Cannabis Inflorescence Yield and Cannabinoid Concentration Are Not Increased With Exposure to Short-Wavelength Ultraviolet-B Radiation

PAR and UV LED Fixture Layout

Photosynthetically active radiation was supplied by 24 LED fixtures (Pro650; Lumigrow Inc.) arranged evenly over the CB in 2 rows of 12 fixtures. The LED composition and spectrum of the PAR fixtures were described in Rodriguez-Morrison et al. (2021a) and the relative spectral photon flux distribution is provided in Figure 1A. Single UV LED fixtures were centered between adjacent PAR fixtures (within each row), resulting in 2 rows of 11 UV fixtures. The 22 UV LED fixtures were a custom design (10 × 90 cm), comprised of UVB LEDs with a peak wavelength of 287 nm (Figure 1B) and adjustable intensity (with analog, constant current dimmers). According to the conventional definitions of the different UV wavebands, the photon flux ratio of UVB (280 to 315 nm) to UVC (100 to 280 nm) was UVB(93):UVC(7). Additionally, 30% of the UV photon flux was at wavelengths >290 nm and there was no photon flux between 310 and 400 nm or <270 nm. The UV treatments (described below) were applied daily, in the last 3.5 hours (16:00 HR to 19:30 HR) of the PAR photoperiod, for 60 d from the day that the plants were transferred to the CB and then harvested.

Experimental Setup

The experiment was arranged and carried out as a gradient design, which can outperform treatment × replication designs when evaluating biological responses along a continuous independent variable (Kreyling et al., 2018), such as radiation intensity (Rodriguez-Morrison et al., 2021a). With a gradient design, regression analyses are performed on the response variables (i.e., measured parameters) against all different levels of the independent variable.

For each cultivar, 44 uniform representative plants were selected from the larger populations to be experimentally evaluated. Plots, each consisting of 4 plants of a single cultivar, were arranged where 2 plants were positioned directly below each UV LED fixture, and 2 plants were adjacent. Three UV LED fixture settings were randomly assigned (within each cultivar) to each plot: off, half power, and full power. Within each plot, the 2 plants positioned below the UV LED fixtures had relatively higher UV exposure than the 2 adjacent treatment plants. This configuration allowed for each cultivar to be exposed to a wide array of UV photon flux densities (UV-PFD); evenly-distributed across the 0.01 to 0.8 μmol⋅m^–2⋅s^–1 range (Figure 2).

At the start of the UV treatments, experimental plants of LT had heights from the substrate surface to the shoot apex ranging from 14 to 23 cm and experimental plants of BW had heights ranging from 14 to 20 cm. Experimental plants were surrounded by plants of the same cultivar to maintain canopy uniformity. The LT cultivar populated the south half of the CB, while BW populated the north half.

The average distance from the bottom of the fixtures to the top of the treatment plants was maintained at 50.5 cm by adjusting the height of the light racks weekly using a system of pulleys and cables. Canopy-level PPFD and UV-PFD were measured at the apex of each plant weekly, after the light rack height adjustment, using a PAR meter (LI-180; LI-COR Biosciences, Lincoln, NE, United States) and a radiometrically calibrated spectrometer (UV-Vis Flame-S-XR, Ocean Optics, Dunedin, Florida), respectively. The UV-PFDs were measured with the PAR LEDs turned off. A MS Excel tool developed by Mah et al. (2019) was used to integrate spectrometer data into UV-PFD, biologically-effective UV-PFD (Flint and Caldwell, 2003), and daily biologically-effective UV dose (kJ⋅m^–2⋅d^–1) (Table 1). At the end of the trial, average PPFD and average UV-PFD were calculated for each treatment plant by determining the corresponding total light integrals (TLI; mol⋅m^–2) and then dividing by accumulated time, as described in Rodriguez-Morrison et al. (2021a). The experiment-wise average (± SE, n = 88) PPFD was 408 ± 6.5 μmol⋅m^–2⋅s^–1. The average UV-PFD for each plant was used as the independent variable (i.e., x-axis) in regressions of UV exposure vs. the measured growth, yield and quality parameters.

Plant husbandry and environmental controls followed the cultivator’s standard operating procedures except for the UV radiation. The air temperature and relative humidity set points were 25°C and 60%. There was no CO₂ supplementation, with typical concentrations of ≥400 ppm when the PAR lights were on. Air was continuously circulated throughout the room with wall-mounted axial fans and the HVAC circulation rate was ≈2 air changes per hour (ACH) with ≥25% refresh with pre- conditioned outside air. The aquaponic solution was maintained within normal levels of nutrient concentrations, pH, electrical conductivity, and dissolved oxygen; as described in Rodriguez-Morrison et al. (2021a).

Growth Measurements

The height (i.e., length of main stem from substrate surface to the highest point) and widths (i.e., the widest part and its perpendicular width) of each experimental plant were measured in week 6. Plant height and widths were used to calculate growth index [(height × width₁ × width₂)/300 (Ruter, 1992)] for each plant, where width₁ was the widest part of the plant and width₂ was perpendicular to width₁.

Leaf Chlorophyll and Fluorescence Measurements

Foliar chlorophyll content index [CCI; the ratio of % transmission 569 at 931 nm and % transmission at 653 nm (Parry et al., 2014)] was measured in upper and lower canopy leaves in week 3. Measurements of CCI were taken from the center leaflet of the three youngest fully-expanded fan leaves and from three fan leaves at the bottom of each plant using a chlorophyll meter (CCM-200; Opti-Sciences, Hudson, NH, United States). The CCI measurements were averaged, for the upper and lower canopy leaves, respectively, on a per plant basis.

The ratio of variable to maximum fluorescence (F_v/F_m) emitted from photosystem II in dark-acclimated leaves exposed to a light-saturating pulse is an indicator of maximum quantum yield of photosystem II photochemistry (Murchie and Lawson, 2013). In week 5, during the first 8.5 h of the PAR photoperiod (i.e., before daily UV exposure), the middle leaflet of the youngest fully-expanded fan leaf from each plant was dark acclimated for 15 min and then F_v/F_m measurements were taken with a fluorometer (FluorPen FP 100; Drasov, Czech Republic).

Leaf Gas Exchange, Leaf Size and Specific Leaf Weight

Quantifications of leaf gas exchange of the middle leaflet of the youngest, fully-expanded fan leaf on each treatment plant was performed in week 5 during the first 8.5 h of the PAR photoperiod using a portable photosynthesis machine (LI-6400XT; LI-COR Biosciences, Lincoln, NE, United States) equipped with the B and R LED light source (6400-02B; LI-COR Biosciences). In situ net CO₂ exchange rate (NCER) was measured with the leaf cuvette environmental conditions set to: PPFD of 500 μmol⋅m^–2⋅s^–1, block temperature of 26.7°C, CO₂ concentration of 400 ppm, and air flow rate of 500 μmol⋅s^–1. Because the leaflets were not wide enough to cover the entire cuvette, the section of each leaflet that was clamped in the cuvette gasket was marked along the outside of the gasket with a permanent marker so that leaf area inside the cuvette could be calculated post hoc (described below). After removing the leaflets from the cuvette, whole leaves were excised from the plant and scanned (CanoScan LiDE 25; Canon Canada Inc., Brampton, ON, Canada) at 600 dpi resolution. Each leaf was oven dried to constant weight at 65°C (Isotemp Oven 655G; Fisher Scientific, East Lyme, CT, United States). The scanned images were processed using ImageJ 1.42 software (National Institute of Health; https://imagej.nih.gov/ij/download.html) to determine the leaflet area within the gas exchange chamber (by subtracting the width of the chamber gaskets from the marks made during gas exchange measurements) and the total individual leaf size. The NCER for each leaf was corrected for measured leaf area inside the cuvette. The dry weight (DW) of each entire scanned leaf was measured using an analytical balance (MS304TS/A00; Mettler-Toledo, Columbus, OH, United States) to determine specific leaf weight [SLW; leaf DW/leaf size (g⋅m^–2)].

Visual Observations

Weekly observations were performed on each entire plant to visually evaluate observable changes in morphological attributes, including: upward curling of the leaflet margins, leaf shine, browning of stigmas, leaf epinasty, leaf necrotic patches, and appearance of powdery mildew on the adaxial sides of the leaves. Except for week 1 observations, which occurred 4 d after the start of the UV treatments, all weekly observations occurred on 7-d intervals thereafter. The absence or presence of each respective parameter was evaluated for each plant weekly, except where noted in the results (Table 2). While these are observational data, the minimum UV-PFDs under which individual attributes were observed were reported, on per cultivar and per week bases, regardless of whether all plants exposed to higher UV-PFDs displayed the observed responses.

At various points throughout the trial, photos of representative whole plants and specific tissues of each cultivar growing under different UV exposure levels were taken with a digital camera (iPhone XR iOS 14.4.1; Cupertino, CA, United States) or flat bed scanner (CanoScan LiDE 25). Photos of whole plants grown under minimum and maximum UV exposure levels were taken in week 2. Photos of the inflorescences grown under minimum and maximum exposure levels, of each cultivar, were taken in week 3. Photos of whole plants grown under minimum, low, moderate and maximum UV exposure levels (described in Table 1) were taken in week 3. In week 5 [i.e., approximately when vegetative growth in cannabis ceases (Rodriguez-Morrison et al., 2021a)], fully-expanded leaves from plants under minimum, moderate, and high UV exposure were excised from the plants and scanned at 600 dpi resolution. Photos of whole plants and apical inflorescences grown under minimum, low, moderate, and maximum UV exposure levels were taken at harvest (i.e., week 9). All photos were processed using ImageJ 1.42 software to add appropriate scale bars.

Yield and Quality

After 60 d of UV exposure, all treatment plants were harvested by cutting the stems at substrate level. The LEDs were turned off prior to harvest and plants were harvested, randomly, one at a time to minimize any harvesting effects on fresh biomass assessments. The aboveground tissue of each plant was separated into stems, leaves, and inflorescences. The inflorescences were further subdivided into apical (i.e., grouping of terminal inflorescences at the top of the main stem, located above the uppermost side-branch) and non-apical groupings. All inflorescence tissues were trimmed of foliar materials, according to the cultivator’s normal practices. The fresh weights (FW) of stems, leaves, and apical and non-apical inflorescence were separately recorded for each plant using a precision balance (EG 2200-2NM; Kern, Balingen, Germany). The apical inflorescences for 18 plants of each cultivar that were representative of the entire range of UV-PFD exposure levels were air dried at 15°C and 40% relative humidity for 7 d. Following air drying, ≈2 g sub-samples (actual weights were recorded) from each plant were submitted to an independent laboratory (RPC Science & Engineering; Fredericton, NB, Canada) for analysis of concentrations [reported in mg⋅g^–1_(DW)] of cannabinoids using ultra-high-performance liquid chromatography and variable wavelength detection (HPLC-VWD), terpenes using gas chromatography and mass spectrometry detection (GC-MSD), and moisture content. The total equivalent (annotated with: eq) Δ⁹-THC, CBD, and cannabigerol (CBG) concentrations were determined by assuming complete carboxylation of the acid-forms of the respective cannabinoids, whose concentrations were adjusted by factoring out the acid-moiety from the molecular weight of each respective compound [e.g., Δ⁹-THCeq = (Δ⁹-THCA × 0.877) + Δ⁹-THC]. The remaining apical tissues from the air-dried samples and the entire apical inflorescences of the non-air-dried plants were re-combined with non-apical inflorescences to make up total inflorescence groupings, on a per-plant basis. All separated aboveground tissues of all plants were oven-dried at 65°C to constant weight (Isotemp Oven 655G) and the DW of the respective tissues were recorded. Moisture content of each separated aboveground tissue grouping was calculated as: [(FW – DW) / FW] × 100%. The sub-sampled apical tissues were accounted for in this calculation using their respective sample weight and moisture content measurements for each sample, provided by RPC.

UV-Induced Cannabis Morphology and Physiology Changes

While the aboveground portions of each entire plant was observed for UV-induced changes in morphology, the recorded effects occurred primarily in recently-developed tissues. When UV effects were also seen in older tissues, this has been highlighted in the text. The data in Table 2 are provided to show how the development of temporal trends in these observed parameters related to each other, and how crop sensitivity to UV exposure increased over time.

The first observed UV-induced changes in cannabis morphology appeared within the first few days of the initiation of the UV treatments where the leaflet margins on leaves that had developed in the vegetative stage (i.e., prior to the initiation of the UV exposure) curled upwards under UV-PFDs ≥0.33 and ≥0.37 μmol⋅m^–2⋅s^–1 in LT and BW, respectively during week 1 (Table 2). Leaves also appeared to accumulate epicuticular wax, as demonstrated by the increase in shiny appearance of adaxial surfaces, shortly after UV exposure began and persisted henceforth (data not shown). Leaf shine appeared to be more prevalent in plants exposed to higher UV-PFDs, and the prevalence appeared to be greater in BW vs. LT. In week 2 there were no changes in the extent of upward curling in mid-canopy foliage (i.e., leaves that had developed during the vegetative stage) however, newly expanded leaves did not appear to present this symptom with the same level of severity (Figure 3). In week 3, which was about one week after the presence of inflorescence tissues were readily apparent, stigmas of terminal inflorescences began to turn from white to brown on LT and BW plants exposed to UV-PFDs ≥0.69 and ≥0.30 μmol⋅m^–2⋅s^–1, respectively (Figure 4 and Table 2). In week 3, early symptoms of foliar epinasty (i.e., interveinal tissues that were raised in the middle) started to appear in upper canopy leaves only of plants grown under higher UV-PFDs (Figure 4). Some leaves at the bottom of the plants started to become yellow and drop off in week 3 for both cultivars under higher UV-PFDs (data not shown). Fallen leaves appeared to be predominantly the same leaves that showed upward curling in week 1. There were no UV treatment effects on the CCI of the upper canopy leaves of LT in week 3, but the CCI of the upper canopy leaves of BW decreased linearly by 42% from the lowest to highest UV-PFD (Figures 5A,B). The CCI in the lower canopy leaves decreased linearly by 60% and 46% from the lowest to highest UV-PFD in LT and BW, respectively (Figures 5C,D). In week 4, the minimum UV-PFD at which plants exhibited stigma browning were lower than the previous week (Table 2). In week 5, the in situ NCER of the youngest fully-expanded leaves decreased linearly with increasing UV exposure, with 31% and 27% lower NCER at highest vs. lowest UV-PFD in LT and BW, respectively (Figures 5E,F). In week 5, the F_v/F_m values of the youngest fully-expanded leaves decreased linearly by 9% and 19% in LT and BW, respectively (Figures 5G,H). The severity of UV-induced epinasty was elevated in plants exposed to higher UV exposure levels, as shown in images of whole plants in week 3 (Figure 6) and single-leaf scans in week 5 (Figure 7) of representative plants grown under different UV exposure levels. In week 5, upper canopy leaves in particular showed upward curling under UV-PFDs ≥0.16 and ≥0.34 μmol⋅m^–2⋅s^–1 in LT and BW, respectively (Table 2 and Figure 7). In week 5, brown stigmas were observed on plants grown under the lowest UV-PFD in LT and ≥0.14 μmol⋅s⋅m^–2 in BW (Table 2). In week 5, leaf epinasty was predominantly evident in youngest fully-expanded leaves in plants exposed to UV-PFDs ≥0.13 μmol⋅m^–2⋅s^–1 in LT and ≥0.14 μmol⋅m^–2⋅s^–1 in BW (Table 2). In week 5, the size of the youngest fully-expanded leaves decreased linearly with increasing UV-PFD, with ≈45% reductions in young leaf size in plants grown under highest vs. lowest UV-PFD (Figures 8A,B). There were corresponding linear increases in SLW with increasing UV exposure, with 27% and 21% increases in LT and BW, respectively, in plants grown under the highest vs. lowest UV-PFD (Figures 8C,D). Brown stigmas were observed in all experimental plants starting in week 6 (Table 2). Starting in week 7, upper canopy leaves on a few LT plants grown under intermediate UV-PFDs, ranging from 0.12 to 0.69 μmol⋅m^–2⋅s^–1, began to show brown (necrotic) patches (Table 2). The minimum UV-PFDs under which leaf epinasty was evident were marginally lower in week 7 vs. week 5, and substantially lower in week 8 vs. week 7 (Table 2) in both cultivars. The prevalence of leaves exhibiting necrotic patches increased in BW in week 8 vs. week 7 (Table 2). Investigating the effects of UV exposure on foliar powdery mildew was not one of the objectives at the outset of this study, however, some potential treatment effects were observed. For example, evidence of powdery mildew was visible on the adaxial leaf surfaces on plants exposed to low UV-PFDs at harvest, but was not observed on any plants exposed to UV-PFDs ≥0.090 μmol⋅m^–2⋅s^–1 (Figure 9). By harvest, there also appeared to be greater incidences of foliar chlorosis in both cultivars, especially in the lower canopy, with increasing levels of ultraviolet exposure (Figure 9).

FIGURE 6

Representative (A) ‘Low Tide’ and (B) ‘Breaking Wave’ Cannabis sativa plants demonstrating (from left to right) minimum, low, moderate, and high UV exposure levels. The images were taken in week 3 after the initiation of the UV treatments. The black scale bar at the lower right of each image is 5.0 cm.

FIGURE 9

Gross plant morphology of representative (A) ‘Low Tide’ and (B) ‘Breaking Wave’ Cannabis sativa plants grown under (from left to right) minimum, low, moderate, and high UV exposure levels. Images were taken just prior to harvest (i.e., 9 weeks after the initiation of UV treatments). Note the white spots (powdery mildew) on the adaxial sides of leaves on the far-left plants in both images. The black scale bar at the upper left of each image is 5.0 cm.

Growth Responses to UV

Exposure to UV generally suppressed plant growth, which was recorded as increase in height and growth index in week 6. The majority of vegetative growth had ceased by week 6 given that height only increased 1.76 ± 0.41 cm and 0.62 ± 0.48 cm (mean ± SE) for LT and BW respectively, between week 6 and harvest (data not shown). Both increase in height and growth index had negative linear relationships with increasing UV-PFD in both cultivars. Increases in height were 31% and 26% lower in plants grown under the highest vs. lowest UV-PFDs in LT and BW, respectively (Figures 8E,F). Growth indices were 61% and 33% lower in plants grown under the highest vs. lowest UV-PFDs in LT and BW, respectively (Figures 8G,H). There were no UV treatment effects on the moisture content of any aboveground tissues in either cultivar (Table 3).

UV Radiation Alters Cannabis Morphology and Physiology

Leaf morphology demonstrated substantial plasticity in response to UV radiation exposure throughout the 9-week flowering stage. The first observed morphological response to UV was upward curling of leaflet margins during the first week of UV exposure. Upward leaf curling was most evident under higher UV-PFDs, and it occurred primarily on the youngest leaves (i.e., that developed in the vegetative stage, just prior to UV exposure). Upward leaf curling under UV stress is not a commonly-reported morphological response, although it has been observed in cotyledons of canola (Wilson and Greenberg, 1993). Upward leaf curling has been a more commonly- reported response to pathogen infection in some crops (Taliansky et al., 2003; Halldorson and Keller, 2018) and to various physiological stresses in tomato, including light stress (Powell et al., 2014). The recently-developed leaves in the present study may also have lacked the acclimative plasticity of the leaves that later developed under UV exposure, which exhibited more typical UV-induced morphology responses such as epinasty, reduced leaf size and increased leaf thickness indicated by SLW (Wilson, 1998; Searles et al., 2001; Zlatev et al., 2012; Fierro et al., 2015). Further, the apparent increase in leaf shine shortly after the initiation of UV exposures indicates an accumulation of epicuticular wax, which is a common response to UV exposure in other crops (Steinmüller and Tevini, 1985; Cen and Bornman, 1993; Fukuda et al., 2008; Valenta et al., 2020). All of these observed morphological responses to UV exposure may reduce the potential for damage to the photosynthetic machinery.

UV radiation accelerated plant senescence (i.e., deterioration with age) symptoms in both inflorescence and foliar tissues. Female inflorescence maturation can normally be characterized by carpel swelling and the transition of stigmas from white to brown in the final days before harvest (Punja and Holmes, 2020). Although the number of days between the initiation of the 12-h photoperiod and appearance of inflorescences was unaffected by UV exposure (data not shown), plants exposed to higher UV-PFDs exhibited earlier stigma browning (Figure 4). It is unknown if premature stigma senescence has any knock-on effects on other inflorescence development parameters, such as production of secondary metabolites. However, since the rate of stigma browning depended on UV exposure levels, this attribute could not be used reliably to determine the “optimum harvest maturity” for the plants in this trial.

Foliar chlorophyll content is often negatively correlated to UV exposure level (Neugart and Schreiner, 2018). Both cultivars showed increasing SLW with increasing UV exposure; similar to cannabis exposed to high PAR intensity (Rodriguez-Morrison et al., 2021a). While the CCI levels measured in this study were within the ranges of cannabis leaves reported by others (Caplan et al., 2017; Yep et al., 2020; Rodriguez-Morrison et al., 2021a), there were only UV treatment effects on CCI in the upper canopy leaves of BW. However, LT showed relatively greater increases in SLW with increasing UV exposure. Since foliar thickness affects a leaf’s optical properties, hence CCI measurements (Parry et al., 2014), LT’s enhanced increases in leaf thickness may have offset any UV-induced reductions in chlorophyll concentration on a biomass basis (e.g., mg⋅g^–1) in this cultivar. While lower vs. upper canopy leaves in indoor-grown cannabis may have lower CCI, regardless of plant age (Rodriguez-Morrison et al., 2021a), the reductions in lower-canopy chlorophyll content with increasing UV-PFD in both cultivars may be another indication of the UV damage done to young leaves at onset of UV exposure. This damage eventually manifested as higher rates of early leaf-drop and increased leaf chlorosis at harvest in plants exposed to higher UV-PFDs. Both of these phenomena have also been observed in UV-stressed sweet basil (Dou et al., 2019). Nitrogen from lower canopy leaves is normally remobilized to more active upper canopy foliage as plants age (Havé et al., 2017); this appeared to be accelerated by UV exposure given the reductions in lower-canopy CCI after only 3 weeks of exposure. Foliar necrosis is also a commonly-observed symptom of UV damage in many species (Maffei and Scannerini, 2000; Zhao et al., 2003; Dou et al., 2019). While the severity of most of the observed UV stress responses increased with increasing UV exposure, necrotic patches were observed on upper canopy leaves that were predominantly exposed to intermediate UV-PFDs (primarily in LT) in the latter weeks of the trial. The acclimation (e.g., epinasty, curling, reduced size) of leaves exposed to the highest UV-PFDs may have mitigated foliar damage, while the leaves grown under intermediate UV-PFDs may not have been sufficiently acclimated for long-term UV exposure. Unstressed leaves normally have F_v/F_m values of ≈0.8 (Björkman and Demmig, 1987). While the reductions in F_v/F_m in upper canopy leaves of both cultivars were similar to cannabis plants exposed high PAR intensity in Rodriguez-Morrison et al. (2021a), the opposite effects of increasing UV vs. PAR radiation on NCER is strongly indicative of UV-induced damage to the photosynthetic machinery.

Lydon et al. (1987) reported no UV treatment effects on the cannabis morphology and physiology parameters they measured, which is in stark contrast to the copious morphological and physiological UV-induced stress responses observed in the present study. Evidently, the plants in the present study were subjected to more efficacious UV radiation treatments than in Lydon et al. (1987) despite similar reported maximum biologically-effective UV doses in both studies. This may be partly due to the shorter-wavelength UV spectrum in the present study. Further, the plants in Lydon et al. (1987) may have experienced lower than reported doses due to rapid UV-induced reductions of UVB transmissivity of the cellulose acetate filters they used to eliminate UVA and PAR wavelengths from their UV spectrum treatments (Middleton and Teramura, 1993). Additionally, their plants grew for several months under greenhouse conditions (likely including some UV) prior to exposure to UV treatments, whereas there was no UV exposure of the plants prior to initiation of the UV treatments in the present study. Therefore, light history (e.g., spectrum and intensity) and plant age may affect how plants to acclimate to new UV stresses.

UV Radiation Suppresses Cannabis Growth and Yield

While increasing UV radiation exposure suppressed overall vegetative plant growth (e.g., height and growth index) in both cultivars, the responses were more severe in LT than BW. However, these are in contrast with the UV-induced reductions in foliar biomass, which were substantially greater in BW. This was particularly surprising given that there were no consequent reductions in total inflorescence biomass in BW. In fact, despite some leaf senescence observed in both cultivars, harvest index – which is the ratio of inflorescence DW to total aboveground DW – went up by ≈10% in BW and went down by ≈10% in LT as UV-PFD increased from lowest to highest. Under low UV exposure, the harvest index for both cultivars was ≈0.6, which was similar to a different cultivar grown under the same PPFD in the same production system without UV (Rodriguez-Morrison et al., 2021a). Given that there were no UV exposure effects on inflorescence DW in BW, earlier and/or elevated foliar senescence in BW may have contributed to its relatively elevated harvest index.

Reduced aboveground biomass and lower yields are commonly observed effects of UV radiation on some other plant species (Teramura et al., 1990; Fiscus and Booker, 1995; Caldwell et al., 2003; Liu et al., 2005). The UV-induced alterations in leaf morphology and physiology probably contributed to the general reductions in growth and overall biomass in both cultivars. For example, reduced leaf area is a typical response to radiative stresses such as high PAR intensity and UV exposure (Wargent and Jordan, 2013; Poorter et al., 2019). In the present study, the reductions in individual leaf size, total foliar biomass, and leaf-level NCER with increasing UV exposure, would have limited the plants’ capacity to convert PAR into biomass (Kakani et al., 2003; Zlatev et al., 2012).

Total inflorescence DW and the proportion of that DW which is comprised of apical tissues are two major considerations for commercial cannabis production. The apical proportion may be of particular interest since these tissues are normally considered premium quality due to their relatively large size and potentially higher cannabinoid concentrations compared to higher-order (i.e., on lower branches) inflorescences (Namdar et al., 2018). Despite the UV-induced limitations to biomass accumulation seen in both cultivars, increasing UV exposure only reduced inflorescence DW in LT. Within this context, the various growth habits of common indoor-grown cannabis cultivars may influence their yield responses to UV stress. In the present study, BW and LT had disparate whole-plant reproductive macro-morphology (i.e., the distribution of inflorescence biomass within the canopy) under normal indoor conditions. For example, under minimum UV exposure, the apical inflorescence comprised 24% of the total inflorescence DW in LT compared to only 11% in BW. Apparently, growth habit may have predisposed BW’s mitigation of UV-induced yield reductions by partitioning relatively more inflorescence biomass to positions farther away (i.e., more protected from the UV) from the top of the plant. However, while this may be a self-protective response to reduce UV exposure to reproductively important (from an ecological sense) tissues, it still came at commercially-objectionable reductions in inflorescence quality, such as visually unappealing morphology (Figure 10).

To prevent UV-induced yield losses, such as are reported in the present study, it is conceivable that cannabis plants could be exposed to UV only after the majority of vegetative growth has completed [i.e., a few weeks after the visual appearance of inflorescences (Potter, 2014)]. This strategy would shorten the accumulated period of exposure to UV stress and may minimize some UV-induced foliar acclimations that could inhibit biomass accumulation. However, there is a risk that later-term UV exposure might also sufficiently stress unacclimated foliar tissues to provoke rapid-onset whole-plant senescence before the inflorescences reach optimum maturity. This strategy warrants further exploration.

UV Radiation Alters the Secondary Metabolite Composition of Cannabis Inflorescences

The most economically relevant cannabinoids (i.e., Δ⁹-THC and CBD) are predominantly found in their acid forms in mature female inflorescence tissues, which are converted to the psychoactive and medicinal neutral forms through decarboxylation (Eichler et al., 2012; Zou and Kumar, 2018). The neutral forms also exist in relatively low quantities in the fresh inflorescences and tend to increase in proportion to the acid forms as the inflorescences mature (Aizpurua-Olaizola et al., 2016). While the Δ⁹-THC concentration increased in BW with increasing UV-PFD, it was a relatively small proportion of the Δ⁹-THCeq; maximized at 3.3% at the highest UV-PFD. Further, CBN was undetectable in the inflorescences, which is an indicator that the crops were not past peak maturity at the time of harvest since Δ⁹-THC naturally degrades to CBN (Russo, 2007). There were no UV-induced enhancements to Δ⁹-THCeq, CBDeq, and CBGeq in either cultivar. These results are consistent with a recent study that found no UV treatment effects on Δ⁹-THCeq content in a Δ⁹-THC-dominant cultivar (Llewellyn et al., 2021), but contrast with studies on older genotypes (Pate, 1983; Lydon et al., 1987). For example, Lydon et al. (1987) found that inflorescence Δ⁹-THC concentrations increased linearly from 32 to 25 mg⋅g^–1 in greenhouse-grown cannabis as UV exposure increased from their no-UV control up to biologically-effective UV doses (based on Caldwell, 1971) of 13.4 kJ⋅m^–2⋅d^–1. These contrasting results may be due to the disparate growing conditions (both before and during UV exposure), plant age at the time of UV exposure, and the relative magnitude of cannabinoid concentrations. Further, while the proportional increases in Δ⁹-THC content (28%) presented in Lydon et al. (1987) appeared to be substantial, the magnitude of their increase (i.e., only 7 mg⋅g^–1) is probably inconsequential in the context of cannabinoid composition in modern genotypes which can have Δ⁹-THC concentrations that exceed 200 mg⋅g^–1 (Dujourdy and Besacier, 2017).

Pate (1983) reported an increase in the ratio of Δ⁹-THC to CBD in inflorescence tissues of cannabis ecotypes grown in global positions with naturally higher UV exposures, which suggests that the production of Δ⁹-THC may be upregulated and CBD downregulated as adaptations (i.e., over multiple generations) to the localized environment. However, the results of the present study do not support this trend, at least as an acclimation response to UV stress of a single generation. Additionally, De Meijer et al. (2003) showed that cannabinoid profiles are largely genetically predetermined (e.g., a CBD-dominant cultivar is lacking the genetic predisposition to generate abundant Δ⁹-THC). This favors the concept that the upregulation of Δ⁹-THC under UV stress may be an adaptive response (i.e., over generations) rather than an acclimation response (i.e., during a single production cycle). Over the past few decades, there have been radical increases in inflorescence cannabinoid concentrations, which is often attributed to intensive breeding programs (Chouvy, 2015; Dujourdy and Besacier, 2017; Aliferis and Bernard-Perron, 2020) and the “sinsemilla” cultivation method that eliminates seeds and chiefly produces high potency female inflorescences (ElSohly et al., 2016). Thus, these factors may have a larger impact on cannabis inflorescence cannabinoid composition in indoor production than environmental factors such as UV stress.

While cannabinoids comprise the primary psychoactive and medicinal compounds in cannabis inflorescences, volatile terpenes are also economically valuable; both for the aromas that influence consumer preference and potential medicinal properties (Nuutinen, 2018; Booth and Bohlmann, 2019). UV exposure equivocally altered the terpene composition in the present study, with disparate responses within the different terpenes and between cultivars. However, total terpene concentrations in both cultivars decreased linearly with increasing UV exposure, which would tend to depreciate the overall quality of aromas and extracts (McPartland and Russo, 2001; Nuutinen, 2018).

While UV exposure did not result in any economically relevant increases in cannabinoid or terpene concentrations in cannabis inflorescences under the conditions of the present study, UV radiation has been shown to increase concentrations of UV-absorbing secondary metabolites (e.g., flavonoids and phenolic compounds) in many species (Huché-Thélier et al., 2016; Robson et al., 2019), including economically important essential oil producing crops (Schreiner et al., 2012; Neugart and Schreiner, 2018). However, UV-induced increases in secondary metabolite concentrations are often concurrent with biomass reductions (Fiscus and Booker, 1995; Caldwell et al., 2003). This paradox must be evaluated when considering the use of UV radiation to manipulate secondary metabolite composition in indoor cannabis production, since the simultaneous yield reduction may offset any improvements in secondary metabolite composition.

Compared to the UV spectra employed in most other studies, the biologically effective doses in the present study were dramatically higher for a given photon flux density due to the very short peak wavelength of the UV LEDs. In fact, ≈70% of the UV photon flux were at wavelengths below 290 nm, and thus outside of the solar spectrum that plants would naturally be exposed and adapted to Nikiforos et al. (2011). Therefore, cannabis may respond dramatically differently to UV from slightly longer wavelength LEDs (e.g., 300 to 315 nm).

We thank Derek Bravo, Tim Moffat, and Madeline Baker for technical support throughout the experiment. We also thank Angus Footman and Erica Emery for their logistical support. This article was first published as a preprint (Rodriguez-Morrison et al., 2021b).