Zinc ions are shown to be an efficient inhibitor of mitochondrial cytochrome c oxidase activity, both in the solubilized and the liposome-reconstituted enzyme. The effect of zinc is biphasic. First there occurs rapid interaction of zinc with the enzyme at a site exposed to the aqueous phase corresponding to the mitochondrial matrix. This interaction is fully reversed by EDTA and results in a partial inhibition of the enzyme activity (50-90%, depending on preparation) with an effective K(i) of approximately 10 microM. The rapid effect of zinc is observed with the solubilized enzyme, it vanishes upon incorporation of cytochrome oxidase in liposomes, and it re-appears when proteoliposomes are supplied with alamethicin that makes the membrane permeable to low molecular weight substances. Zinc presumably blocks the entrance of the D-protonic channel opening into the inner aqueous phase. Second, zinc interacts slowly (tens of minutes, hours) with a site of cytochrome oxidase accessible from the outer aqueous phase bringing about complete inhibition of the enzymatic activity. The slow phase is characterized by high affinity of the inhibitor for the enzyme: full inhibition can be achieved upon incubation of the solubilized oxidase for 24 h with zinc concentration as low as 2 microM. The rate of zinc inhibitory action in the slow phase is proportional to Zn(2+) concentration. The slow interaction of zinc with the outer surface of liposome-reconstituted cytochrome oxidase is observed only with the enzyme turning over or in the presence of weak reductants, whereas incubation of zinc with the fully oxidized proteoliposomes does not induce the inhibition. It is shown that zinc ions added to cytochrome oxidase proteoliposomes from the outside inhibit specifically the slow electrogenic phase of proton transfer, coupled to a transition of cytochrome oxidase from the oxo-ferryl to the oxidized state (the F --> O step corresponding to transfer of the 4th electron in the catalytic cycle).

Purpose: To investigate empirically the energy dependence of the detector response of two in vivo luminescence detectors, LiF:Mg,Cu,P (MCP-N) high-sensitivity TLDs and Al 2 O 3 :C OSLDs, in the 40-300-kVp energy range in the context of in vivo surface dose measurement. As these detectors become more prevalent in clinical and preclinical in vivo measurements, knowledge of the variation in the empirical dependence of the measured response of these detectors across a wide spectrum of beam qualities is important. Method: We characterized a large range of beam qualities of three different kilovoltage x-ray units: an Xstrahl 300 Orthovoltage unit, a Precision x-Ray X-RAD 320ix biological irradiator, and a Varian On-Board Imaging x-ray unit. The dose to water was measured in air according to the AAPM's Task Group 61 protocol. The OSLDs and TLDs were irradiated under reference conditions on the surface of a water phantom to provide full backscatter conditions.To assess the change in sensitivity in the long term, we separated the in vivo dosimeters of each type into an experimental and a reference group. The experimental dosimeters were irradiated using the kilovoltage x-ray units at each beam quality used in this investigation, while the reference group received a constant 10 cGy irradiation at 6 MV from a Varian clinical linear accelerator. The individual calibration of each detector was verified in cycles where both groups received a 10 cGy irradiation at 6 MV. Results: The nanoDot OSLDs were highly reproducible, with AE1.5% variation in response following >40 measurement cycles. The TLDs lost~20% of their signal sensitivity over the course of the study.The relative light output per unit dose to water of the MCP-N TLDs did not vary with beam quality for beam qualities with effective energies <50 keV (~150 kVp/6 mm Al). At higher energies, they showed a reduced (~75-85%) light output per unit dose relative to 6 MV x rays.The nanoDot OSLDs exhibited a very strong (120-408%) dependency of the light output relative to 6 MV x rays. Variations up to 15% between different x-ray units with equivalent effective energies were also observed. Conclusions: While convenient for clinical use, nanoDot OSLDs exhibit a strong variation in their measured light output per unit dose relative to 6 MV in the 40-300 kV x-ray range. This variability differs unit-to-unit, limiting their effective use for in vivo dosimetry applications in the kilovoltage x-ray energy range. MCP-N TLDs offer a much more stable response, but suffer from variations in sensitivity over time dependent on radiation history, which requires careful experimental handling.

The purpose of this study was to assess the performance of structure‐guided deformable image registration (SG‐DIR) relative to rigid registration and DIR using TG‐132 recommendations. This assessment was performed for image registration of treatment planning computed tomography (CT) and magnetic resonance imaging (MRI) scans with Primovist® contrast agent acquired post stereotactic body radiation therapy (SBRT). SBRT treatment planning CT scans and posttreatment Primovist® MRI scans were obtained for 14 patients. The liver was delineated on both sets of images and matching anatomical landmarks were chosen by a radiation oncologist. Rigid registration, DIR, and two types of SG‐DIR (using liver contours only; and using liver structures along with anatomical landmarks) were performed for each set of scans. TG‐132 recommended metrics were estimated which included Dice Similarity Coefficient (DSC), Mean Distance to Agreement (MDA), Target Registration Error (TRE), and Jacobian determinant. Statistical analysis was performed using Wilcoxon Signed Rank test. The median (range) DSC for rigid registration was 0.88 (0.77–0.89), 0.89 (0.81–0.93) for DIR, and 0.90 (0.86–0.94) for both types of SG‐DIR tested in this study. The median MDA was 4.8 mm (3.7–6.8 mm) for rigid registration, 3.4 mm (2.4–8.7 mm) for DIR, 3.2 mm (2.0–5.2 mm) for SG‐DIR where liver structures were used to guide the registration, and 2.8 mm (2.1–4.2 mm) for the SG‐DIR where liver structures and anatomical landmarks were used to guide the registration. The median TRE for rigid registration was 7.2 mm (0.5–23 mm), 6.8 mm (0.7–30.7 mm) for DIR, 6.1 mm (1.1–20.5 mm) for the SG‐DIR guided by only the liver structures, and 4.1 mm (0.8–19.7 mm) for SG‐DIR guided by liver contours and anatomical landmarks. The SG‐DIR shows higher liver conformality as per TG‐132 metrics and lowest TRE compared to rigid registration and DIR in Velocity AI software for the purpose of registering treatment planning CT and post‐SBRT MRI for the liver region. It was found that TRE decreases when liver contours and corresponding anatomical landmarks guide SG‐DIR.

ВВЕДЕНИЕХромогенные субстраты ферментов часто используются для определения активностей последних с помощью фотометрических методов [1,2]. Так как данные методы широко применяются в научных исследованиях, клинической диагностике, при проведении анализов в биотехнологической промышленности, экологии и иных областях для выявления ферментов и оценки их активности, потребности в специфичных хромогенных субстратах весьма высоки, и многие из этих субстратов есть в каталогах зарубежных компаний -производителей и поставщиков реактивов.Тромбин, он же фактор свёртывания II, играет одну из ключевых ролей в процессе свёртывания крови. Нарушения процесса свёртывания, вызывающие усиленную активацию тромбина, приводят к тромбозам и являются одними из основных причин развития сердечно-сосудистых заболеваний, нарушения кровообращения и кровоснабжения органов и тканей [3][4][5]. Связь серьезных патологий с нарушением регуляции активности тромбина требует контроля активности этого фермента. Медикаментозная регуляция повышенной свёртываемости крови также должна сопровождаться контролем активности тромбина, поскольку длительное и излишнее снижение её может привести к опасным для жизни кровотечениям [5]. Помимо этого, разработка новых лекарственных средств, регулирующих активность тромбина, сопровождается исследованием их воздействия на кинетические параметры катализируемых тромбином реакций. Потребность в субстратах тромбина, позволяющих просто, надежно и в условиях прямой регистрации во времени определять активность этого фермента, привела к разработке ряда коротких пептидов, модифицированных по С-концевой карбоксильной группе остатками хромофора (п-нитроанилина) или флуорофора (7-амино-4-метилкумарина) [6,7]. п-Нитроанилидные субстраты ранее получали путём присоединения хромофора непосредственно к готовому пептиду либо к С-концевой аминокислоте в растворе [2,[8][9][10], однако разработаны и более удобные так называемые one-pot способы синтеза п-нитроанилидов пептидов на твёрдой фазе. В ряде способов аминокислотный остаток присоединяется к смоле за функциональную группу в боковой цепи [9][10][11]. Это требует обязательного наличия таких остатков вблизи С-конца пептида и использования особым образом функционализированных смол, обычно не применяемых в пептидном синтезе [11,12], что весьма неудобно и непрактично. Использование связанного со смолой п-фенилендиамина позволило проводить наращивание на нём пептидной цепи с помощью стандартных методик твердофазного пептидного синтеза с последующим окислением остатка п-аминоанилида (рАА) до п-нитроанилида (pNA) в отщепленных пептидах, однако применяемые смолы подвергались дополнительным модификациям для обеспечения возможности присоединения п-фенилендиамина [13,14]. Наиболее удобным оказался способ с модификацией остатком п-фенилендиамина смолы с легко замещаемым атомом хлора (тритилхлоридной или 2-хлортритилхлоридной, обычно используемых для синтеза защищенных пептидов) с последующим наращиванием на нём пептидной цепи с помощью Fmoc-аминокислот и мягким окислением остатка п-аминоанилида до п...

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