Использование микробных биосенсоров для анализа углеводов, ксенобиотиков и параметров состояния окружающей среды тема диссертации и автореферата по ВАК РФ 03.01.06, кандидат биологических наук Ильясов, Павел Владимирович

• Ильясов, Павел Владимирович
• кандидат биологических наук
• 2012

Оглавление диссертации кандидат биологических наук Ильясов, Павел Владимирович

Содержание.

1. ОБЩАЯ ХАРАКТЕРИСТИКА РАБОТЫ.

1.1. Актуальность проблемы.

1.2. Цель и задачи исследования.

1.3. Научная новизна работы.

1.4. Практическая значимость работы.

2. ОБЗОР ЛИТЕРАТУРЫ.

2.1. Общие сведения о микробных сенсорах.

2.2. Используемые типы преобразователей.

2.3. Строение биорецептора.

2.4. Микробные биосенсоры и нанотехнологии.

2.5. Методы повышения селективности анализа.

2.6. Практическое использование микробных сенсоров.

2.6.1. Детекция углеводов.

2.6.2. Анализ спиртов и органических кислот.

2.6.3. Определение антибиотиков.

2.6.4. Детекция аминокислот, холестерина и других соединений, не представляющих опасности с точки зрения окружающей среды.

2.6.5. Использование микробных биосенсоров в природоохранных целях - общие аспекты.

2.6.6. Определение ароматических соединений.

2.6.7. Определение ионов неорганических кислот и тяжелых металлов.

2.6.8. Анализ других экологически опасных соединений.

2.6.9. Определение общей токсичности и генотоксичности.

2.6.10. Биосенсорное определение БПК.

3. МАТЕРИАЛЫ И МЕТОДЫ.

3.1. Регистрация сигналов.

3.2. Особенности использованных в работе штаммов. Культивирование клеток.

3.3. Иммобилизация микроорганизмов.

3.4. Гипероксигенация измерительного раствора.

3.5. Обработка данных.

4. РЕЗУЛЬТАТЫ.

4.1. Сенсор для определения углеводов на основе О. охуйат.

4.1.1. Скрининг штаммов рода аисопоЬа^ег.

4.1.2. Исследование влияния концентрации растворенного кислорода на сигналы амперометрического биосенсора.

4.1.3. Использование сенсора на основе (7. охус1ат для анализа глюкозы в сыворотке крови человека.

4.1.4. Оценка содержания глюкозы в ферментационной среде.

4.2. Микробный биосенсор для детекции анионных ПАВ.

4.2.1. Скрининг штаммов и общие характеристики моделей.

4.2.2. Оценка оптимального метода иммобилизации.

4.2.3. Оценка возможности качественного анализа проб сложного состава.

4.2.4. Тестирование моделей сенсоров на образцах модельных и реальных сточных вод

4.3. Детекция нафталина.

4.4. Детекция ионов нитрита.

4.5. Создание БПК-сенсора на основе клеток Агхи1а айептоуогат и оценка его характеристик.

4.5.1. Проведение испытаний анализаторов на модельных и реальных образцах.

5. ВЫВОДЫ.

Заключение диссертации по теме «Биотехнология (в том числе бионанотехнологии)», Ильясов, Павел Владимирович

5. ВЫВОДЫ

1. Создана и исследована модель микробного биосенсора на основе G. oxydans для детекции ряда углеводов и спиртов. Модель характеризуется чувствительностью к этанолу, глюкозе, ксилозе, арабинозе, глицерину и сорбиту. Показана перспективность данной модели как возможного прототипа анализатора глюкозы. Продемонстрирована возможность использования созданного биосенсора для анализа проб сыворотки крови.

2. Протестирован ряд моделей микробных биосенсоров для детекции анионных и неионогенных ПАВ. Наиболее перспективным среди них признан сенсор на основе штамма P. rathonis Т, характеризующийся преимущественной чувствительностью анализа к ДСН и АБС. Упомянутый сенсор был использован для анализа модельных сточных вод, содержащих ПАВ.

3. Произведена оценка возможности повышения селективности биосенсорной детекции нафталина. Найдено, что элиминантный штамм P. putida BS238 при культивировании на бензоате обладает субстратной специфичностью, близкой к специфичности того

JIIII» ■1 ■ 1 '-1-1-1IIIIIIIIIIIII1 1 1 ■ ■IIIIIIL же штамма, несущего плазмнду pBS2; вместе с тем, наличие ответа на нафталин не позволяет использовать биосенсор на основе элиминантного штамма в качестве электрода сравнения в системе для детекции нафталина. Представляются перспективными дальнейшие исследования, направленные на поиск штаммов, позволяющих осуществлять дифференциальную детекцию нафталина.

4. Продемонстрирована эффективность модели микробного биосенсора для детекции ионов нитрита на основе миксотрофного штамма N. vulgaris. Показана возможность биосенсорной детекции нитрита в диапазоне концентраций 0.01-10 мМ, что в сочетании с высокой селективностью также делает разработанную модель перспективной как прототип промышленно выпускаемого анализатора.

5. Искусственное повышение концентрации растворенного кислорода приводит к значительному увеличению сигналов сенсора при высокой плотности иммобилизованных в биорецепторе клеток и/или больших концентрациях детектируемого субстрата, т. е. в случае, когда биорецепторный элемент находится в условиях недостатка кислорода. При малых концентрациях иммобилизованной биомассы и субстрата упомянутый эффект незначителен. Полученный результат расширяет представления о режимах функционирования биорецепторного элемента.

6. Разработанный на основе культуры A. adeninovorans биосенсорный анализатор БПК характеризуется широкой субстратной специфичностью и устойчивостью к действию токсических компонентов муниципальных сточных вод и позволяет выполнять оценку БПК в речной воде и сточных водах различного происхождения.

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