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Impact of tropospheric Ozone on Food and Feed Quality of Brassica species (OFFQ)

Tropospheric ozone - CODA-CERVA


Ground-level ozone (O3) is an example of how a local problem has grown in scale as a result of global industrialisation and the interactions of emissions over large geographical areas. When photochemical smog, of which ground-level O3 is the main component, was first recognised in southern California after World War II, it was considered to be a local phenomenon unique to the Los Angeles basin (Swedish Environmental Protection Agency and the Authors, 2009). Currently, tropospheric O3, that is present in the atmosphere 0-10 km above the earth surface (Fig. 1), is the third most important greenhouse gas and simulations for the period 2015 through 2050 even predict increases in tropospheric O3 of up to 25% (Meehl et al, 2007).




Tropospheric Ozone - CODA-CERVA
Fig. 1: Variation in O3 concentration with altitude in the atmosphere (ref: Swedish Environmental Protection Agency and the authors, 2009)

Besides its role as a direct greenhouse gas, O3 represents an important risk for human health, materials and vegetation (Kucera and Fitz, 1995, Meleux et al., 2007, Mills et al., 2011). There is a strong demand from policy makers for the quantification of these O3 damages to be fed into cost-benefit analysis of emission control strategies (Holland et al, 2006). O3 has proven to decrease crop production and change its quality, with the former aspect being more extensively studied in the past than the latter (Ashmore, 2005). The focus on quantitative yield changes may however result in a misleading risk assessment and unreliable economic extrapolations (Shortle et al., 1988) especially in those cases where the quality of the harvested product is crucial for industrial processing and consumer's health.


Therefore, the objective of the OFFQ project, funded by the Belgian Federal Science Policy, was to develop a quantitative relationship between the level of O3 exposure and the impact on yield and quality of two economically important Brassica species: spring oilseed rape (Brassica napus L.) (Fig. 2) and broccoli (Brassica oleracea L. cv. Italica) (Fig. 3). World production of oilseed rape is growing rapidly, with estimates of 58.4 million tons in the 2010-2011 season (USDA, 2011) and the world production of cauliflowers and broccoli reached over 19 million tons in 2009 (FAOSTAT, 2011).


Tropospheric ozone - CODA-CERVA Tropospheric ozone - CODA-CERVA
Fig. 2: Oilseed rape at flowering
Fig. 3: Broccoli, ready for harvest

Tropospheric Ozone - CODA-CERVA


This research required experimental facilities were both crops could be subjected to elevated O3 concentrations during their entire growth, from sowing or planting until harvest, under circumstances as close as possible to reality such as ambient climatic conditions, native soil, application of common agricultural practices etc. This was achieved by the use of so-called "Open-Top Chambers" (OTCs), located at the Veterinary and Agrochemical Research Centre (CODA-CERVA) in Tervuren (Fig. 4, on the right).



Tropospheric Ozone - CODA-CERVAHow to determine the critical ozone exposure or dose to prevent crop losses?

As for human exposure and health related impact studies, there is a fundamental difference between air pollutant concentrations as measured by physico-chemical methods and the effective dose absorbed by the recipient depending on e.g. physiological activity and fitness, influence of other environmental conditions, time and duration of exposure etc. As humans inhale O3 through normal breathing, plants absorb this gas through small openings in their leaves, the stomata (Fig. 5, on the left).

The actual diffusion of gasses (including O3) through the stomata is expressed by the stomatal conductance and strongly controlled by the stomatal aperture (Fig. 6). For the quantification of O3 responses a relationship based on the O3 uptake into the leaves, is preferred on the grounds that it estimates yield losses and quality effects against the received dose of O3, rather than against simple exposure to ambient concentrations. However, unlike O3 concentrations, the actual O3 uptake is hard to measure in the field and therefore the stomatal O3 fluxes are calculated through mathematical models (Op de Beeck et al., 2010). The influence of plant phenology, irradiance, temperature, air humidity and soil moisture on stomatal O3 uptake is incorporated in these models through their interference with the stomatal aperture (Jarvis, 1976). Based on the data provided by the OFFQ project, the research group of Plant & Vegetation Ecology of the University Antwerp was able to develop such a model for spring oilseed rape and broccoli (Op de Beeck et al., 2010).


Tropospheric ozone - CODA-CERVA Tropospheric ozone - CODA-CERVA
Fig. 6: Gas exchange measurements on plant leaves

Subsequently, the seed and oil yield of oilseed rape and fresh marketable broccoli yield were related to both the concentration based AOT40 (Accumulated Ozone exposure over a Threshold of 40 ppb) and the modelled O3 uptake POD6 (Phytotoxic O3 Dose above a threshold of 6 nmol s-1 m-2 projected leaf area) accumulated from emergence or planting until harvest (Fig. 7).

Tropospheric ozone - CODA-CERVA


Fig. 7: Correlation between relative seed yield and AOT40 (a) or POD6 (b) for spring oilseed rape, calculated from 50% emergence until harvest. Relative seed yield is the ratio of the seed yield at a specific level of accumulated O3 exposure/dose relative to the reference value (100% yield) at an AOT40 or POD6 equal to zero (De Bock et al., 2011)



Based on these O3 dose relationships, the critical O3 exposure/dose above which 5% seed yield reduction for oilseed rape may be expected, was calculated to be 3.7 ppm h AOT40 and 4.4 mmol m-2 POD6 (De Bock et al., 2011). The critical levels for 5 % oil yield reduction were lower due to an additional decrease of the oil percentage (3.2 ppm h and 3.9 mmol m-2). These concentration- and flux-based critical levels can then be compared to modelled O3 concentrations and fluxes for 50 km x 50 km grid squares across Europe as supplied by EMEP (European Monitoring and Evaluation Programme) to identify those areas that are most at risk for O3 damage, not only at present but also in the future.

The OFFQ project revealed that economic losses can be expected for oil seed rape if O3 concentrations continue to rise. In comparison to the current situation, seed yield losses of spring oilseed rape may reach 30% within 100 years if future ambient 7 or 12 hr average O3 concentrations increase to a range of 51 - 75 ppb, as predicted by Assessment Report Four (Meehl et al, 2007). For broccoli, no adverse O3 effects on fresh vegetable yield quantity are to be expected (De Bock et al., 2011).


Consequences of ozone pollution for the food and feed quality


Crop quality may be affected by O3 pollution either by changes in primary metabolite production and/or assimilate allocation and transport (carbohydrates, proteins,...) but also by changes in secondary plant metabolism. Due to the strong oxidative character of O3, these effects are induced by an increased production of reactive oxygen species, both outside and inside the plant cell, which is a common feature of biotic (pathogens, insects) and edaphic stresses (drought, high light, UV, cold...) in plants. The increased oxidative stress activates signal transduction pathways that drive plant defence responses.


In Brassica species, the phytochemicals arising from such changes in plant defence responses include not only powerful antioxidants such as vitamin C (ascorbic acid, ASC) and E (tocopherol, TOC) (Iriti and Faoro, 2009), but also glucosinolates. These compounds are commonly termed "natural pesticides" because they are often toxic to predators and diseases, such as insects , animals, fungi and bacteria, thereby conferring a competitive advantage to the plant that produces them (Talalay and Fahey, 2001). In animal feed, glucosinolates decrease digestibility and may cause goitre and haemolytic anaemia if supplemented at excessive rates (Stoewsand, 1995). For the human diet on the other hand, glucosinolates are of particular importance because the protective effect of cruciferous vegetables (cauliflower, broccoli, Brussels sprouts...) against cancer has been suggested to be partly due to these compounds (van Poppel et al., 1999).

The results of the OFFQ project showed that increased tropospheric O3 had an influence on the qualitative attributes of the harvested products of oilseed rape and broccoli, which may have economic effects and consequences for the nutritional value (Vandermeiren et al., 2012). Rapeseed oil, derived from the seeds of oilseed rape, is the third most important world source of vegetable oil (Luhs and Friedt, 1994) whilst the residual seed meal is used as feed supplement because of its high protein content. Exposure to more elevated O3 concentrations caused a decrease of the seed oil percentage, but the protein concentration was increased. Consequently, to obtain a reliable estimation of the economic consequences of increasing tropospheric O3 levels such a qualitative shift in seed composition needs to be taken into account in combination with the predicted yield losses (De Bock et al., 2011).


Next to a reduction of the oil concentration, there was also a shift in the fatty acid composition of seeds of oilseed rape. Oleic acid, a monounsaturated fatty acid which constitutes about 60% of the total fatty acid content, declined significantly in favour of linoleic acid. There was no change in the relative proportion of linolenic acid. The suppression of unsaturated oleic acid is of nutritional importance since mammalian metabolism does not allow the build-up of these double bonds. Linoleic and linolenic acids are also essential polyunsaturated fatty acids and precursors of the Omega 6 and Omega 3 fatty acid families.

Also a decrease in vitamin E content of the seeds was observed. Such a decrease of vitamin E could be considered detrimental to the nutritional value of the oil. However, this effect was mainly due to a decrease of γ-TOC and not of α-TOC, which exhibits the greatest activity in the prevention of vitamin E deficiency abnormalities (San Andrés et al., 2011). γ-TOC mainly plays a role in relation to oil stability (Gliszczynska-Swiglo and Sikorska, 2004) since TOCs are powerful antioxidants reducing the autoxidation of unsaturated fatty acids, the production of off-flavours and rancidity (Raclaru et al., 2004). So, although α-TOC is healthwise most important in vegetable oils, γ-TOC is qualitatively equally or maybe even more relevant to preserve the oxidative stability and increase their storage life.


O3 exposure did not result in any significant changes in glucosinolate content or composition of Brassica napus seeds; so no consequences are to be expected with regard to feed safety. The spring oilseed rape cultivar that was used in our experiments is a so-called double low variety which implies that the glucosinolate level in the seeds is very low (less than 20 μmol g-1 seed dry matter). For ruminants, such low-glucosinolate rapeseed meal can be used as the sole protein supplement without any apparent adverse effects on animal health (EFSA, 2008).


ITropospheric ozone - CODA-CERVAn broccoli however, an important shift occurred from indolic to aliphatic glucosinolates although the total glucosinolate concentration was not changed. The increase in the aliphatic/indolic glucosinolate ratio (Fig. 8) may be important in relation to the anticarcinogenic properties of these vegetables. The prevailing mechanism for the anticarcinogenic activity of these compounds is considered to be the induction of mammalian detoxication and antioxidant (phase II) enzyme activity by their breakdown products: the isothiocyanates. Sulphoraphane, the isothiocyanate from the aliphatic glucosinolate glucoraphanin, is still considered the most potent inducer of phase II enzyme (Verkerk et al., 2009).


The isothiocyanate indole-3-carbinol, derived from indolic glucosinolates, on the other hand, is considered to both inhibit and promote carcinogenesis (Stoner et al., 2002) and appears responsible for the modulation of estrogen receptor activity (Rahman and Sarkar, 2002). Such findings complicate conclusions in relation to the health effects of glucosinolates for humans but overall the increase of the ratio of aliphatic/indolic glucosinolates in response to O3 may be considered beneficial.

Main conclusion

The OFFQ project provided clear evidence that not only yield effects, but also changes in food and feed quality of the harvested products are essential to determine the economic costs and risks of a further increase in tropospheric O3. Depending on the crops, the specific quality parameter and the nature of the response, O3 may have either a beneficial or detrimental impact on the quality and safety of the food and feed chain.



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