An overview of potential cardioprotective benefits of xanthophylls in atherosclerosis:an evidence-based review

Atherosclerosis,as the most prevalent form of cardiovascular disease,is characterized by oxidized lowdensity lipoprotein(ox-LDL)accumulation in the vascular wall,increased inflammation of the large arteries,dysfunction of the endothelial cells(ECs)and vascular smooth muscle cells(VSMCs),which may eventually lead to the formation of plaques.Xanthophylls,one of the main groups of carotenoids,have been proposed as preventive agents or adjunct therapies to prevent and slow the progression of atherosclerosis due to their cardioprotective properties.However,the underlying preventive mechanism of action of xanthophylls on the pathogenesis of atherosclerosis remains unclear,and clinical evidence of the effect of xanthophylls on atherosclerosis have not yet been summarized and critically reviewed.In this regard,we conducted a comprehensive literature search in four scientific databases(Pub Med,Google Scholar,Science Direct and Web of Science)and carefully analyzed the existing evidence to provide meaningful insights on the association between xanthophylls and atherosclerosis from various aspects.Based on the evidence from in vitro and in vivo studies,we explored several potential mechanisms,including antioxidant effect,anti-inflammatory effect,regulation of lipid metabolism,and modulation of ECs and VSMCs dysfunction,and we found that a clear picture of regulatory pathways of xanthophylls on atherosclerosis prevention and treatment is still lacking.In addition,epidemiological studies suggested the possible relationship among high dietary intake of xanthophylls,high plasma/serum xanthophylls and a reduced risk of atherosclerosis.Direct evidence from interventional studies investigating the effect of xanthophylls on atherosclerosis is very sparse,whilst indirect clinical evidence was only limited to astaxanthin and lutein.Therefore,well-designed long-term randomized controlled trials(RCTs)are highly recommended for future studies to investigate the effective dose of different xanthophylls on atherosclerosis prevention and their possible ancillary effect in conjunction with drug therapies on different stages of atherosclerosis.

Inflammation/bioenergetics-associated neurodegenerative pathologies and concomitant diseases:a role of mitochondria targeted catalase and xanthophylls

Various inflammatory stimuli are able to modify or even"re-program"the mitochondrial metabolism that results in generation of reactive oxygen species.In noncommunicable chronic diseases such as atherosclerosis and other cardiovascular pathologies,type 2 diabetes and metabolic syndrome,these modifications become systemic and are characterized by chronic inflammation and,in particular,"neuroinflammation"in the central nervous system.The processes associated with chronic inflammation are frequently grouped into"vicious circles"which are able to stimulate each other constantly amplifying the pathological events.These circles are evidently observed in Alzheimer's disease,atherosclerosis,type 2 diabetes,metabolic syndrome and,possibly,other associated pathologies.Furthermore,chronic inflammation in peripheral tissues is frequently concomitant to Alzheimer's disease.This is supposedly associated with some common genetic polymorphisms,for example,Apolipoprotein-Eε4 allele carriers with Alzheimer's disease can also develop atherosclerosis.Notably,in the transgenic mice expressing the recombinant mitochondria targeted catalase,that removes hydrogen peroxide from mitochondria,demonstrates the significant pathology amelioration and health improvements.In addition,the beneficial effects of some natural products from the xanthophyll family,astaxanthin and fucoxanthin,which are able to target the reactive oxygen species at cellular or mitochondrial membranes,have been demonstrated in both animal and human studies.We propose that the normalization of mitochondrial functions could play a key role in the treatment of neurodegenerative disorders and other noncommunicable diseases associated with chronic inflammation in ageing.Furthermore,some prospective drugs based on mitochondria targeted catalase or xanthophylls could be used as an effective treatment of these pathologies,especially at early stages of their development.

Potential ability of xanthophylls to prevent obesityassociated cancer

Obesity-associated cancers, including colon cancer and breast cancer, are increasing in Asian countries with Westernized lifestyles as exemplified by reduced physical activity and increased fat/sugar consumption. An excessive accumulation of visceral adipose tissue causes insulin resistance, dyslipidemia and adipocytokine imbalance, and these factors are suggested to be involved in cancer promotion. To prevent obesityassociated cancers, researcher attention is increasing on the so-called "functional foods". In addition, new approaches to cancer control are in high demand, and using "functional foods" as supplemental or adjuvant agents in chemotherapy is thought to be a promising approach. One of these functional ingredients is xanthophylls, which are natural fat-soluble pigments found in fruits, vegetables, algae and other plants. Xanthophylls belong to the carotenoid class and have struc-tures containing oxygen. Some studies have revealed that xanthophylls improve the inflammation status, serum triglyceride levels, blood pressure levels and liver function test values. Furthermore, recent studies show that xanthophylls possess high anti-cancer, antidiabetic, anti-obesity and anti-oxidant properties. In this review, we highlight the recent findings for five xanthophylls, namely astaxanthin, β-cryptoxanthin, fucoxanthin, neoxanthin and zeaxanthin/lutein, and their relevance to cancer prevention.

Optimization of xanthophylls extraction from Marigold extractum with supercritical CO_(2) by response surface methodology

In this work,xanthophylls were extracted from Marigold using supercritical CO_(2).A response surface experimental design was used to optimize operating conditions for extraction pressure(22-30 MPa),extraction temperature(50-60℃),CO_(2) flow rate(9-15 kg/h).The maximum concentration of xanthophyll in extracts was determined to be 35.8%by high performance liquid chromatography(HPLC),and optimum xanthophylls extraction was obtained at 55℃,26 MPa and 12 kg/h.The optimization results demonstrated that for xanthophylls extraction,all variables(temperature,pressure and CO_(2) flow rate)were to be the influential variables,with the statistical significant effect p-value was smaller than 0.05.

Responses of Chlorophyll Fluorescence and Carotenoids Biosynthesis to High Light Stress in Rice Seedling Leaves at Different Leaf Position

In the present study, we investigated the changes of photosynthesis, chlorophyll fluorescence and the content of carotenoid pigments in rice (Oryza sativa L.) seedling leaves and their responses to high light. The results showed that the rate of photosynthesis, the contents of individual and total carotenoids and the pool size of xanthophyll cycle decreased with age increasing of the leaf. When the leaves were exposed to high light for 2 h, the qN of mature leaf (5th leaf) increased more significantly than that of younger (6th leaf) and older leaves (3rd and 4th leaf). Comparing with the leaves before exposure to high light, the excitation pressure on PSⅡ (1- qP ) increased by 44%, 57%, 19% and 45% in the 3rd, 4th, 5th and 6th leaf under high light, respectively. The highest content of carotenoids and the greatest conversion of violaxanthin to zeaxanthin were found in the 5th leaf, and it was consistent with the 5th leaf exhibiting the strongest resistance to high light. Our results suggested that the ability of rice leaf to resist photoinhibition is related to the level of carotenoids and the ability of carotenoids biosynthesis.

Cloning and Expression Analysis of Violaxanthin De-Epoxidase (VDE) cDNA in Wheat

Violaxanthin de-epoxidase (VDE) is the key enzyme in the xanthophyll cycle and protects plant photosynthetic apparatus from the damage of excessive light. A wheat (Triticum aestivum L cv. Xiaoyan 54) VDE cDNA was obtained using RT-PCR method. Its deduced protein sequence shares high identity with that of Arabidopsis and rice. Southern blot revealed that there are three copies of VDE gene per haploid genome of wheat. VDE transcript levels were higher in green leaf than in root, seed and etiolated leaf. Northern blotting analysis indicated that VDE mRNA level is induced during greening process of etiolated wheat seedling and increased by intense light illumination.

Photoinhibition in Shaded Cotton Leaves After Exposing to High Light and the Time Course of Its Restoration

Chlorophyll fluorescence emission, pigment composition and photosynthetic rate of shade-grown cotton ( Gossypium hirsutum L.) plants were measured immediately after suddenly exposing to full sunlight and at regular intervals there after within 15 d. Photoinhibition occurred in shade-grown cotton leaves immediately after exposed to full sunlight. The chlorophyll fluorescence parameter F-v/F-m and PhiPS II, which reflect the efficiency of PS II,obviously decreased in shade-grown leaves, much lower than that of the full sunlight-grown leaves. On the contrary, F-o value was sharply increased. Neither of these parameters could completely recover till next morning. The photoinhibition was chronic and continued for about 4 d, while the F-v/F-m and the net photosynthetic rate ( P-n) continued to decline, then began to increase gradually 6 d later and turned stable after 10 - 12 d, appearing as an acclimation phenomenon. However, the final value of F-v/F-m and P-n did not reach the level as in those leaves grown in the full sunlight ever before. The final P-n was higher by 60% than that before exposure, but lower for more than 40% than that of the full sunlight-grown leaves. The most notable response of chloroplast pigment composition was a pronounced increase in the pool size of carotenoids in xanthophyll cycle over a period of 3 d. The results indicated that when shade-grown cotton seedlings were suddenly transferred to the full sunlight, the decline of F-v/F-m and P-n might associate with the damage of the PS II reaction center. During the light acclimation, photoprotective mechanisms such as the xanthophyll cycle-dependent energy dissipation were increased, so that photodamage in leaves transferred from low to high light might be reduced.

Xanthophyll Cycle and Its Molecular Mechanism in Photoprotection

When plants absorb more light than that can be used for photosynthesis, the excessive energy can cause photoinhibition and even photooxidation of photosynthetic apparatus. Xanthophyll cycle-dependent photo-protection is believed to be the main mechanism for plants to deal with excessive light energy. This review focuses on molecular biological aspects and regulations of violaxanthin de-epoxidase and zeaxanthin epoxidase involved in xanthophyll cycle. We will summarize the functions of xanthophyll cycle, especially recent advances in its thermal dissipation mechanism of photoprotection. Some interesting issues deserving further study will be discussed.

Photoinhibition and Photoprotection in Ginkgo biloba leaves: Influence of Temperature, CO 2 and O 2

In midday ginkgo ( Ginkgo biloba L.) leaves have to bear photon flux density over 1400 μmol·m -2 ·s -1 in combination with high temperatures around 35 ℃ at natural habitat. They show typical midday depression of stomatal conductance and of CO 2 assimilation rate. The zeaxanthin changes with light intensity during the day. The influence of the combination of strong light and temperature on photoinhibition was also examined in the laboratory. A low CO 2 internal conductance (31 mmol·m -2 ·s -1 ) was found in ginkgo leaves, which had been exposed to excessive light at temperature between 15 ℃ and 35 ℃ with reduced CO 2 (80 μL·L -1 ) or oxygen (2%) for 2 h, causing a low CO 2 concentration at the carboxylation site and a high proportion of photorespiration. The ratio of electron transport to CO 2 fixation was rather high in ginkgo (16 e -/CO 2 at 25 ℃) as compared with other plants. It increased with temperature also in 2% O 2 which could not be explained solely as due to change of photorespiration. The reduction of oxygen in 340 or 80 μL·L -1 CO 2 had no effect on the extent of photoinhibition at all temperatures, which indicated that electron flow caused by photorespiration in excess light was negligible in protective effect in ginkgo leaves. However, a decreased CO 2 concentration increased photoinhibition, especially at high temperature. It is concluded that the dissipation of excessive excitation energy in the PSⅡ antennae through the xanthophyll cycle may be the major protective mechanism to preventing from the deteriorated effects of strong light in ginkgo leaves.

Photochemical Efficiency of PSⅡ and Membrane Lipid Peroxidation in Leaves of indica and japonica Rice (Oryza sativa) Under Chilling Temperature and Strong Light Stress Conditions

Relationships between fluorescence parameters and membrane lipid peroxidation in leaves of indica and japonica rice (Oryza sativa L.) during later growth stage were studied under chilling temperature and strong light stress conditions. Results showed that D1 protein contents of PSⅡ in photosynthetic apparatus dropped, the generation of antheraxanthin (A) and zeaxanthin (Z) of xanthophyll cycle were inhibited partly, PSⅡ photochemical efficiency (F v/F m)and non-photochemical quenching (q N) were also decreased obviously. In addition, endogenous active oxygen scavenger—superoxide dismutase (SOD) reduced, superoxide anion radical (O -· 2) and malondialdehyde (MDA) accumulated, as a result, photooxidation of leaves occurred under chilling temperature and strong light stress conditions. Obvious differences in the changes of the above mentioned physiological parameters between indica and japonica rice were observed. Experiments in leaves treated with inhibitors under chilling temperature and strong light conditions showed that indica rice was more sensitive to chilling temperature with strong light and subjected to photooxidation more than japonica rice. Notable positive correlation between D1 protein contents and F v/F m or (A+Z)/(A+Z+V), and a marked negative correlation between F v/F m and MDA contents were obtained by regression analysis in indica and japonica rice during chilling temperature and strong light conditions. According to the facts mentioned above, it was inferred that PSⅡ photochemical efficiency(F v/F m) was the key index to forecast for the prediction of photooxidation under stress circumstances and the physiological basis were the synthetic capacity of D1 protein and the protection of xanthophyll cycle.

Photosynthesis of Resurrection Angiosperms

Resurrection plants which are able to quickly reactivate after falling into a period of anabiosis caused by dehydration have been very rare among angiosperms, especially among dicotyledons whose chlorophyll content and chloroplast structure little changed in the course of desiccation, therefore has been called homoiochlorophyllous desiccation-tolerant plants (HDTs). Another type of resurrection angiosperms that lost its chlorophyll dining desiccation is called poikilochlorophyllous desiccation-tolerant plants (PDTs). HDTs have been received more attention because of simplicity of protection mechanism which is much easy to the study and utilization of the desiccation tolerance of resurrection angiosperms. Recent advances in studies of photosynthesis of resurrection angiosperms indicate that photochemical activities are sensitive indicators for the study of physiological state of resurrection angiosperms during desiccation and rehydration. Photochemical activities of resurrection angiosperms are inhibited with loss of water similar to those of general plants, however, the magic thing is that they could reactivate rapidly during rehydration even losing more than 95% water. Up-regulations in xanthophyll cycle and antioxidative systems as well as preservation in integrity and stability of photosynthetic membranes during desiccation may be very important to desiccation tolerance of resurrection angiosperms. The fact that phosphate treatment in rehydration stage also strongly influences resurrection indicated importance of studies on rehydration stages of resurrection angiosperms.

Changes in Violaxanthin Deepoxidase Activity and Unsaturation of Thylakoid Membrane Lipids in Indica and Japonica Rice Under Chilling Condition and Strong Light

To explore the differences of sensitivities to chilling and strong light in indica and japonica rice (Oryza sativa L), the changes in unsaturation of thylakoid membrane lipids and xanthophyll cycle were studied under chilling condition and strong light. The contents of unsaturated fatty acids of thylakoid membrane lipids decreased and that of the saturated ones increased with the time of chilling and strong light treatment, resulting in the reduction of the index of unsaturation of fatty acids (IUFA). The activity of violaxanthin deepoxidase (VDE), a key enzyme of xanthophyll cycle, also reduced. The content of violaxanthin (V) increased, and the contents of antheraxanthin (A) and zeaxanthin M decreased, the ratio of (A+Z)/ (A+Z+V) decreased correspondingly. Arrhenius analysis showed that VDE was sensitive to both chilling and unsaturation level of thylakoid membrane lipids. Correlation analysis showed that there was distinctly positive relationships between IUFA of thylakoid membrane lipids and the activity of VDE, Fv/Fm, and D, protein content. Lower IUFA values, less fluidity and stability of thylakoid membrane lipids, lower VDE activity and (A+Z)/(A+Z+V) ratio were found in indica rice cv. Shanyou 63 than in japonica rice cv. 9516 under chilling and strong light.

Photoinhibition and Photooxidation in Leaves of indica and japonica Rice Under Different Temperatures and Light Intensities

Physiological indices related to the efficiency (F-v/F-m) of light energy conversion in PS II and the peroxidation of membrane lipid were measured in leaves of Oryza sativa L. sp. indica rice cv. 'Shanyou 63' and sp. japonica rice cv. '9516'' under different temperatures and fight intensities for 4 days. No changes in F-v/F-m and membrane lipid peroxidation product (MDA) were observed, so neither photoinhibition nor photooxidation happened in both rice cultivars under moderate temperature and medium light intensity. However, F-v/F-m dropped obviously with no change in MDA contents, and photoinhibition appeared in indica rice cv. 'Shanyou 63' under medium temperature and strong light intensity. Furthermore, both photoinhibition and photooxidation were observed in two rice cultivars under chilling temperature and strong light intensity. Experiments with inhibitors under chilling temperature and strong light intensity showed that indica rice had a decrease in DI protein content and SOD activity, and the extent of inhibition of xanthophyll. cycle and nonphotochemical quenching (qN) was larger, and a higher level of MDA was observed. The photoinhibition and photooxidation in indica rice were more distinct as compared with japonica rice. The authors suggested that PS II light energy conversion efficiency (F-v/F-m) and membrane lipid peroxidation were the key indices for the detection of photooxidation.

Overflow phenomenon in serum lutein after supplementation:a systematic review supported with SNPs analyses

Lutein,a type of carotenoids,is found to delay the onset and progression of age-related macular degeneration(AMD).Several lutein supplementation studies showed that after an initial increase,lutein serum levels demonstrated a subsequent decrease despite continuous supplementation.In this systematic literature review,this obscure phenomenon was tried to be explained.The subsequent drop in lutein levels was postulated due to down-regulation of lutein receptors scavenger receptor class B typeⅠ(SR-BI)in the gastrointestinal tract,upregulation of lutein degrading enzymeβ-carotene dioxygenase(BCDO2),or perhaps a combination of both.Some single nucleotides polymorphisms(SNPs)that could have influence on the occurrence of this phenomenon.To date,an exact scientific explanation for this phenomenon has not been established.Further research is needed to investigate this phenomenon in depth to reach an irrefutable explanation,giving that lutein is proven to be effective in delaying the onset and progression of AMD and its metabolism in the human body becomes of equal importance.

Dehydrolutein:a metabolically derived carotenoid never observed in raptors

Carotenoids are fat-soluble pigments synthesised by photosynthetic organisms(Brush,1990).Conversely,animals are incapable of synthesizing carotenoids de novo,and they must obtain them through their diet.However,some animal species are able to make some alterations to the basic chemical structure,converting ingested carotenoids into more oxidized and differently coloured forms(Schiedt,1998).

Photosynthetic Pigments Affected by Fertilizer Source in Four Brassica Vegetables

Effect of fertilizer source （mineral fertilizer or organic manure） on photosynthetic pigments leaves content of four brassica vegetables （Pak choi, Kohlrabi, Cauliflower and Cabbage） was tested. Among treatments, cauliflower fertilized with chicken manure had the highest content of chlorophyll a and b （10.08 and 9.37 μm·g^-1）, while, pak choi had the lowest values （2.00 and 2.44 μm·g^-1） in non-fertilized plants. These differences in chlorophyll a and b content may have a great impact on photosynthetic rate and activity. Total carotenoids and total xanthophylls （lutein, zeaxanthin, antheraxanthin and violaxanthin） were the highest in cauliflower while the lowest was in pakchoi, β-carotene, on the other hand, was the highest percentage of total carotenoids （65.7%） in pak choi while Kohlrabi had the lowest percent （57.4%）. This high percentage offl-carotene of total carotenoids may compensate for lower content of chlorophylls and assist for efficient light harvesting process. Lutein showed to be the major components of xanthophyll pigments brassica vegetables and ranged from 0.462μm·g^-1 in cabbage to 0.626μm·g^-1 in kohlrabi but not differed significantly which may refer to genetically controlled. However, lutein, antheraxanthin and zeaxanthin content were increased significantly by the application of chicken manure regardless ofbrassica vegetable type. Violaxanthin was only detected in pak choi and not affected by fertilizer source. It can be concluding from the increasing contents of total xanthophylls pigments that may increase photosynthetic efficiency through non-photochemical quenching process.

Comparative Study on Photoinhibition Between Two Wheat Genotypes

Changes in the efficiency of the primary light energy conversion, fluorescence quenching parameters and contents of photosynthetic pigments were compared between two wheat ( Triticum aestivum L.) genotypes in response to high light stress. The contents of chlorophyll and carotenoid in “Jing_411' were slightly higher than those in “Xiaoyan_54'. Under high light stress, photoinhibition as indicated by a sustained decrease in PSⅡ photochemical efficiency was more pronounced in “Jing_411' than in “Xiaoyan_54'. The content of ascorbate and the activity of the deepoxidase were higher in “Xiaoyan_54' than in “Jing_411'. The genotypic difference in resistance to photoinhibition is related to the capacity to dissipate the excess energy nonradiatively.

PSⅡ Photochemistry and Xanthophyll Cycle in Two Superhigh-yield Rice Hybrids, Liangyoupeijiu and Hua-an 3 During Photoinhibition and Subsequent Restoration

PSⅡ photochemistry and xanthophyll cycle during photoinhibition (exposed to strong light of 2 000 μmol photons·m -2 ·s -1 ) and the subsequent restoration were compared between two superhigh_yield rice hybrids (Liangyoupeijiu and Hua_an 3, the newly developed rice hybrids from two parental lines) and the traditional rice hybrid Shanyou 63 developed from three parental lines. The results showed that the maximal efficiency of PSⅡ photochemistry ( Fv/Fm ), the efficiency of excitation energy capture by open PSⅡ centers ( Fv′/Fm′ ), and the yield of PSⅡ electron transport ( Φ PSⅡ ) of the three rice hybrids decreased during photoinhibition. However, a greater decrease in Fv/Fm , Fv′/Fm′ , and Φ PSⅡ was observed in Shanyou 63 than in Liangyoupeijiu and Hua_an 3. At the same time, the components of xanthophyll cycle, antherxanthin (A) and zeathanxin (Z) increased rapidly while violaxanthin (V) decreased considerably. Both the rate of accumulation and the amount of A and Z in the two superhigh_yield rice hybrids were higher than that in Shanyou 63. The de_epoxidation state (DES) of xanthophyll cycle increased rapidly with the fast accumulation of A and Z, and reached the maximal level after first 30 min during photoinhibition. Of the three hybrids, the increasing rate of DES in Liangyoupeijiu and Hua_an 3 was higher than that in Shanyou 63. After photoinhibition treatment, the plant materials were transferred to a dim light (70 μmol photons·m -2 ·s -1 ) for restoration. During restoration, both chlorophyll fluorescence parameters and xanthophyll cycle relaxed gradually, but the rate and level of restoration in the two superhigh_yield rice hybrids were higher than those in Shanyou 63. Our results suggest that Liangyoupeijiu and Hua_an 3 had higher resistance to photoinhibition and higher capacity of non_radiative energy dissipation associated with xanthophyll cycle, as well as higher rate of restoration after photoinhibition, than Shanyou 63 when subjected to strong light.

Photosynthetic Characteristics of a Super High Yield Cultivar of Winter Wheat During Late Growth Period

The mechanism of high yield of winter wheat in the field at late growth period was investigated by measuring the photosynthetic characteristics of photosystem Ⅱ （PSⅡ） and xanthophylls cycle, which could provide physiological reference for breeding. Weimai 8 （W8）, a super high yield cultivar, and Lumai 14 （L14）, a control cultivar were object. The photosynthetic rate （Pn）, parameters of chlorophyll fluorescence and chlorophyll content were measured. The Pn, maximum photochemical efficiency of PSII （Fv/Fm）, quantum yield of PSII electron transport （ΦPSⅡ）, efficiency of excitation energy capture by open PSII reaction centers （Fv＇/Fm＇）, and photochemical quenching coefficient （qP） were higher in Weimai 8 compared to that in Lumai 14, a commercial high yield cultivar. Furthermore, Weirnai 8 showed a lower non- photochemical quenching coefficient and a lower de-epoxidized ratio of the xanthophyll cycle pigments than of Lumai 14 at late growth period. At mature stage, chlorophyll content of different leaves decreased both in Weimai 8 and Lumai 14. Chlorophyll content in flag, second and third leaf from the top of plant decreased more in Lumai 14 than in Weimai 8. These results suggested that Weimai 8 had more antenna pigments to absorb light energy, and had higher photosynthetic capability and photochemical efficiency of PSⅡ. The yield of Weimai 8 was also higher than that of Lumai 14.

Responses of Transgenic Tobacco Plants with Increased Proline Content to Drought and/or Heat Stress

Transgenic tobacco plants (M51-1) constitutively over-expressing a modified gene for the proline biosynthetic enzyme △2-pyrroline-5-carboxylate synthetase (P5CSF129A) and the corresponding wild-type plants (WT) were compared during drought or heat stress and under combination of both stresses. The proline content in M51-1 was several times higher than in WT plants. Under optimal conditions, the transpiration rate and stomatal conductance of M51-1 plants were lower than those in WT plants. The differences in net photosynthetic rate were not significant and water use efficiency and contents of chlorophyll and xanthophyll cycle pigments were higher in M51-1 than in WT plants. Drought induced by cessation of watering for 7 d resulted in decrease of all gas exchange parameters and chlorophyll content, but in an increase of the content of xanthophyll cycle pigments and degree of their de-epoxidation. After application of heat stress (40°C/60 min) to control or water-stressed plants the gas exchange parameters decreased considerably. Short-term heat stress alone, however, did not affect pigment contents. The responses of M51-1 and WT plants to the tested stresses did not differ significantly. Therefore, a decisive contribution of elevated proline content to drought or heat stress tolerance of tobacco was not proved.