Mantar örneklerinde toplam antioksidan kapasite tayini
Determination of total antioxidant capacity of mushrooms
Yazar: HAFİZE REVA BAYRAKTAR
Danışman: PROF. DR. BİRSEN ÖZTÜRK
İstanbul Teknik Üniversitesi / Fen Bilimleri Enstitüsü / Kimya Ana Bilim Dalı / Kimya Bilim Dalı
Derece: Yüksek Lisans, 2019
Konu: Kimya = Chemistry

Özet:
Canlıların metabolik faaliyetleri esnasında oluşan kimyasal reaksiyonlar, özellikle
oksidasyon reaksiyonları, serbest radikallerin oluşmasına sebep olur.
Fazlasıylareaktif olma özelliğine sahip olan serbest radikaller birçok molekül ile
rahatça reaksiyona verebilir, hücreler bu reaksiyonlar yüzünden tahrip olabilir,
enzimlerin ve DNA'nın yapısı bozulabilir. Ayrıca gıdalar, kozmetik ürünler, ilaçlar,
sigara dumanı gibi faktörler serbest radikallerin oluşmasına sebebiyet verirler ve bu
serbest radikaller zincir reaksiyonunun başlangıcını oluştururlar. Antioksidanlar
serbest radikalleri inhibe ederek zincir reaksiyonunu engellerler. Gıdalar ile
organizmaya alınan antioksidanların en önemli etkisi "oksidatif stresi"
engellemesidir. Serbest radikallerin miktarının giderek yükselmesi ve hücre yapısına
zarar vermesi ile kendini gösteren oksidatif stresin kendisi direkt bir hastalık özelliği
taşımaz fakat çeşitli hastalıklara sebep olur.
Antioksidanların kimyasal çeşitliliği, sebze, meyve ve bitki matrikslerinden
ayrılmalarını ve miktarlarının tayinini zorlaştırmaktadır. Toplam antioksidan kapasite
(TAC), karmaşık bir örnekteki tüm antioksidanların antioksidan kapasitelerinin
toplamsallığını içeren bir parametredir. TAC (reaksiyon termodinamiğine veya
dönüşüm verimliliğine dayalı) veya aktivitenin (reaksiyon kinetiğine dayalı)
ölçülebilmesi için çeşitli analitik yöntemler geliştirilmiştir. Mantarlar, bitki grubunda
yer almakla birlikte yaprak gibi organlara sahip değildirler ve besinlerini ortamda
bulunan organik maddelerden sağlarlar (fotosentezle değil). Bu nedenle mantarlar
önemli bir gıda kaynağı olmalarının yanında, tıbbi amaçlarla da yetiştirilmektedir.
Bu çalışma da Türkiye'nin Antalya şehrinde yetişen doğal kestane mantarının
(Agaricus Spp.) saf su ve % 80 metanol-su kullanılarak dört farklı ektraksiyon
yöntemi (ultrasonik ekstraksiyon, mikrodalga ekstraksiyon, ultrasonik-mikrodalga
ekstraksiyon ve maserasyon) ile elde edilen ekstraktların antioksidan özellikleri
incelenmiştir.

Abstract:
In living beings, chemical reactions formed during metabolic activities, particulary
oxidation reactions, results in formation of free radicals, which are very reactive and
can easily react with different molecules, cells can be damaged due to that reactions
and structure of enzymes can be destroyed, moreover even chemical composition of
DNA can be changed. On the other hand, foods, drugs, cosmetics, smoke give rise to
occuring of free radicals. Chain reactions are initiated by means of free radicals
which are formed through oxidation reactions. These chain reactions are inhibited in
presence of antioxidants compounds, which have the ability of binding free radicals.
Antioxidants are beneficial compounds to health that fight reactive oxygen and
nitrogen species and free radicals that may eventually give rise to various diseases.
Plant foods, fruit and vegetable are rich in antioxidant vitamins, phenolic and
hydroxycinnamic acids, flavonoids, carotenoids, and anthocyanins. Consumption of
vegetables and fruits in the diet is one of the most efficient ways of preventing DNA
and protein damage, lipid peroxidation, cancer, atherosclerosis, ageing, and
inflammatory diseases.The most significant property of antioxidants, that are
ingested through nutrians, is equilibration of ''oxidative stress''. The oxidative stress
arises with escalating quantity of free radicals, which devastate structure of cells and
that oxidative stres is not a disease only it‟s self it also paves the way to other healt
disorders.
Total antioxidant capacity is a parameter including the cumulative action of all
antioxidants in a complex sample. Several analytical methods have been developed
to measure total antioxidant capacity (based on reaction thermodynamics or
conversion efficiency) and total antioxidant activity (based on reaction kinetics).
There are twenty analitical methods developed to determine antioxidant capacity.
Litrature investigations reveal that the antioxidant capacity of an antioxidant
compound determined with a certain method, may be different compared to another
method. Therefore it is necessary to use more than one antioxidant determination
method and compare them to analyze of antioxidant capacity of a nutrient. A
classification of total antioxidant capacity methods according to mechanism is the
type of reaction: Electron transfer (ET)- and hydrogen atom transfer (HAT)- based
assays. ET-based assays include ABTS/TEAC (trolox-equivalent antioxidant
capacity), DPPH (2,2- diphenyl-1-picrylhydrazeyl) assay, FCR (Folin-Ciocalteu
reagent) assay, FRAP (ferric ion reducing antioxidant power), CERAC (cerium (IV)
ions reducing antioxidant capacity) and CUPRAC (Cupric Reducing Antioxidant
Capacity) methods. Generally, ET-based methods are commonly used than HATbased ones, because they are cheaper and require less laboratory equipment, more
flexible, and thus more suitable to routine testing of natural products. However, ET
based assays may be less reproducible due to strong dependency on the protocol,
such as reagent concentrations, pH, solvent, and time of incubation. The problem can be likely solved by standardization of protocols. In this study, total antioxidant
capacity is tried to be determined with five different methods. These methods are
CERAC method, CUPRAC method, DPPH method, ABTS/TEAC method and lastly
Modifiye Folin- Ciocalteu method, These methods are cheap, more flexible, less
laborious and widely used for routine testing of natural products.
The chemical variety of antioxidants makes it hard to separate and quantify
antioxidants from the vegetable, fruit and plant matrix . Extraction is the first part of
separation of natural antioxidant compound from the plant materials. Most common
extraction methods are percolation and reflux extraction, solvent extraction,
distillation method, pressing and sublimation, usually organic solvents and large
volume of solvents are used for these methods, moreover a long extraction time is
consumed. Solvent extraction is the most widely used method. Furthermore there are
some contemporary and environment sensitive extraction methods such as super
critical fluid extraction, pressurized liquid extraction, ultrasound wave assisted
extraction and microwave assisted extraction, they are applied in natural products
extraction, and they have some advantages such as less organic solvent consumption,
shorter extraction time and higher selectivity. The extraction process of vegetable,
fruit and plant have got four steps, firstly the solvent penetrates into the solid matrix,
secondly the solute dissolves in the solvent, after that the solute is diffused out of
the solid matrix and lastly the extracted solutes are collected.
Factors affecting the extraction efficiency are the properties of the extraction solvent,
the particle size of the raw materials, the solvent-to-solid ratio, the extraction
temperature and the extraction time. The selection of the solvent is crucial for solvent
extraction. Selectivity, solubility, cost and safety should be considered in selection of
solvents. Based on the law of similarity which states that polar compounds are likely
to dissolve in polar solvents and apolar compounds are likely to dissolve in apolar
solvents as well. Alcohols such as ethanol and methanol are universal solvents in
solvent extraction for phytochemical investigation.
Generally, the smaller the particle size is, the more efficiency the extraction is. The
extraction efficiency will be enhanced by the small particle size due to the enhanced
penetration of solvents and diffusion of solutes.
High temperatures increase the solubility and diffusion, but too much high
temperature levels lead to solvents to be lost and lead to extraction of undesirable
impurities, additionally the decomposition of thermo sensitive components.
The extraction efficiency increases with the increase in extraction duration in a
certain time range. Increasing time will not affect the extraction after the equilibrium
of the solute is reached inside and outside the solid material.
While mushrooms are included in plant group they don‟t have organs such as leaf
and they obtain their foods by organic materials from the environment (not by
photosynthesis). For this reason muchrooms are significant nutrient source and also
grown for medical aims as well.
Ultrasonic extraction, also called ultrasonic extraction or sonication, uses ultrasonic
wave energy in the extraction. Ultrasound in the solvent producing cavitation
accelerates the dissolution and diffusion of the solute as well as the heat transfer,
which improves the extraction efficiency. The other advantage of ultrasonic
extraction is less solvent and energy consumption, and the reduction of extraction temperature and time. It is applicable for the extraction of thermolabile and unstable
compounds and commonly used in the extraction of many types of natural products.
Microwave extraction generate heat by interacting with polar compounds such as
water and some organic components in the plant matrix according to the ionic
conduction and dipole rotation mechanisms. The transfers of heat and mass are in the
same direction in microwave extraction, which accelerates and improve extraction
yield. The application of microwave extraction provides many advantages, such as
increasing the extract yield, decreasing the thermal degradation and selective heating
of vegetal material. It is also regraded as a green chemistry because it lowers the
usage of organic solvent.
This is a very simple extraction method with the disadvantage of long extraction time
and low extraction efficiency. It could be used for the extraction of thermolabile
components.
In this study antioxidant features of extractions, which are obtained by four different
extraction methods (ultrasonic extraction, microwave extraction, ultrasonicmicrowave extraction and maceration) of natural chestnut mushrooms(Agaricus
Spp.), which is grown in Antalya city of Turkey, are studied by using distilled water
and % 80 methanol-water.