Abstract
The present research project has been designed to explore microwave-enzyme-assisted methods for extraction of conventional, non-conventional and high-value vegetable oils from indigenous oilseed crops of Pakistan without the use of volatile organic solvents (VOS). In this context, enzyme-assisted cold pressing (EACP) and enzyme-assisted aqueous extraction (EAAE) processes are in use. The optimization of enzymatic process variables such as pH, temperature, hydrolysis time, seed/water ratio, enzyme/substrate ratio etc. is under investigation to accomplish an inclusive protocol for improving the yield/recovery and nutritive quality of produced oils and the oilseed residues. Furthermore, efforts are underway to evaluate the effects of short-term microwave heating pretreatment of seeds, prior to enzymatic applications, so that a fast and an effective extraction procedure can be devised. The analyses of quality-oriented physico-chemical characteristics and antioxidant attributes of the oils, produced with enzyme-adjuvant, is designed employing classical (routine wet chemical assays) and non-classical (modern analytical equipments such as UV/VIS, FTIR and HR-GLC, HPLC and Rancimat) techniques. The development of the proposed microwave-enzyme-assisted oil extraction methods can lead to improving the oil quality, nevertheless, it might offer valuable by-products for food and feed industry.
Problem Statement
The flammability, explosiveness, accelerated temperature and environmental concerns of conventionally used hexane-oilseed extraction methods have urged the edible oil industry to seek some suitable alternative techniques for oilseed extractions. In this regard enzyme-assisted extraction technique is emerging as an alternative and eco-friendly approach to extract oils from variety of seeds. Efficacy of such methods may also be further enhanced by microwave pretreatment of seeds. Such microwave and enzyme adjuvant help break up the structure of seed cell wall cotyledon and the complex lipoprotein and lipopolysaccharide molecules into simpler molecules offering higher oil yields. In the present project multi-enzymes with diverse activity are in use for improvement of oil recovery and nutritive quality of oils from selected oilseed crops of Pakistan using EAAE and EACP.
Objectives

1. Exploration of eco-friendly microwave-enzyme-assisted methods for extraction of conventional, non-conventional and high-value vegetable oils from indigenous oilseed crops of Pakistan.
2. Appraisal of the efficacy of different enzyme mixtures for optimum oil recovery.
3. Investigatation of the potential of short-term microwave heating pretreatment of seeds, prior to enzymatic applications, for a fast and effective extraction system.
4. Optimization of process variables such as pH, temperature, water/seed ratio, enzyme concentration and incubation time to develop a comprehensive protocol for improved oil recovery and quality.
5. Detailed characterization of enzyme-extracted vegetable oils for physico-chemical, sensory and antioxidant attributes

Methodology
i)          Collection of Sample
Indigenously available raw materials/oilseeds: Sohanjna seeds, Safflower seeds, Low-erucic acid rapeseed, Maize seeds, Cottonseed, Rice bran, Soybean, Blackcurrant seeds and flaxseed etc. will be collected/procured from local agricultural resources. These will be then subjected to preliminary pretreatment (drying, dehulling, flaking, crushing and sieving) etc.
ii)         Extraction of Oil
The seeds materials will be treated with different enzymes mixtures and then the oil will be extracted by aqueous extraction and cold pressing. Experimental variables such as pH, temperature, solid/liquid ratio, enzyme/substrate ratio, and incubation time etc. for the enzyme-assisted extraction process will be optimized for development of comprehensive protocol leading to achieving optimum oil yield with better oil quality. Furthermore, the effects of short-term microwave heating pretreatment of seeds, prior to enzymatic applications, will also be studied.   
iii)        Determination of Physico-chemical Characteristics of Oils                  
The oils, extracted with enzyme-adjuvant and conventional methods, and the oilseed residues/cakes, left after oil extraction, will be characterized for physical, chemical and nutritional properties, using classical and non-classical analytical methods. Routine quality-oriented parameters such as density, refractive index, iodine values, free fatty acids, oxidation parameters, saponification value and unsaponifiable matter etc. of the extracted oils will be investigated by standard AOCS/IUPAC methods. Spectroscopic (UV/VIS, FTIR), chromatographic (HR-GLC, HPLC), tintometer and Rancimate techniques will  be used for the evaluation of oxidative stability parameters, fatty acids, tocopherols, sterols, and triglyceride profiles of the oils under investigation. 
iv)        Determination of Antioxidant Attributes of Oils
The antioxidant attributes of the oils, produced with enzyme treatments, will be evaluated using different in-vitro antioxidant assays such as measurements of total phenolic contents (TPC), DPPH radical scavenging capacity, percent inhibition of linoleic acid oxidation etc.
c)         Results & Discussion
NA  (The project is in the process of initial execution).
d)        Conclusions
N/A    
e)         Recommendations
N/A     
f)          One to two photographs of project     N/A