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Wattanakul, Jutarat (2020) Nutrient value and stability of chloroplast-rich fraction (CRF) material from pea vine haulm, and the role of galactolipolytic enzymes during digestion. PhD thesis, University of Nottingham.

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Abstract

Underutilised green plant material is a rich source of chloroplasts, organelles in plants and other photosynthetic organisms, which are responsible for the conversion of light energy into chemical energy. The removal of intact chloroplasts from their cell wall confinement offers a novel way to obtain lipophilic nutrients and galactolipids from green biomass, especially carotenoids, vitamin E, and galactolipids (the main membrane lipids in plants and they represent a major source of the essential alpha-linolenic acid (18:3, ALA)). We have developed a physical method of recovering a chloroplast-rich fraction (CRF) from postharvest, pea vine field residue (haulm) by using slow-screw twin-gear juicer without added water. The CRF from pea vine haulm (PVH), like CRF from spinach leaves, is a good source of lipophilic nutrients, including beta-carotene (provitamin A), lutein, alpha-tocopherol (vitamin E), phylloquinone (vitamin K), and essential fatty acids.

Steam sterilisation of pea vine haulm prior to generating CRF was used to inactivate enzymes after harvesting and extend the shelf life of nutrients. The stability of selected nutrients (beta-carotene, lutein, and alpha-tocopherol) in the freeze-dried CRF material was measured over 84 days; the impact of storage temperature (-20°C, 4°C, 25°C and 40°C), light and air on nutrient stability was established. All three nutrients were stable at -20°C and 4°C in the presence or absence of air; this stability was lost at higher temperatures in the presence of air. The extent and rate of nutrient breakdown significantly increased when the CRF samples were exposed to light. beta-Carotene appeared to be more susceptible to degradation than lutein and tocopherol at 40°C in the presence of air, but when CRF was exposed to light all three nutrients measured were significantly broken-down during storage at 25°C or 40°C, whether exposed to air or not.

The bioaccessibility (% nutrient available for uptake) of some essential lipophilic nutrients, contained in a chloroplast-rich fraction (CRF) recovered from post-harvest pea vine field residue (haulm) (PVH), was measured using an in vitro gastrointestinal human digestion model with and without additional rapeseed oil. The impact on nutrient stability during digestion (in-digesta) and bioaccessibility of postharvest heat treatment of the biomass (HPVH), or the juice (HJ) derived from the biomass, was determined. The results show that both heat treatments stabilised beta-carotene, lutein, alpha-tocopherol and alpha-linolenic acid in the CRF material in-digesta. The presence of oil during digestion appears to improve retention of beta-carotene, and the bioaccessibility of beta-carotene in CRF from HPVH and HJ, except in the case of lutein and alpha-tocopherol. Conversely, the combination of heat treatment both HPVH and HJ and the presence of oil in PVH CRF enhanced the retention and bioaccessibility of beta-carotene, lutein, and alpha-tocopherol.

The release of nutrients that are embedded into the chloroplast structure is likely to be affected by the extent to which thylakoid membranes are hydrolysed during digestion. The PVH CRF and spinach CRF were therefore measured in vitro under simulated gastrointestinal conditions. Using a two-step static model, CRF from both spinach leaves and postharvest pea vine field residue (haulm) was first exposed to enzymes from rabbit gastric extracts and then either to pancreatic enzymes from human pancreatic juice (HPJ) or to porcine pancreatic extracts (PPE). The lipolysis of monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) was monitored by thin layer chromatography and gas chromatography of fatty acid methyl esters. For both CRF preparations, MGDG and DGDG were converted to mono galactosylmonoacylglycerol (MGMG) and digalactosylmonoacylglycerol (DGMG), respectively, during the intestinal phase and ALA was the main fatty acid released. Galactolipids were more effectively hydrolysed by HPJ than by PPE, and PPE showed a higher activity on MGDG than on DGDG. These findings may be explained by the higher levels of galactolipase activity in HPJ compared to PPE, is probably due to presence of pancreatic lipase-related protein 2 (PLRP2) in HPJ. Thus, we showed that CRF galactolipids are well digested by pancreatic enzymes and represent an interesting vehicle for ALA supplementation in human diet.

Item Type:	Thesis (University of Nottingham only) (PhD)
Supervisors:	Gray, David Foster, Tim
Keywords:	chloroplast, pea vine haulm, beta-carotene, lutein, alpha-tocopherol, linoleic acid, alpha-linolenic acid, galactolipid, digestion
Subjects:	S Agriculture > SB Plant culture T Technology > TX Home economics
Faculties/Schools:	UK Campuses > Faculty of Science > School of Biosciences
Item ID:	63487
Depositing User:	Wattanakul, Jutarat
Date Deposited:	04 Oct 2023 12:08
Last Modified:	04 Oct 2023 12:08
URI:	https://eprints.nottingham.ac.uk/id/eprint/63487

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