Plant factories with artificial lighting (PFAL) are a relatively new method of efficient agricultural cultivation that are becoming more and more important in combatting increasingly serious global food supply problems [1
]. Unusual weather, water shortages, and reduction of cultivated land area all threaten to reduce the production of the crops worldwide; however, these environmental factors do not threaten PFAL crop growth as they are thermally insulated, artificially lit, air conditioned, and have air circulation fans along with supplies of CO2
and nutrient solution, making it so the environment can be fully controlled [2
]. Crops in PFALs always rely on artificial lighting in that the light energy drives photosynthesis, which means that electricity accounts for 25%–30% of the total production costs and that light sources account for the majority of all energy consumption, about 60%–80% [1
]. Therefore, improving the efficiency of light sources would greatly reduce the cost of PFALs, which would further encourage their sustainable development because costs and ecological impacts could be reduced.
In PFALs, light energy can be provided for crops at any time because day and night no longer determine lighting schedules. This means that any photoperiod can be chosen to optimize the growth and quality of many crops. Gaudreau [5
] reported that lengthening the photoperiod resulted in substantial gains in the fresh weight of lettuce. Longer photoperiods not only increased the growth of the lettuce under low CO2
concentrations at the same day light integral (DLI), but also compensated for weak photosynthetic photon flux density (PPFD) [6
]. One study revealed that increasing photoperiod under weak light cultivation conditions compensated for weak light stress [7
]. Vlahos et al. [8
] reported that the growth of Achimenes
species can be promoted under weak light intensities at the same DLI compared with those under high light intensities, as the lower light intensities combined with longer photoperiods resulted in higher light-use efficiencies. Overall, these results have suggested that under the same DLI, longer photoperiods promote the growth of lettuce.
Light is not only an energy source, but also an important environmental signal [9
]. Morphological adaptations and light-dependent adjustments optimize plants for photosynthesis in a given environment [11
]. One previous study has shown that the formation and accumulation of photosynthetic pigments in leaves was affected by light quality, which either increases light harvesting under low-light conditions or acts as screening pigments and free-radical scavengers under high-light conditions [13
]. There has been plenty of research about the effect of different ratios of red and blue light [11
], different light intensities [11
], and various light intensities combined with different photoperiods on plant growth [11
]. However, these studies have ignored the effect of light as an environmental signal affecting the circadian rhythm, which in turn affects the metabolism in many plants. Breathing in plants is regulated daily, alternately cycling between high activity and low activity in roughly 24 h, determined by the circadian rhythm, which also has a great influence on the growth of crops [17
]. Dodd [20
] reported that the circadian rhythm confers an advantage to plants, benefitting photosynthesis, growth, survival, and competitive advantage. Most plants use circadian oscillators to coordinate physiological and developmental processes such as photosynthesis, respiration, and cell wall synthesis. In addition, the rhythm of these processes can change periodically during a single day. Previous lighting strategies have ignored the effects of photoperiod on circadian rhythm, missing the opportunity to use the circadian rhythm to benefit the growth of crops in PFALs [21
In this study, three different lighting strategies were used, with varying photoperiods while maintaining the same DLI. The goal of the study was to determine the effects of different lighting strategies on growth of lettuce and to clarify the underlying mechanisms by investigating growth parameters, chlorophyll content, photosynthesis parameters, chlorophyll fluorescence parameters, and circadian rhythm.
Previous research has shown that higher light intensities promote growth and increase the production of crops [31
], and that lettuce yields were highest under a light intensity of 600 μmol photons m−2
]. However, in a plant factory, higher light intensity means higher energy consumption, and thus light intensities no higher than 300 μmol photons m−2
are usually used. Previous results have also shown that the qP and PhiPSII values of lettuce were low in the 100 μmol photons m−2
treatment, which translated to low light use efficiency and plant yields [24
]. Thus, the light intensity must be higher than 100 μmol photons m−2
in PFALs. To maximize the yield to cost ratio, the light intensities used in plant factories usually end up being below the optimal range, that is, natural light conditions, causing the plant to suffer from mild light stress that can limit plant production. In this study, lettuce growth was improved in both multi-segment light intensity and extended photoperiod lighting strategies, proving that both lighting strategies are effective at improving crop performance under weak light conditions in PFALs. In addition, the extended photoperiod lighting strategy improved the root/shoot ratio and chlorophyll content (Figure 2
). One study reported that the root/shoot ratio decreased as PPFD increased [6
], and Mozafar [33
] reported that the root/shoot ratio increased along with a photoperiod increase from 12 to 18 h. This study supports the results that the root/shoot ratio under the extended photoperiod lighting strategy is promoted when light intensity is reduced with an increased photoperiod. In addition, the increase in length of photoperiod under weak light conditions compensated for the low-light stress, promoting increases in the chlorophyll content of the crop and increasing competitiveness in weak lighting conditions. Increased chlorophyll content is beneficial to plants, helping to effectively absorb sufficient light energy at weaker light intensities by maximizing photosynthetic efficiency [34
]. From the results, we concluded that the extended photoperiod lighting strategy promoted the growth of roots and increased chlorophyll content, which enhanced the light-use efficiency, and thus resulted in better growth of lettuce.
According to the photosynthetic rate curve over 60 consecutive hours (Figure 3
), it can be seen that after transplanting the lettuce from natural light conditions to constant artificial light and temperature conditions, the circadian rhythm was maintained for a short period of time before gradually fading. The circadian clock of the plant regulates the circadian rhythm, which synchronizes internal physiological and biochemical processes with the external day and night cycle. Circadian rhythms enable plants to maintain high photosynthetic rates during the photoperiod and maximize biomass yields, providing a competitive advantage [17
]. According to the photosynthetic rates and the integral values of A, B, and C from this study, we observed that the multi-segment light intensity lighting strategy significantly increased diurnal photosynthetic capacity. This was because the multi-segment lighting strategy mimicked the changes of sunlight during a day, stimulating the crop to maintain its circadian rhythm, which maximizes photosynthetic rates despite the environmental changes. Thus, while the short photoperiod constant light intensity lighting strategy cleared the crop’s circadian rhythm (Figure 4
), the multi-segment lighting strategy took advantage of the circadian rhythm and increased photosynthesis when the light was strong, which enhanced the photosynthetic rate, and ultimately led to an increased yield. Therefore, this study suggested that the multi-segment lighting strategy can take advantage of circadian rhythms and increase crop yields while not increasing energy costs.
-PPFD curve, PNmax
, and AQY of the samples were determined to investigate the photosynthetic response of lettuce plants to the aforementioned three lighting strategies. The results revealed that the light saturation point of B and C were higher than A; this was due to the adaptation and adjustment of the photosynthetic mechanism to the various light energy regimens. The weak light treatment reduced the demand for light energy by the crop, and through adaptation to the weak light environment, crops increased their ability to use weak light. Moreover, the AQY reflected the photosynthetic capacity of the leaves under weak light. Higher AQY values indicate that there are more pigment protein complexes for the plant to use to absorb and convert light energy [35
]. Our results suggested that the lettuce plants in the extended photoperiod lighting strategy (A) had a higher ability to use weak light than the lettuce cultivated under the short photoperiod lighting strategy (B). In addition, the PNmax
of A was lower than B and C, reflecting reduced leaf photosynthetic capacity in lower light treatments, which results from reduced RuBP (Ribulose-1,5-bisphosphate) carboxylase activity in leaves. These results indicated that, even though this lighting strategy reduced the photosynthetic capacity of crops, it increased the ability of crops to utilize light under weak light conditions, which reduced the impact of weak light on crop yields. Thus, overall the extended photoperiod lighting strategy (A) increased the yields of the lettuce.
Chlorophyll fluorescence parameters can be used to assess the effects of many environmental factors on photosynthesis. The value of Fv
was shown to vary in the range of 0.8–0.84 for the majority of the C3 plants when the crops were not exposed to environmental stress. When the Fv
value was below 0.8, this indicated that the plant had been exposed to some environmental stressor, such as light stress [38
]. All the Fv
values were slightly below 0.8 (Figure 7
, panel 3) for all three lighting strategies, which showed that lettuce plants suffered a mild light stress under these three lighting conditions. The Fv
value in the lighting strategy A was the highest, indicating that increasing the photoperiod in a weak light environment somewhat compensated for weak light stress. Also, the values of Fv
′ were not different between the three lighting strategies, suggesting that the integrity of the photosynthetic apparatus was not affected. However, the quantum yield of PSII electron transport (Figure 7
, panel 3), the quantum yield of carboxylation rate (Figure 7
, panel 4), and photochemical quenching (Figure 7
, panel 5) were all improved as the lettuce adapted to the multi-segment lighting strategy (C). Fu [24
] reported that an appropriate increase in light intensity below the light saturation point can improve the efficiency of light energy utilization. Light energy absorbed by chlorophyll molecules goes through three consecutive processes [28
]: first, it is used to drive photosynthesis; second, excess light energy is dissipated as heat; third, it is re-emitted as chlorophyll fluorescence [29
]. It has been suggested that any increase in efficiency of one of these processes will lead to a decline in the other two, which is why the NPQ of strategy C was lower than the other two strategies [36
]. In other words, the multi-segment lighting strategy exposed lettuce to a higher light intensity level for a period of time each day, which meant the lettuce was better adapted to high light intensity conditions and was able to use more of the light energy to drive photosynthesis rather than it dissipating as heat. This also explains why the PNmax
of lighting strategy C was much higher than the other two strategies.
In addition, by examining the dynamics of Fo
, we concluded that the Fo
value in lighting strategy A was the largest, whereas Fo
in the B and C strategies were not significantly different from each other. These values reflected the differences in chlorophyll content. This is supported by Figure 2
, panel 8, where we can see that the chlorophyll content of lettuce in lighting strategy A was significantly higher than that of the other two strategies. The Fo
in all of the three different lighting strategies increased significantly after transplanting; there are two potential explanations for this. First, the white fluorescent lamps were used during the nursery period, and the red and blue LED light sources were used after transplanting. We checked the ratios of blue light in the two light sources and there was higher ratio of blue light in the red and blue LED lamp (33%) than the fluorescent lamp (21%). It has been shown that increasing the ratio of blue light can significantly increase the chlorophyll content of lettuce and increase the ratio of chlorophyll a/b [42
]. Secondly, because the lettuce was in the seedling stage, the plants had lower demand for light energy, and the lower chlorophyll content was able to meet the photosynthesis demands. After transplanting, however, with the change of light environment the lettuce was able to adapt by increasing chlorophyll content for better photosynthesis. Following the adaptation to the new light environment and the associated increase in growth, the demand for light energy also began to gradually increase. The lettuce increased the efficiency of light energy utilization by regulating the chlorophyll content, and thus Fo
also gradually increased. However, Fo
in strategy A decreased between the 7th and 14th days; this was likely due to the longer photoperiod of strategy A, as the longer photoperiod can help the lettuce partially compensate for the weak light. The lettuce in strategy A was better able to adapt to its light environment, therefore, it did not need to stimulate photosynthesis by increasing chlorophyll content. However, as the plants grew, the bigger leaves created more and more shading, resulting in even more severe weak light stress. Therefore, the lettuce in our experiment increased its chlorophyll content to cope with the weak light stress.
The dynamics of Fv/Fm for lettuce in all three lighting strategies were synchronous, with the maximum value appearing on the 14th day, and the minimum value appearing on the 28th day. However, the values of Fv/Fm in the different lighting strategies were significantly different, especially on the 28th day when the Fv/Fm value in strategy A was greater than the other two strategies, which was consistent with the previous conclusion that the extended photoperiod somewhat compensated for the effects of weak light stress. We also found that the Fv/Fm of lettuce in all three strategies was higher than 0.8 before the 14th day, before gradually decreasing until they were slightly lower than 0.8 on the 28th day. This showed that the lettuce in all treatment groups grew better before the 14th day, when they were not stressed by the external environment (mainly weak light stress in this study). After 14 days, however, the environmental stress gradually increased as the lettuce grew and the light intensity was not sufficient to meet the growth requirements. This trend was further exacerbated by the rapid expansion of leaf area, resulting in more and more self-shading as the number of leaves increased. Shading further reduced the light that is intercepted by the lettuce, whereas at the same time the demand for light energy rapidly increased, leading to the increased occurrence and severity of weak light stress. In addition, the values of Fv/Fm varied with the growth stage, indicating that the lettuce had different requirements for light energy at different growth stages.