonary exposure. Conversely, enzymatic biodegradation of CNTs by inflammatory cells has also been reported. The aim of this study was to study the degradation of oxidized single-walled CNTs (ox-SWCNTs) by lactoperoxidase (LPO), a secreted peroxidase present in the airways, and whether pulmonary surfactant affects this biodegradation. To this end, ox-SWCNTs were incubated in vitro with recombinant bovine LPO + H₂O₂ + NaSCN in the presence and absence of porcine lung surfactant (Curosurf®) and biodegradation was monitored using UV–Vis–NIR spectroscopy, Raman spectroscopy, and scanning electron microscopy. The interaction of recombinant LPO with bundles of ox-SWCNTs was confirmed by atomic force microscopy. Cell-free biodegradation of ox-SWCNTs was also observed ex vivo in murine bronchoalveolar lavage fluid in the presence of H₂O₂ + NaSCN. Our study provides evidence for biodegradation of ox-SWCNTs with a lung surfactant ‘bio-corona’ and expands the repertoire of mammalian peroxidases capable of biodegradation of ox-SWCNTs. These findings are relevant to inhalation exposure to these materials, as LPO serves as an important component of the airway defense system.     

Lactoperoxidase activity in milk is correlated with somatic cell count in dairy cows

Lactoperoxidase (LPO) is a milk protein with antimicrobial function. The present study was undertaken to examine the correlation between LPO activity and somatic cell count (SCC) in milk to use LPO activity as an indicator of mastitis. Composite milk of 36 cows and quarter milk of 3 cows were collected once per week from 0 to 300 d postpartum and twice per day for 1 wk, respectively. For the measurement of LPO activity, milk was mixed with tetramethylbenzidine solution and incubated at 37°C for 30 min, followed by the measurement of optical density. When only milk with low SCC (132 ± 12 × 10³ cells/mL) was used, a significant decrease in LPO activity was detected in primiparous cows from 0 to 4 mo postpartum. Lactoperoxidase activities of primiparous cows in mo 1, 2, and 3 postpartum were significantly higher than those in multiparous cows. When composite milk was divided based on LPO activity, the SCC was significantly higher in the groups with LPO activity >5 and from 3 to 3.9 U/mL in the second- and fourth-parity cows, respectively, compared with the group with LPO activity <2 U/mL. Extremely high SCC were found in the ≥fifth-parity cows, even in low-LPO activity groups. In the case of quarter milk, higher LPO activity was associated with increased SCC in all 3 cows. The percentage of quarter milk samples with high SCC (4,062 ± 415 × 10³ cells/mL) increased with an increase in the LPO activity. The percentage of quarter milk samples with high SCC was 50.0 to 100% in the milk with LPO activity ≥5 U/mL. These results indicate that the correlation of LPO activity to the SCC in bovine milk may point to the potential use of the former as an indicator of SCC.     

Antimicrobial activity of nisin, reuterin, and the lactoperoxidase system on Listeria monocytogenes and Staphylococcus aureus in cuajada, a semisolid dairy product manufactured in Spain

The inhibitory activity of nisin (N), reuterin (R), and the lactoperoxidase system (LPS), added individually or in combination, against Listeria monocytogenes and Staphylococcus aureus was investigated in “cuajada” (curdled milk), a semisolid dairy product manufactured in Spain. Cuajada was manufactured from UHT skim milk separately inoculated with L. monocytogenes and Staph. aureus, each at approximately 4 log cfu/mL, and held under conditions of temperature abuse (10°C). On d 3, a synergistic bactericidal activity was observed for the combinations of biopreservatives assayed, with L. monocytogenes counts of only 0.30 log cfu/mL in cuajada made with N + R + LPS vs. 8.31 log cfu/mL in control cuajada. After 12 d, L. monocytogenes could not be detected in cuajada made with added N + LPS or N + R + LPS. Staphylococcus aureus was more resistant than L. monocytogenes to biopreservatives added individually. On d 3, the synergistic effect of the 3 biopreservatives against Staph. aureus resulted in counts of 3.03 log cfu/mL in cuajada made with N + R + LPS vs. 6.40 in control cuajada. After 12 d, Staph. aureus counts were 2.61 log cfu/mL in cuajada made with N + R + LPS, whereas they ranged from 6.11 to 7.70 log cfu/mL in control cuajada and in cuajada made with other combinations of biopreservatives. The most pronounced decrease in pathogen counts was achieved by the triple combination N + R + LPS, which acted synergistically on the inactivation of L. monocytogenes and Staph. aureus in cuajada over 12 d at 10°C. The treatment combining these 3 natural biopreservatives at low concentrations, within the hurdle concept of food preservation, might be a useful tool to control the growth of pathogenic microorganisms in nonacidified dairy products.     

Utilization of microfiltration or lactoperoxidase system or both for manufacture of Cheddar cheese from raw milk

The objective of the present study was to determine if application of microfiltration (MF) or raw milk lactoperoxidase system (LP) could reduce the risk of foodborne illness from Escherichia coli in raw milk cheeses, without adversely affecting the overall sensory acceptability of the cheeses. Escherichia coli K12 was added to raw milk to study its survival as a non-pathogenic surrogate organism for pathogenic E. coli. Five replications of 6 treatments of Cheddar cheese were manufactured. The 6 treatments included cheeses made from pasteurized milk (PM), raw milk (RM), raw milk inoculated with E. coli K12 (RME), raw milk inoculated with E. coli K12 + LP activation (RMELP), raw milk inoculated with E. coli K12 + MF (MFE), and raw milk inoculated with E. coli K12 + MF + LP activation (MFELP). The population of E. coli K12 was enumerated in the cheese milks, in whey/curds during cheese manufacture, and in final Cheddar cheeses during ripening. Application of LP, MF, and a combination of MF and LP led to an average percentage reduction of E. coli K12 counts in cheese milk by 72, 88, and 96%, respectively. However, E. coli K12 populations significantly increased during the manufacture of Cheddar cheese for the reasons not related to contamination. The number of E. coli K12, however, decreased by 1.5 to 2 log cycles during 120 d of ripening, irrespective of the treatments. The results suggest that MF with or without LP significantly lowers E. coli count in raw milk. Hence, if reactivation of E. coli during cheese making could be prevented, MF with or without LP would be an effective technique for reducing the counts of E. coli in raw milk cheeses. The cheeses were also analyzed for proteolysis, starter and nonstarter lactic acid bacteria (NSLAB), and sensory characteristics during ripening. The concentration of pH 4.6 soluble nitrogen at 120 d was greater in PM cheese compared with the other treatments. The level of 12% trichloroacetic acid-soluble nitrogen at 120 d was greater in RM, RME, and RMELP cheeses compared with PM, MFE, and MFELP cheeses. This could be related to the fact that cheeses made from raw milk with or without LP (RM, RME, and RMELP) had greater levels of NSLAB compared with PM, MFE, and MFELP cheeses. Cheeses at 60 d, as evaluated by 8 trained panelists, did not differ in bitterness, pastiness, or curdiness attributes. Cheeses at 120 d showed no differences in acid-taste, bitterness, or curdiness attributes. Sensory analysis at 60 d showed that PM and MFELP cheeses had greater overall sensory acceptability than RM and RME cheeses. The overall sensory acceptability of the cheeses at 120 d showed that PM, MFE, and MFELP cheeses were more acceptable than RM and RME cheeses.     

Inhibition of Penicillium commune by Edible Whey Protein Films Incorporating Lactoferrin, Lacto-ferrin Hydrolysate, and Lactoperoxidase Systems

ABSTRACT: Effects of lactoferrin (LF), lactoferrin hydrolysate (LFH), and lactoperoxidase systems (LPOS), both directly and incorporated into edible whey protein isolate (WPI) films, on the inhibition of Penicillium commune were studied. Mechanical, oxygen-barrier, and color properties of WPI films with and without LPOS were also compared. Antimicrobial effects were examined by turbidity, disc diameter, surface spreading, and film surface inoculation tests. Film elastic modulus, tensile strength, percent elongation, oxygen permeability, and Hunter L, a , and b values were tested. LF and LFH at 10 mg/mL or higher inhibited P. commune in 1% peptone water but not in potato dextrose broth. WPI films incorporating LPOS inhibited growth of P. commune . The properties of WPI films were not significantly changed by incorporation of LPOS ( P > 0.05).     

乳过氧化物酶的分离纯化和酶学性质研究

采用超滤-离子交换色谱分步洗脱法,对牛初乳中的乳过氧化物酶进行了分离和纯化。经SDS-PAGE测定,分离出的乳过氧化物酶显示为单一区带,相对分子质量为75035D。该酶酶活回收率为76.17%,其最适pH为5.0～5.5,最适温度为55℃。在70℃、75℃时LP酶活的热失活曲线呈现一般植物过氧化物酶失活的双相特征。     

Psychrotrophs in dairy products: Their effects and their control

Health concerns and technological effects of psychrotrophic bacteria in dairy products are reviewed, as well as methods to control their presence and development. The various Gram‐negative and Gram‐positive psychrotrophic species are listed and, with respect to pathogenic psychrotrophs, emphasis is given on Listeria monocytogenes, Yersinia enterocolytica, and Bacillus cereus. The influence of psychrotrophic bacteria on the quality of raw milk, pasteurized and UHT milks, butter, ice cream, cheese, and powders is examined. Public health considerations of Listeria monocytogenes, Yersinia enterocolytica, and Bacillus cereus of these various dairy products are also presented. Methods that can be used to eliminate or control the development of psychrotropic bacteria include low or high temperatures, chemicals, gases, the lactoperoxidase system, lactic acid bacteria, microfiltration, bactofugation, lactoferrin‐related proteins, sanitation, flavors, and naturally occurring spore germinants.     

口服牛初乳及其组分的保健功能

综述了口服牛初乳的动物、人体功能性实验结果。基于牛初乳内各种生长因子、乳铁蛋白与乳过氧化物酶体系，口服牛初乳具有抗感染、强化机体免疫力、促进生长发育、改善运动性能、促进胃肠组织创伤愈合等功能。     

乳源抗菌物质对益生菌生长的影响

含益生菌的乳制品研究开发是一种发展趋势,而天然乳源抗菌组分对这些益生菌的影响尚缺乏系统性研究。综述了4种主要的乳源抗菌物质(免疫球蛋白、乳铁蛋白、乳过氧化物酶、溶菌酶)对益生菌生长的影响及其抗菌机理研究进展。     

Characterizing endogenous and exogenous peroxidase activity for bleaching of fluid whey and retentate

The lactoperoxidase (LP) system may be used to achieve the desired bleaching of fluid whey with the addition of low concentrations (<50 mg/kg) of hydrogen peroxide. The addition of an exogenous peroxidase (EP) to whey may also be used to aid in whey bleaching when the LP system is not fully active. The objectives of this study were to monitor LP activity in previously refrigerated or frozen milk, fluid whey, and whey retentate (10% solids) and to evaluate peroxidase activity in fluid whey and whey retentate (10% solids), with and without additional EP (2, 1, or 0.5 dairy bleaching units), over a range of pH (5.5–6.5) and temperatures (4–60°C). Subsequent experiments were conducted to determine the relationship between enzyme activity and bleaching efficacy. Raw and pasteurized milk, fat-separated pasteurized whey, and whey retentate (10% solids) were evaluated for LP activity following storage at 4 or −20°C, using an established colorimetric method. A response surface model was applied to evaluate both endogenous and EP activity at various temperatures and pH in freshly manufactured whey and retentate. Refrigerated or frozen storage at the parameters evaluated did not affect LP activity in milk, whey, or retentate. In fluid whey, with and without added EP, as pH decreased (to 5.5) and temperature increased (to 60°C), peroxidase activity increased. Retentate with EP exhibited behavior similar to that of fluid whey: as pH decreased and temperature increased, activity increased. However, in retentate without EP, as pH increased and temperature increased, activity increased. Enzyme activity was negatively correlated to bleaching time (time for >80% norbixin destruction) in fluid whey but a linear relationship was not evident in retentate. When fluid whey is bleached enzymatically, if pH is decreased and temperature is increased, the rate of reaction increases (e.g., bleaching occurs in less time). When bleaching in retentate, a higher pH (pH 6.5 vs. pH 5.5) is desired for optimal bleaching by the LP system. Due to processing restraints, this may not be possible for all dairy producers to achieve and, thus, addition of EP could be beneficial to improve bleaching efficacy.