Гальченко Е.С. “Влияние замораживания и сублимационного высушивания на сохранность молозива коров”.-Рукопись.
Диссертация на соискание ученой степени кандидата биологических наук по специальности 03.00.19 – криобиология. – Институт проблем криобиологии и криомедицины НАН Украины, Харьков, 2003.
В работе исследовалось влияние замораживания, сублимационного высушивания и условий хранения молозива коров на его рН, кислотность и характер перекисных процессов в липидах молозива, а также на сохранность антиоксидантных свойств лактоферрина, входящего в его состав. Была также исследована эффективность сублимационно высушенного молозива при экспериментальном сахарном диабете и влияние лактоферрина на жизнеспособность и репарационные процессы в ДНК меланомных клеток линии Arn8 после УФ облучения.
Установлено, что исследованные показатели качества молозива после замораживания, сублимационного высушивания и хранения сухого молозива расфасованным в желатиновые капсулы при 4°С не изменяются на протяжении 12 мес.
На модельных системах показано, что лиофилизация молозива не изменяет хелаторные и антиоксидантные свойства лактоферрина.
Результаты исследования эффективности сухого молозива при сахарном диабете показали, что его пероральное введение способствует уменьшению концентрации глюкозы в крови и интенсивности ПОЛ в организме.
Установлено также, что ЛФ проявляет цитотоксическое действие по отношению к меланомным клеткам Arn8 в культуре. При этом он не способствует продвижению клеток с поврежденной ДНК в цикл пролиферации.
Полученные в работе результаты могут быть положены в основу разработки технологии получения пищевой добавки из молозива коров, а также при определении показаний к его применению.
Ключевые слова: молозиво коров, замораживание, сублимационное высушивание, лактоферрин, перекисное окисление липидов, гипотермическое хранение, хелатор, сахарный диабет, ультрафиолетовое облучение.
Galchenko K.S. “Effect of freezing and lyophilization on the integrity of bovine colostrum”.- Manuscript.
Thesis for conferring a scientific degree of Candidate of Biological Sciences, speciality 03.00.19- Cryobiology.-Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv, 2003.
The effect of freezing, lyophilization and storage conditions on bovine colostrum (first milk) quality has been studied in the work. The quality of native, frozen and lyophilized bovine colostrum was estimated by such indices as pH, acidity and intensity of induced chemilumunescence (Chl). It was defined that duration of native colostrum storage at positive temperatures (20°C and 4°C) is limited by 7 and 14 days correspondingly. Storage of colostrum at -12°C is advisable not longer than 35 days. It should be noted that the storage at -196°C does not result in negative changes of quality indices during this storage term. It allows to recommend this temperature for long-term storage of the product. The Chl method may be used as the express-method of estimation of changes in lipids of colostrum under its storage and processing. Lyophilization of colostrum was done with the device for lyophilization made in Constructing and Technical Bureau with Design Unit of the Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine. The lyophilized colostrum was packed into gelatinous capsules and then to plastic containers and stored in refrigerator at 4°C. It was shown that storing of lyophilized colostrum under these conditions does not result in statistically significant decrease of quality indices up to 12 months term.
Studies on the effect of freezing and lyophilization of colostrum on the integrity of biological properties of lactoferrin (LF) were carried out.
Antioxidative properties of LF were studied in the model systems containing Fe2+, H2O2, mannitol, egg yolk lipoproteids, rat liver homogenate using Chl method. It was shown that LF derived from native, frozen and lyophilized colostrum is an efficient iron chelator. LF reduces the concentration of hydroxyl radicals and thus decreases the intensity of peroxidative processes in the model systems. Freezing, lyophilization and storage of lyophilized bovine colostrum within 12 months under certain conditions does not lead to the decrease of LF antioxidative properties.
Researches on the effect of LF derived from native and lyophilized bovine colostrum on the intensity of peroxidative processes under hypothermal storage (4°C) of rat liver fragments have been carried out. The intensity of peroxidation processes in liver fragments was estimated with Chl method as well as by TBA-active products (TBAAP) accumulation with spectrophotometric method.
The presence of LF in the storage medium reduced the development of peroxidative processes and exhaustion of endogenic antioxidative systems up to 48 h of hypothermal storage, and up to 72 h storage in the case of LF extra adding to the medium.
The efficiency of the lyophilized bovine colostrum under experimental diabetes mellitus in rats has been also investigated. It was shown that in the experimental group of rats (which were given colostrum during one month before alloxan injection and during all experiment) the death rate and the number of animals with diabetes were lower than in the group of diabetic rats. The dynamics of the glucose level in blood and the intensity of lipid peroxidation in blood and liver were studied on the 7th, 15th and 30th days. Glucose level was estimated using the chromooxygenic method. Lipid peroxidation intensity was defined with the Chl method and by the initial, spontaneous, Fe2+- and ascorbate-induced TBAAP level. It was shown that introduction of colostrum to the animals has protective effect decreasing glycemia level in blood and lipid peroxidation activation in blood and liver. Thus, the lyophilized bovine colostrum possesses protective activity under experimental diabetes mellitus and could be used in curing this disease.
The comparative studies on the effect of LF from bovine colostrum (bLF) and human LF (hLF) on the viability of Arn8 melanoma cell line in vitro have been also carried out. It was defined that the presence of bLF in the incubation medium resulted in much more loss of cell viability (80%) than in cells incubated with hLF (10-20%). It testifies to the fact that bLF is more cytotoxic to melanoma cells in dose-dependent manner.
LF effect on Arn8 cells with intact DNA as well as on the processes of DNA reparation after damaged effect of ultraviolet (UV) irradiation has been studied. Irradiation of cells with low dose of UVB (wavelengths 280-315 nm) resulted in decreasing cell number by 70%. The number of cells with damaged DNA (“blue cells”) made 20% 24 h after irradiation. Incubation of cells in the medium with LF did not lead to increasing or decreasing “blue cells” number. After irradiation of cells with UVA (wavelengths 315-400 nm) the dose-dependence of total cell number in culture and “blue cell” number was noted. LF adding to the incubation medium had no effect on these indices. This testifies to the fact that LF has no direct protective effect on Arn8 cell viability and cell genomic material under damaged effect of UV radiation. At the same time LF does not change the number of cells with damaged DNA arrested in G1-phase and, thus, does not promote the progression of cells with damaged genome into S-phase and duplication of the DNA.
Key words: bovine colostrum, freezing, lyophilization, lactoferrin, lipid peroxidation, hypothermal storage, chelator, diabetes mellitus, ultraviolet irradiation.
Відповідальний за випуск Г.О. Бабійчук
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Підписано до друку 15.04.2003 р. Формат 60x90 1/16
Тираж 100 прим. Авт.арк. 0.9. Замовлення №742
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