Ph.D IN AGRICULTURAL AND FOOD SCIENCES XXXI CYCLE NUTRIENT DELIVERY AND WATER MANAGEMENT FOR PRODUCING LETTUCE (Lactuca sativa L.) FOR BIOREGENERATIVE LIFE SUPPORT SYSTEMS COORDINATER CANDIDATE Prof. Guido D’Urso Christophe El-Nakhel PROMOTER Prof. Youssel Rouphael Co-PROMOTER Prof. Stefania De Pascale 1 Table of content 1 Chapter 1- Introduction .......................................................................................... 6 1.1 BLSS history .............................................................................................................. 6 1.2 Plant role in BLSS ..................................................................................................... 8 1.3 Candidate crops for BLSS ......................................................................................... 9 1.4 Influence of lettuce genetic material ........................................................................ 11 1.5 Nutrient solution management: Macro-elements concentration & composition ..... 12 1.6 Importance of biofortification .................................................................................. 13 1.6.1 Iron biofortification ................................................................................................. 14 1.6.2 Selenium biofortification ......................................................................................... 14 1.7 Research objective ................................................................................................... 15 1.8 References ................................................................................................................ 17 2 Chapter 2: Performance and bioactive profile of diverse lettuce genotypes grown under optimal and low light conditions: attempt for energy limitation in future Bioregenerative Life Support Systems (BLSSs) ...................................... 19 2.1 Abstract …………………………………………………………………………...19 2.2 Introduction .............................................................................................................. 20 2.3 Materials and methods ............................................................................................. 23 2.3.1 Standards and chemicals .......................................................................................... 23 2.3.2 Growth chamber environmental control, plant material and closed soilless system management ………………………………………………………………………23 2.3.3 Sampling, morphological and physiological parameters ......................................... 24 2.3.4 Mineral composition analysis .................................................................................. 25 2.3.5 Antioxidant compounds determination .................................................................... 25 2.3.6 Extraction and quantification of total ascorbic acid and lipophilic antioxidant activity analysis ....................................................................................................... 26 2.3.7 Phenolic acids and anthocyanins identification and quantification ......................... 26 2.3.8 Target carotenoids extraction and quantification .................................................... 27 2.3.9 Statistical analysis and principal component analysis ............................................. 28 2.4 Results and discussion ............................................................................................. 29 2.5 Conclusion ............................................................................................................... 45 2.6 References ................................................................................................................ 46 3 Chapter 3: Water requirements, physiological and qualitative aspects of red and green butterhead lettuce grown in closed soilless system: better understanding of Salanova as a candidate cultivar for life support systems ... 51 3.1 Abstract …………………………………………………………………………...51 3.2 Introduction .............................................................................................................. 52 3.3 Materials and methods ............................................................................................. 53 3.3.1 Plant material, growth chamber conditions, experimental design and harvesting schedule …………………………………………………………………………..53 3.3.2 Sampling, growth analysis, and SPAD index measurement .................................... 54 3.3.3 Water uptake, water use efficiency and relative growth rate ................................... 55 2 3.3.4 Water potential and relative water content .............................................................. 55 3.3.5 Dry matter, total nitrogen and mineral content analysis .......................................... 56 3.3.6 Analysis of lipophilic antioxidant activity ............................................................... 56 3.3.7 Analysis of total ascorbic acid and total phenols ..................................................... 56 3.3.8 Statistical analysis .................................................................................................... 57 3.4 Results …………………………………………………………………………….57 3.4.1 Growth response, water uptake and water use efficiency ........................................ 57 3.4.2 Leaf macro-mineral composition ............................................................................. 64 3.4.3 Relative water content, leaf water potential and SPAD index ................................. 66 3.4.4 Qualitative parameters ............................................................................................. 68 3.4.5 Heat map analysis .................................................................................................... 70 3.4.6 Principal component analysis .................................................................................. 72 3.5 Discussion ................................................................................................................ 73 3.6 Conclusion ............................................................................................................... 77 3.7 References ................................................................................................................ 78 4 Chapter 4: Macronutrient deprivation eustress elicits differential secondary metabolites in red and green-pigmented butterhead lettuce grown in closed soilless system ......................................................................................................... 82 4.1 Abstract …………………………………………………………………………...82 4.2 Introduction .............................................................................................................. 82 4.3 Materials and methods ............................................................................................. 85 4.3.1 Plant Material, Growing Conditions, Experimental Design and Nutrient Solution Management ………………………………………………………………………85 4.3.2 Sampling, growth analysis, and light use efficiency ............................................... 86 4.3.3 Leaf dry matter and mineral content analysis .......................................................... 86 4.3.4 Analysis of hydrophilic antioxidant activity ............................................................ 87 4.3.5 Analysis of total ascorbic cid ................................................................................... 87 4.3.6 Phenolic acids and anthocyanins extraction, identification and quantification ....... 87 4.3.7 Carotenoids extraction, identification and quantification ........................................ 88 4.3.8 Statistics and principal component analysis ............................................................ 89 4.4 Results and discussion ............................................................................................. 90 4.4.1 Growth, fresh yield, leaf dry matter and light use efficiency .................................. 90 4.4.2 Mineral Profile and Nitrate Content ........................................................................ 93 4.4.3 Hydrophilic antioxidant activity, ascorbic acid and phenolic compounds .............. 98 4.5 Principal component analysis ................................................................................ 105 4.6 Conclusion ............................................................................................................. 108 4.7 References .............................................................................................................. 108 5 Chapter 5: The bioactive profile of lettuce produced in a closed soilless system as configured by combinatorial effects of genotype and macrocation supply composition ........................................................................................................... 112 5.1 Abstract ………………………………………………………………………….112 5.2 Introduction ............................................................................................................ 112 5.3 Material and methods ............................................................................................ 114 5.3.1 Standards and chemicals ........................................................................................ 114 3 5.3.2 Growth chamber conditions, plant material, experimental design and nutrient solution management ............................................................................................. 115 5.3.3 Growth parameters, marketable yield, leaf colour measurements and collection of lettuce samples