It has been demonstrated that freezing-induced acidity changes have an impact on the structural integrity, degree of aggregation, and chemical stability of frozen food and pharmaceutical products. The stability of the compounds in solutions is maintained by the presence of buffers. However, many buffers are unsuitable for applications involving freezing as this process substantially alters the acidity. In this study, we determine the effect of initial pH, concentration, and cooling rate on the freezing-induced change in acidity of phosphate buffered saline (PBS) in the frozen state via UV-VIS spectroscopy. Furthermore, we examine the impact of individual salts present in PBS and discuss the mechanisms affecting the resulting acidity that we approximate via Hammett acidity function (H2-).

• Klíčová slova
• Acidification, Buffering capacity, Freezing, PBS, Preferential precipitation,
• MeSH
• fosfáty * chemie   MeSH
• koncentrace vodíkových iontů MeSH
• pufry MeSH
• solný roztok * chemie   MeSH
• zmrazování MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfáty * MeSH
• pufry MeSH
• solný roztok * MeSH

• MeSH
• aminokyseliny MeSH
• Bacillus megaterium cytologie   účinky léků   růst a vývoj   metabolismus   MeSH
• bakteriální proteiny biosyntéza   MeSH
• bakteriální RNA biosyntéza   MeSH
• bakteriologické techniky MeSH
• buněčná stěna MeSH
• buněčné dělení MeSH
• DNA bakterií biosyntéza   MeSH
• elektronová mikroskopie MeSH
• fosfáty * MeSH
• glykosaminoglykany biosyntéza   MeSH
• hypertonické roztoky farmakologie   MeSH
• izotopy uhlíku MeSH
• koncentrace vodíkových iontů MeSH
• kultivační média MeSH
• kyseliny pimelové metabolismus   MeSH
• leucin metabolismus   MeSH
• pufry * MeSH
• thymidin metabolismus   MeSH
• uracil metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• aminokyseliny MeSH
• bakteriální proteiny MeSH
• bakteriální RNA MeSH
• DNA bakterií MeSH
• fosfáty * MeSH
• glykosaminoglykany MeSH
• hypertonické roztoky MeSH
• izotopy uhlíku MeSH
• kultivační média MeSH
• kyseliny pimelové MeSH
• leucin MeSH
• pufry * MeSH
• thymidin MeSH
• uracil MeSH

Solutions of three Good's buffers (HEPES, MOPS, and MES), both pure and mixed with sodium phosphate buffers (Na-P), are investigated in terms of the freezing-induced acidity changes in their operational pH ranges. The Good's buffers have the tendency to basify upon freezing and, more intensively, at lower pHs. The acidity varies most prominently in MES, where the change may reach the value of two. Importantly, the Good's buffers are shown to mitigate the strong acidification in the Na-P buffer. Diverse concentrations of the Good's buffers are added to cancel out the strong, freezing-induced acidity drop in 50 mM Na-P that markedly contributes to the solution's acidity; the relevant values are 3 mM HEPES, 10 mM MOPS, and 80 mM MES. These buffer blends are therefore proposed to be applied in maintaining approximately the acidity of solutions even after the freezing process and, as such, should limit the stresses for frozen chemicals and biochemicals.

• Klíčová slova
• Cooling rate, Cryopreservation, Freeze-concentrated solution, Hammett acidity function, Stabilization, Sulfonephthalein indicators, pH shift,
• MeSH
• fosfáty * MeSH
• ionty MeSH
• koncentrace vodíkových iontů MeSH
• pufry MeSH
• zmrazování MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfáty * MeSH
• ionty MeSH
• pufry MeSH
• sodium phosphate MeSH Prohlížeč

The concept of "pH memory" has been established in the literature for the correlation between the pH of a pre-lyophilization solution and the ionization state of freeze-dried powder (lyophile). In this paper, the concept of "pH memory" is explored for the system of an aqueous solution, a frozen solution, and a lyophile. Sodium and potassium phosphate buffers in the pH range of 5-9 were frozen and lyophilized with sulfonephthalein indicators as acidity probes, and their Hammett acidity functions were compared to the initial pH of the aqueous solution. The results show that the acidities of the lyophiles are somewhat changed compared to the initial pHs, but the acidities in the frozen state differ more substantially. The Hammett acidity functions of the frozen buffers were found to be markedly dissimilar from the initial pH, especially in the sodium phosphate frozen at 233K, where an increase in the initial pH led to a decrease in the Hammett acidity function of the frozen state at a certain pH range. The large acidification observed after freezing the sodium phosphate buffer was not detected in the lyophiles after the sample had been dried; the phenomenon is explained considering the formed crystals analyzed by X-ray powder diffraction. The results suggest that monitoring the final acidity of a lyophile is not sufficient to predict all the acidity changes throughout the whole lyophilization process. The importance of well-controlled freezing and lyophilization conditions follows from the results of the research.

• Klíčová slova
• Formulation buffers, Freezing, Freezing stress, Lyophilization, Protein cryopreservation, pH memory, pH shift,
• MeSH
• fosfáty chemie   MeSH
• koncentrace vodíkových iontů * MeSH
• lyofilizace MeSH
• pufry * MeSH
• zmrazování MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• fosfáty MeSH
• pufry * MeSH

• MeSH
• anestetika lokální * MeSH
• fosfáty * MeSH
• isotonické roztoky * MeSH
• kokain farmakologie   MeSH
• koncentrace vodíkových iontů MeSH
• králíci MeSH
• pufry MeSH
• rohovka účinky léků   MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• anglický abstrakt MeSH
• časopisecké články MeSH
• Názvy látek
• anestetika lokální * MeSH
• fosfáty * MeSH
• isotonické roztoky * MeSH
• kokain MeSH
• pufry MeSH

Humanin (HN), Met-Ala-Pro-Arg-Gly-Phe-Ser-Cys-Leu-Leu-Leu-Leu-Thr-Ser-Glu-IIe-Asp-Leu-Pro-Val-Lys-Arg-Arg-Ala, recently discovered in the human brain, is an important neuroprotective peptide. Some derivatives of HN show even higher biological activity, for example [G-14]-HN, where Ser at position 14 is replaced with Gly. As structurally related HN peptide derivatives have similar chemical properties, their separation by CE is difficult. In this work, the electrophoretic behaviour of HN derivatives including [G-14]-HN, a tryptophan HN derivative [W-14]-HN, several other HN derivatives and HN fragments was studied. While phosphate buffer was used as the general BGE, the effects of the buffer concentration and various additives were examined, including sulphate, heptane sulphonate, 2-morpholinoethanesulphonic acid N-[tris(hydroxymethyl)methyl]-2-aminoethane sulphonic acid (TES), sulphated-beta-CD and beta-CD. Separation efficiency of 200,000 theoretical plates was achieved in a BGE of 80 mM phosphate at pH 2.5 where seven out of nine major peaks were partially separated. By investigating the influence of concentration of the interrogated ions on peptides migration, the association between positively charged protonated sites of peptides and various anions was proved. Especially a strong interaction with phosphate, sulphate and sulphonate groups was established. Conditional stability constant of the [Pep(z+), (H(2)PO(4)(-))(n)](z - n) ion associate (n = 1) for [G-14]-HN equals to log K approximately 1.78.

• MeSH
• alkylsulfonany chemie   MeSH
• beta-cyklodextriny chemie   MeSH
• elektroforéza kapilární metody   MeSH
• fosfáty chemie   MeSH
• intracelulární signální peptidy a proteiny chemická syntéza   chemie   izolace a purifikace   MeSH
• koncentrace vodíkových iontů MeSH
• lidé MeSH
• molekulární sekvence - údaje MeSH
• neuropeptidy chemická syntéza   chemie   izolace a purifikace   MeSH
• peptidové fragmenty chemická syntéza   chemie   izolace a purifikace   MeSH
• pufry MeSH
• roztoky MeSH
• sekvence aminokyselin MeSH
• sírany chemie   MeSH
• substituce aminokyselin MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alkylsulfonany MeSH
• beta-cyklodextriny MeSH
• fosfáty MeSH
• humanin MeSH Prohlížeč
• intracelulární signální peptidy a proteiny MeSH
• neuropeptidy MeSH
• peptidové fragmenty MeSH
• pufry MeSH
• roztoky MeSH
• sírany MeSH

Changes in the protonation state of lyophilized proteins can impact structural integrity, chemical stability, and propensity to aggregate upon reconstitution. When a buffer is chosen, the freezing/drying process may result in dramatic changes in the protonation state of the protein due to ionization shift of the buffer. In order to determine whether protonation shifts are occurring, ionizable probes can be added to the formulation. Optical probes (dyes) have shown dramatic ionization changes in lyophilized products, but it is unclear whether the pH indicator is uniform throughout the matrix and whether the change in the pH indicator actually mirrors drug ionization changes. In solid-state NMR (SSNMR) spectroscopy, the chemical shift of the carbonyl carbon in carboxylic acids is very sensitive to the ionization state of the acid. Therefore, SSNMR can be used to measure ionization changes in a lyophilized matrix by employing a small quantity of an isotopically-labeled carboxylic acid species in the formulation. This paper compares the apparent pH of six trehalose-containing lyophilized buffer systems using SSNMR and UV-Vis diffuse reflectance spectroscopy (UVDRS). Both SSNMR and UVDRS results using two different ionization probes (butyric acid and bromocresol purple, respectively) showed little change in apparent acidity compared to the pre-lyophilized solution in a sodium citrate buffer, but a greater change was observed in potassium phosphate, sodium phosphate, and histidine buffers. While the trends between the two methods were similar, there were differences in the numerical values of equivalent pH (pHeq) observed between the two methods. The potential causes contributing to the differences are discussed.

• Klíčová slova
• Diffuse reflectance spectroscopy, Lyophilization, Solid-state NMR spectroscopy, Solid-state acidity, UV/VIS,
• MeSH
• fosfáty * chemie   MeSH
• histidin * chemie   MeSH
• koncentrace vodíkových iontů MeSH
• kyselina citronová chemie   MeSH
• lyofilizace * metody   MeSH
• magnetická rezonanční spektroskopie * metody   MeSH
• pufry MeSH
• spektrofotometrie ultrafialová metody   MeSH
• trehalosa * chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfáty * MeSH
• histidin * MeSH
• kyselina citronová MeSH
• pufry MeSH
• trehalosa * MeSH

The aims of the present study were to isolate new yeasts with high extracellular (exo) invertase activity and to investigate the usability of buffer systems as invertase production media by immobilized yeast cells. Among 70 yeast isolates, Cryptococcus laurentii MT-61 had the highest exo-invertase activity. Immobilization of yeast cells was performed using sodium alginate. Higher exo-invertase activity for immobilized cells was achieved in tris-sucrose buffer system (TSBS) compared to sodium acetate buffer system and potassium phosphate buffer system. TSBS was prepared by dissolving 30 g of sucrose in 1 L of tris buffer solution. The optimum pH, temperature, and incubation time for invertase production with immobilized cells were determined as 8.0, 35 °C and 36 h in TSBS, respectively. Under optimized conditions, maximum exo-invertase activity was found to be 28.4 U/mL in sterile and nonsterile TSBS. Immobilized cells could be reused in 14 and 12 successive cycles in sterile and nonsterile TSBS without any loss in the maximum invertase activity, respectively. This is the first report which showed that immobilized microbial cells could be used as a biocatalyst for exo-invertase production in buffer system. As an additional contribution, a new yeast strain with high invertase activity was isolated.

• MeSH
• biotechnologie metody   MeSH
• časové faktory MeSH
• Cryptococcus enzymologie   metabolismus   MeSH
• imobilizované buňky enzymologie   metabolismus   MeSH
• invertasa izolace a purifikace   MeSH
• koncentrace vodíkových iontů MeSH
• kultivační média chemie   MeSH
• pufry MeSH
• sacharosa MeSH
• teplota MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• invertasa MeSH
• kultivační média MeSH
• pufry MeSH
• sacharosa MeSH

• MeSH
• draslík MeSH
• fosfáty analýza   MeSH
• fotometrie * MeSH
• indikátory a reagencie analýza   MeSH
• metody MeSH
• pufry * MeSH
• sodík MeSH
• tartaráty analýza   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• draslík MeSH
• fosfáty MeSH
• indikátory a reagencie MeSH
• pufry * MeSH
• sodík MeSH
• tartaráty MeSH

The paper deals with the characterisation of the bioactive phenomena of glass-ceramic scaffold derived from Bioglass® (containing 77 wt.% of crystalline phases Na(2)O·2CaO·3SiO(2) and CaO·SiO(2) and 23 wt.% of residual glass phase) using simulated body fluid (SBF) buffered with tris-(hydroxymethyl) aminomethane (TRIS). A significant effect of the TRIS buffer on glass-ceramic scaffold dissolution in SBF was detected. To better understand the influence of the buffer, the glass-ceramic scaffold was exposed to a series of in vitro tests using different media as follows: (i) a fresh liquid flow of SBF containing tris (hydroxymethyl) aminomethane; (ii) SBF solution without TRIS buffer; (iii) TRIS buffer alone; and (iv) demineralised water. The in vitro tests were provided under static and dynamic arrangements. SBF buffered with TRIS dissolved both the crystalline and residual glass phases of the scaffold and a crystalline form of hydroxyapatite (HAp) developed on the scaffold surface. In contrast, when TRIS buffer was not present in the solutions only the residual glassy phase dissolved and an amorphous calcium phosphate (Ca-P) phase formed on the scaffold surface. It was confirmed that the TRIS buffer primarily dissolved the crystalline phase of the glass-ceramic, doubled the dissolving rate of the scaffold and moreover supported the formation of crystalline HAp. This significant effect of the buffer TRIS on bioactive glass-ceramic scaffold degradation in SBF has not been demonstrated previously and should be considered when analysing the results of SBF immersion bioactivity tests of such systems.

• MeSH
• biokompatibilní materiály * MeSH
• keramika * MeSH
• koncentrace vodíkových iontů MeSH
• mikroskopie elektronová rastrovací MeSH
• prášková difrakce MeSH
• sklo * MeSH
• spektrofotometrie atomová MeSH
• tělesné tekutiny * MeSH
• tromethamin * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• biokompatibilní materiály * MeSH
• tromethamin * MeSH