Микробиологическая утилизация полимерных отходов

С развитием химической промышленности в биосферу стало регулярно поступать более тысячи различных ксенобиотиков, которые в значительной степени загрязняют окружающую среду.  Известно, что соединения, вносимые человеком в окружающую среду, а именно: инсектициды, гербициды, полимерные детергенты и другие ксенобиотики помимо токсичности, очень устойчивы в среде, что представляет опасность для человека и животных. В настоящее время нагрузка на процессы самоочищения биосферы является избыточными, вследствие чего параллельно с деструкцией загрязнений идёт их постепенное накопление. Таким образом, биодеградация ксенобиотиков становится одной из важных проблем защиты биосферы.

Биодеградация — разрушение сложных веществ в результате деятельности живых систем. Основными биологическими системами, осуществляющими биоразрушения, являются микроорганизмы, обладающие огромным разнообразием ферментных систем и большой лабильностью метаболизма. Именно они способны разлагать широкий спектр химически устойчивых соединений, тем самым, возвращая основные питательные элементы в глобальные циклы и предотвращая накопление «мертвых» остатков в биосфере.

Наиболее активно участвуют в разрушении ксенобиотиков бактерии и грибы, основное количество которых выделено из почвы и воды. Представители бактерий относятся к различным родам Грам — отрицательных и Грам — положительных аэробных и анаэробных организмов. Из наиболее важных аэробных Грам — отрицательных бактерий следует отметить виды родов Pseudomonas, Sphingomonas, Burkholderia, Alcaligenes, Acinetobacter, Flavobacterium, метанокисляющие и нитрифицирующие бактерии, а из Грам — положительных – представителей родовArthrobacter, Nocardia, RhodococcusиBacillus.

Грибы, способные аэробно разрушать такие соединения, относятся к родам Phanerochaete(возбудители «белой гнили»), Penicillium, Aspergillus, Trichoderma, Fusarium (Шлегель,1987).

Особую актуальность, разрушающая способность микроорганизмов приобрела в последние десятилетия в связи с увеличивающимся присутствием в биосфере устойчивых загрязнителей антропогенного происхождения, причем в масштабах, превышающих природную самоочищающую способность. Человеку удалось создать такие соединения, которые не разрушаются в природе в обычных условиях. Это различные синтетические полимеры, красители, пестициды, фармацевтические препараты, моющие средства и т.д. Эти ксенобиотики имеют уникальную биологическую активность уже на уровне микропримесей. В широком смысле к ксенобиотикам могут быть отнесены и вещества природного происхождения, но полученные в сверхколичествах и перемещенные в несвойственные им места (например, нефть). Большинство таких соединений обладает значительной стабильностью, и для их полного разложения при обычных условиях требуются столетия. Происходит непрерывный перенос этих веществ по пищевым цепям и их накопление на конечных этапах, к которым относится и человек. Огромное количество ксенобиотиков чрезвычайно токсично и проявляет мутагенную, канцерогенную, аллергенную и тератогенную активности. Однако понятно, что человечество не может полностью отказаться от использования таких веществ, так как они применяются практически во всех областях деятельности.

Современная стратегия в решении экологических проблем, связанных с загрязнением земли, заключается в восстановлении хозяйственной деятельности на загрязненных территориях, а не в непрерывном освоении все новых чистых территорий.

Очевидно, что развитие эффективных технологий восстановления загрязненных земель, рассчитанных на длительную перспективу, зависит от сложного сочетания различных факторов, таких, как законодательная деятельность, политические инициативы, общественное мнение и подходящие технологии.

Мировое производство одного из наиболее распространенных и доступных полимеров — полиэтилена достигает сегодня десятков миллионов тонн в год. Гигантские молекулы этого полимера, называемые макромолекулами, построены из метиленовых -CH2- групп: … -CH2-CH2-CH2- … -CH2- …, объединенных в линейные цепи. Число фрагментов -CH2-CH2-, называемых мономерными звеньями, изменяется для промышленно производимых полимеров этилена от 1000 до 10000. Значительное количество полиэтилена перерабатывается в пленочные материалы, с которыми мы встречаемся в быту на каждом шагу. В первую очередь это упаковочные материалы — пакеты и сумки для продуктов. Это также различные контейнеры для хранения разнообразных жидкостей — от воды до минеральных масел, например, для моторных масел, или смесей воды и этиленгликоля, используемых в качестве охлаждающих и незамерзающих жидкостей в современных автомобилях.

Отслужившие свой срок, выполнившие свои функции, загрязненные пакеты и контейнеры выбрасываются. Каждый из нас практически ежедневно видит эти выброшенные и ставшие бесполезными вещи в своем доме, на улице, за городом, в лесу. Они не только создают неудобства в обыденной жизни, но и наносят вред окружающей природе, замусоривая землю и препятствуя росту растений из-за нарушения воздухо- и влагообмена в почве.

При указанных масштабах производства только полиэтилена, производимого за год, вполне хватило бы, чтобы покрыть пленкой толщиной 0,05 мм (это типичная пленка), территорию, равную Франции, а если учесть накопленные за последние 5 лет отходы, то и всю Европу. Если же добавить сюда и другие крупнотоннажные полимеры, такие, как полипропилен, полистирол, полиамиды полиэфиры, например, лавсан, каучуки и резины и многие другие, то общее количество отходов индустрии полимеров многократно возрастет.

Промышленные синтетические полимеры, получаемые полимеризацией реакционноспособных соединений — мономеров, являются весьма устойчивыми химическими соединениями. Многие из них, такие, как полиэтилен, способны выдерживать воздействие солнечного излучения и кислорода воздуха в совокупности с воздействием тепла и влаги в природных условиях в течение десятков лет без заметного химического разрушения.

Для борьбы с загрязнением природы, связанным с производством полимеров используют следующие подходы:

  1. Уничтожение отработавших и выброшенных полимеров методом сжигания. Однако при этом уничтожаются в принципе ценные вещества и материалы. Продуктами сжигания в лучшем случае являются вода и углекислый газ, а это значит, что не удается вернуть даже исходных мономеров, полимеризацией которых получали уничтожаемые полимеры. Кроме того, выделение в атмосферу больших количеств углекислого газа CO2 приводит к глобальным нежелательным эффектам, в частности к парниковому эффекту. При этом также образуются вредные летучие вещества, которые загрязняют воздух и, соответственно, воду и землю. В случае поливинилхлорида — это разнообразные низкомолекулярные хлорированные органические вещества, отличающиеся высокой токсичностью, в том числе канцерогенностью. Образуется и газообразный хлористый водород, который, растворяясь в воде, дает соляную кислоту. В окружающую среду выделяются разнообразные соединения, включающие тяжелые металлы, используемые в качестве катализаторов при синтезе полиэтилена. Отходы полимеров, несмотря на это, отчасти уничтожают сжиганием, улавливая образующиеся летучие вредные вещества. Однако это сильно удорожает их уничтожение. Именно это обстоятельство приводит к удорожанию и самих полимеров, в стоимость которых включают и затраты на их уничтожение.
  2. Вторичная переработка отслуживших свой срок полимеров и изделий из них. Однако, мы имеем дело, как правило, с грязными отходами, которые включают, например, частицы песка. Это исключает возможность применения высокопроизводительного и высокотехнологичного оборудования, используемого при первичной переработке исходных полимеров. Это оборудование будет быстро выходить из строя из-за абразивного воздействия твердых частиц минерального происхождения. Но даже при переработке, получаются «грязные» изделия, товарный вид и потребительские свойства которых не могут конкурировать с первичными изделиями. При этом имеется возможность использовать продукты вторичной переработки по другому назначению, предполагающему существенно пониженные требования. В частности, загрязненные изделия из полиэтилена могут быть переработаны в пластины толщиной в несколько миллиметров для применения в качестве кровельного материала, имеющего ряд неоспоримых преимуществ перед традиционными, таких, как низкая плотность, а значит, малый вес, гибкость и коррозионная стойкость, а также низкая теплопроводность и хорошие теплоизолирующие свойства.

В настоящее время количество полимерных отходов, которые реально утилизируются, т.е. идут на вторичную переработку или сжигаются, составляет менее 7 %.

  1. Использование биоразрушающей способности микроорганизмов для очистки окружающей среды от антропогенных загрязнителей.

К синтетическим полимерам, склонным к биоразложению, относятся сложные полиэфиры.

Число звеньев в цепи полимеров n, называемое степенью полимеризации, составляет приблизительно 100. Если эти полимеры закопать в землю на четыре недели, а затем извлечь и взвесить, то окажется, что их вес уменьшается в среднем на 20%. Это характерно для катализируемого ферментами гидролиза, продуктами которого являются низкомолекулярные вещества, растворимые в воде и диффундирующие в окружающее образец пространство. Этот процесс протекает на поверхности твердого образца, поскольку ферменты, будучи белками, то есть природными полимерами, не способны проникать в объем образца полиэфира, который приготавливают в форме пленки. Источником таких ферментов служат грибки, обитающие в почвах. Одновременно с этим в объеме образца протекает и химический гидролиз, то есть разрушение сложноэфирных связей полимерных цепей молекулами воды, способными проникать внутрь образца полимера. Этот процесс сопровождается разрывом цепей в любом месте с равной вероятностью. В результате наблюдается уменьшение молекулярной массы полимера и ухудшение механических свойств образца.

В том случае, если в полимерной цепи нет «слабых связей», способных расщепляться ферментами или подвергаться химическому разрушению в природной среде, дело обстоит гораздо сложнее. Типичный пример такого рода — полиэтилен, цепи которого образованы атомами углерода.

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В том случае, если в полимерной цепи нет «слабых связей», способных расщепляться ферментами или подвергаться химическому разрушению в природной среде, дело обстоит гораздо сложнее. Типичный пример такого рода — полиэтилен, цепи которого образованы атомами углерода.

Научный задел

A wonderful serenity has taken possession of my entire soul, like these sweet mornings of spring which I enjoy with my whole heart. I am alone, and feel the charm of existence in this spot, which was created for the bliss of souls like mine. I am so happy, my dear friend, so absorbed in the exquisite.

For the happiest life, days should be rigorously planned, nights left open to chance.

В настоящее время в ОАО «Биохиммаш» выделены из окружающей среды микроорганизмы способные использовать углерод полиэтилена в своём метаболизме в качестве источника углерода. В результате проведенных исследований было установлено, что полиэтилен может подвергаться деструкции под воздействием микроорганизмов. Среди этих микроорганизмов были обнаружены один вид грибов и пять доминирующих видов бактерий, образующих консорциум.

Условия культивирования

Образцы полиэтилена измельчали до частиц размером не более 5 мм. К навескам измельченного полиэтилена добавляли стерильную синтетическую питательную среду следующего состава:

K2HPO4 — 7,0 г;

KH2PO4 — 0,3 г;

Mg SO4´7H2O – 0,1 г;

(NH4)2SO4 – 1,0 г;

Na – цитрат ´3H2O — 0,5 г;

Микроэлементы: MgO – 0,1 г, FeCl3 – 0,054 г; ZnSO4´7H2O – 0,014 г; MnSO4´4H2O – 0,011 г; CuSO4´5H2O – 0,0025 г; CoSO4´7H2O – 0,0028 г; H3BO3 – 0,0006 г; Na2MoO4´2H2O – 0,0049 г.

Дистиллированная вода —  до 1 л.

В качестве единственного источника углерода использовали измельченный полиэтилен.

В приготовленную питательную среду вносили 1 мл посевного материала, содержащего консорциум микроорганизмов (5 штаммов).

Процесс биодеструкции полиэтилена проходит в анаэробных условиях; на начальной стадии с выделением газа, в дальнейшем — с потреблением выделенного газа.

Микроскопирование фрагментов полиэтиленовой пленки показало, что деструкция полиэтилена начинается в местах механического повреждения пленки с образованием перфорированных участков, затем нитеобразной структуры, и сферических образований, вероятно, содержащих продукты разрушения полиэтилена (Рис.2).

Рис.2. Разрушение полиэтиленовой пленки под воздействием консорциума микроорганизмов (увеличение 400 крат).

По окончании процесса культивирования через 12 месяцев был получен осадок активного ила с незначительными включениями фрагментов неразрушенной полиэтиленовой пленки и культуральная жидкость, цвет которой менялся от светло-желтого до темно-коричневого, опалесцирующего, в зависимости от типов полиэтилена. Структура осадка представлена на рис. 3.

Рис.3. Осадок продуктов биоконверсии полиэтилена (увеличение 400 крат).

Были проведены анализы зрелой культуральной жидкости.

Результаты газохроматографических анализов газовой фазы над жидкими пробами представлены в таблице 1. Концентрации веществ приведены в мг/м3 .

Таблица 1.

пп

Обнаруженные

вещества

1

проба

2

проба

3

проба

4

проба

5

проба

6

проба

7

проба

1 Метан (СН4) 3,11 2,47 2,75 2,68 3,28 2,01 2,76
2 Этан (С2Н6) 0,08 н.о. н.о. 0,05 0,13 0,13 0,09
3 Пропилен (С3Н6) 0,41 0,35 0,31 0,29 0,23 0,27 0,24
4 Пропан (С3Н8) 0,16 н.о. 0,15 н.о. 0,09 0,14 н.о.
5 Гептан (С7Н16) 0,13 н.о. 0,06 0.05 н.о. н.о. н.о.
6 Ацетон (С3Н6О/

СН3СОСН3)

0,15 0,33 0,13 0,28 0,12 0,16 0,10
7 Метанол (СН3ОН) н.о. 0,31 0,24 н.о. н.о. н.о. н.о.
8 Ацетальдегид (СН3СНО) н.о. 6,12 н.о. н.о. н.о. 2,37 н.о.
9 Бутилацетат (С6Н12О2/

СН3(СН2)3ОСОСН3

0,09 1,85 н.о. 0,78 н.о. н.о. н.о.
10 2-метил-1-бутанол

5Н12О/СН3СН2СН(СН3)СН2ОН)

н.о. 2,37 н.о. н.о. 1,26 н.о. н.о.
11 Сумма органических кислот 0,48 4,66 2,21 1,01 2,29 1,55 1,25

Таблица 2.

Состав эфирного экстракта
мкг/мл мкг/мл мкг/мл
Target Compounds MB-CA1 MB-CA4 MB-CA7
1 Lactic 0.7 0.0 0.0
2 Caproic 0.0 0.2 0.0
3 Glycerol 0.5 0.3 0.0
4 C13CD3st 3.0 3.0 3.0
5 Myristic 1.9 2.3 4.4
6 palmitic 14.8 10.6 20.1
7 linolic 2.9 0.0 0.0
8 oleic 0.3 2.2 4.2
9 stearic 15.1 9.5 19.1
10 фенол 0.0 0.9 0.2
11 2-hydroxy-propionic 0.0 4.7 0.0
12 фосфорная к-та 0.0 47.7 1.1
13 этилгексановая 0.0 0.0 11.3
Сумма 39 81 63

Результаты анализов, приведенные в таблицах 1 и 2, позволяют утверждать, что в процессе анаэробного культивирования консорциума микроорганизмов с использованием полиэтилена в качестве единственного источника углерода происходит утилизация полиэтилена с выделением смеси газообразных углеводородов, а также с образованием органических кислот.  Состав микрофлоры, качественное и количественное содержание продуктов метаболизма изменяется с увеличением продолжительности культивирования микроорганизмов. Так, в течение первого месяца культивирования происходит значительное выделение газа при неизменном составе микрофлоры культуральной жидкости и практически неизменном весе полиэтиленовой массы. После трех месяцев культивирования потери в массе полиэтилена составляют до 32,5% в зависимости от исходной концентрации полиэтилена. К 3-му месяцу культивирования потеря в массе полиэтилена замедляется. При длительном культивировании (более 11 месяцев) в культуральной жидкости образуется осадок активного ила, содержащий незначительное количество мелких фрагментов полиэтиленовой крошки.

Из полученных данных следует, что при культивировании с использованием полиэтилена в качестве единственного источника углерода, имеет место конкурентное ингибирование. Для оптимизации процесса в дальнейших исследованиях, на наш взгляд, культивирование следует вести в непрерывном режиме, т.е. с подпитками свежей питательной средой и отводом продуктов метаболизма.

Научная новизна

Нет сведений об аналогичных отечественных и зарубежных разработках.

Потребность в оборудовании

Экспериментальный стенд для исследования биохимических процессов, протекающих при биодеструкции полиэтилена, состоящего из: блока атомно-абсорбционного спектрофотометрирования и блока ультрафильтрационной подготовки образцов.

Ожидаемые результаты

  1. Выявление механизма микробиологической утилизации бытовых полимерных отходов. 2. Разработка технологии микробиологической утилизации бытовых полимерных отходов.
  2. Масштабирование и опытно-промышленные испытания технологии биодеструкции полиэтилена.

Бюджет и сроки

300.000 долл. США, три года

Следующий

Просмотрено 7 788

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