Photosynthetic and Chlorophyll Fluorescence Characteristics of Isodon rubescens (Hemsley) H. Hara

Abstract

The ecological and economic cultivation of Isodon rubescens is currently being carried out. The demand of I. rubescens for light intensity should be made clear to estimate whether the environmental conditions of an area are suitable for cultivating I. Rubescens and improve cultivation techniques. The photosynthetic and chlorophyll fluorescence characteristics of I. rubescens were determined with a Li-6400 photosynthesis system and PAM-2500 portable chlorophyll fluorescence apparatus. The results showed that there was no obvious midday depression of photosynthesis in I. rubescens leaves. The light compensation point and light saturation point of I. rubescens leaves were 21.83482 µmol·m⁻²·s⁻¹ and 802.262 µmol·m⁻²·s⁻¹, respectively. The CO₂ compensation point and CO₂ saturation point of I. rubescens leaves were 101.7199 µmol·mol⁻¹ and 1674.514 µmol·mol⁻¹, respectively. The maximal photochemical efficiency of photosystem II ((Fm-Fo)/Fm) in I. rubescens leaves reached 0.7. The electron transport rate of photosystem II in I. rubescens leaves reached 20 μmol electrons/(m²·s). I. rubescens can tolerate intense light above the light compensation point and utilize low light. I. rubescens leaves have a strong photoprotective capacity. I. rubescens can grow in both sunny and shady places. The most important factor affecting photosynthetic efficiency in I. rubescens leaves is the concentration of CO₂ in air.

Introduction

Isodon rubescens (Hemsley) H. Hara is a perennial subshrub belonging to a genus of the Lamiaceae family¹. There are several bioactive chemical components in I. rubescens, such as oridonin and ponicidin. The dry aerial portions of I. rubescens are named rabdosiae rubescentis herba and are used in traditional Chinese medicine for the treatment of sore throats, inflammation and gastrointestinal problems^2,3.

The ecological and economic cultivation of I. rubescens is currently being carried out. However, there are different environmental conditions in different places. Wild I. rubescens grows on mountains or hills. There are obvious differences between the environmental conditions of mountains and plains. The demand of I. rubescens for light intensity should be made clear. The photosynthetic and chlorophyll fluorescence characteristics of I. rubescens were studied in this research to define the most suitable environmental conditions for I. rubescens cultivation and improve cultivation techniques.

Results

Diurnal variation in I. rubescens leaf photosynthesis

The results of the diurnal variation in I. rubescens leaf photosynthesis are shown in Table 1. Based on the collected data, the curve of the diurnal variation in I. rubescens leaf photosynthesis is shown in Fig. 1.

Table 1.

Diurnal variation in I. rubescens leaf photosynthesis (average).

Time	Photo (µmol CO2·m−2·s−1)	Transpiration (mmol H2O m−2s−1)	PARout (µmol·m−2·s−1)	Tleaf (°C)	CO2 (µmol·mol−1)
8:20	0.134554	0.078752	165.3032	27.48817	374.3593
10:01	0.489859	0.100056	690.494	31.51378	364.5589
11:09	0.677691	0.16249	923.3254	33.61967	384.1379
11:46	2.431713	2.894369	1211.345	34.83849	370.7948
12:33	3.21911	2.629756	1484.135	35.78894	361.1602
13:25	2.385099	2.6251	1697.34	36.47706	363.1944
14:53	2.390117	2.888978	1346.388	35.82655	370.8038
16:31	1.619721	2.060572	524.8741	33.17798	359.4734
17:15	1.067346	1.362125	280.3203	32.90131	359.0663
18:09	0.166573	0.043506	163.6786	31.8848	370.7527
19:08	0.146304	0.063026	145.7678	30.09128	366.364
19:23	0.038786	0.050609	86.26784	29.08384	366.6463

Note: PARout is the PAR out of the leaf chamber.

Figure 1.

Diurnal variation in I. rubescens leaf photosynthesis.

The diurnal variation in I. rubescens leaf photosynthesis indicates that there was no obvious midday depression of photosynthesis. There is still a high net photosynthetic rate in I. rubescens leaves at noon with high light intensity. Leaves of I. rubescens can utilize very faint light, e.g., with a 20 µmol·m⁻²·s⁻¹ intensity. The leaves of I. rubescens can photosynthesize even in the faint light of evening.

Light response curve of I. rubescens leaves

The data from the light response curve of I. rubescens leaf photosynthesis are shown in Table 2. Based on the collected data, the curve of the light response of I. rubescens leaf photosynthesis is shown in Fig. 2.

Table 2.

Light response curve of I. rubescens leaf photosynthesis (average).

PARin (µmol·m−2·s−1)	Photo (µmol·m−2·s−1)	Transpiration (mmol m−2s−1)	Tleaf (°C)	CO2 (µmol·mol−1)
3124.457	2.260209	4.466418	29.3982	395.2443
2198.982	4.721694	3.573953	30.04687	392.505
2000.081	4.965527	3.807614	30.61862	391.8968
1800.327	5.001151	3.645889	29.77489	392.8282
1500.057	5.38657	3.336598	30.33475	392.6403
1200.517	5.329608	2.987329	30.81813	391.6764
1001.321	5.443283	2.672035	30.69158	391.8007
799.9315	5.541983	2.354444	30.53654	391.9345
599.9705	5.559295	2.089824	30.37206	391.9419
499.3938	5.348708	1.890056	30.96318	392.7756
400.1194	5.171424	1.844144	30.15175	393.0815
200.3279	3.963617	1.612985	30.92859	394.4694
149.6206	3.822843	1.509174	30.68748	395.1866
99.69547	2.725072	1.457733	30.33666	396.0288
70.8973	1.200702	1.417435	30.03102	398.5548
30.33922	0.248815	1.142081	29.94907	399.1747
21.26013	0.1307	1.322936	29.69206	399.0078
E	0.050712
M	0.00021
N	0.005891
LCP	21.83482 µmol·m−2·s−1
E·LCP	1.107
LSP	802.262 µmol·m−2·s−1
PLSP	5.7471 µmol·m−2·s−1
R2	0.97336

Note: PARin is the PAR in the leaf chamber. PLSP is the net photosynthetic rate at the light saturation point.

Figure 2.

Light response curve of I. rubescens leaf photosynthesis.

The light response curve of I. rubescens leaf photosynthesis indicates that the net photosynthetic rate was obviously related to the light intensity when the light intensity was low. The net photosynthetic rate of I. rubescens leaves rapidly increased as the light intensity increased from 20–400 µmol·m⁻²·s⁻¹. I. rubescens leaves were able to utilize intense light. With light intensities of 400–2200 µmol·m⁻²·s⁻¹, the net photosynthetic rate of I. rubescens leaves was high. However, the net photosynthetic rate of I. rubescens leaves obviously decreased when the light intensity was above 2200 µmol·m⁻²·s⁻¹.

The results of the light response curve fitted with the modified rectangular hyperbola model are shown in Table 2. The fitted light saturation point and the net photosynthetic rate at this point were very similar to the observed value.

CO₂ response curve of I. rubescens leaves

The CO₂ response curve data of I. rubescens leaf photosynthesis are shown in Table 3.

Table 3.

CO₂ response curve of I. rubescens leaf photosynthesis (average).

CO2 (µmol·mol−1)	Photo (µmolCO2·m-−2·s−1)	Transpiration (mmolH2Om−2s−1)	Tleaf (°C)	PARin (µmol·m−2·s−1)
1981.063	13.28118	1.711614	30.67786	1200.049
1779.07	13.74822	2.076798	30.12322	1200.109
1477.98	13.9082	2.422849	30.99781	1199.903
1381.585	12.22414	2.429738	30.03088	1199.725
1181.821	11.9153	2.542288	30.79212	1198.881
983.2891	11.6798	2.593174	30.62088	1199.717
785.1578	10.43091	2.605263	30.6294	1199.769
588.2518	8.370093	2.598826	29.70778	1199.477
392.7176	5.688908	2.572764	29.82269	1199.592
361.7573	3.743928	2.235932	30.30235	1200.262
286.5714	2.378747	2.31753	30.69262	1200.178
256.2721	2.153212	2.340206	30.60159	1199.589
197.9524	1.113644	1.946278	30.54132	1201.117
148.4747	0.958305	2.032948	30.34006	1201.138
99.09576	0.396179	2.113611	30.11915	1201.403
69.56865	0.095351	2.193027	29.91523	1201.294
E	0.050712
M	0.00021
N	0.005891
CCP	101.7199 µmol·mol−1
E·CCP	1.107
CSP	1674.514 µmol·mol−1
PCSP	13.62882 µmol·m−2·s−1
R2	0.98526

Note: CCP is the CO₂ compensation point. CSP is the CO₂ saturation point. PCSP is the net photosynthetic rate at the CO₂ saturation point. PARin is the PAR in the leaf chamber.

Based on the collected data, the curve of the CO₂ response of I. rubescens leaf photosynthesis is shown in Fig. 3.

Figure 3.

CO₂ response curve of I. rubescens leaf photosynthesis.

The CO₂ response curve of I. rubescens leaf photosynthesis indicates that the net photosynthetic rate was obviously related to the concentration of CO₂ in the air when the CO₂ concentration was below 1000 µmol·mol⁻¹. However, the effect of the CO₂ concentration on the net photosynthetic rate was not obvious when the concentration of CO₂ was above 1000 µmol·mol⁻¹.

The results of the CO₂ response curve fitted with the modified rectangular hyperbola model are shown in Table 3. The fitted CO₂ saturation point and the net photosynthetic rate at this point were very similar to the observed value.

Chlorophyll fluorescence characteristics of I. rubescens leaves

The results of the slow kinetics of chlorophyll fluorescence are shown in Table 4.

Table 4.

Slow kinetics of chlorophyll fluorescence.

PAR (µmol·m−2·s−1)	(Fm-Fo)/Fm	Y(II)	Y(NPQ)	Y(NO)	qN	qP	ETR (μmol electrons·m−2·s−1
198	0.725319	0.25	0.468	0.282	0.755	0.528	20.8
198	0.75982	0.239	0.534	0.227	0.835	0.57	19.9
198	0.727212	0.274	0.453	0.273	0.731	0.527	22.8
Average	0.7374	0.2543	0.485	0.261	0.7737	0.5417	21.167

The slow kinetics of chlorophyll fluorescence of I. rubescens leaves indicates that the maximal photochemical efficiency of photosystem II ((Fm-Fo)/Fm) in I. rubescens leaves reached 0.7. The electron transport rate of photosystem II in I. rubescens leaves reached 20 μmol electrons/(m²·s). The fraction of energy dissipated as heat via the regulated photoprotective NPQ mechanism (Y(NPQ)) was much more than that passively dissipated in the form of heat and fluorescence (Y(NO)).

The results of the rapid light curves of chlorophyll fluorescence in I. rubescens leaves are shown in Table 5. The rapid light curve of chlorophyll fluorescence in I. rubescens leaves is shown in Fig. 4.

Table 5.

Rapid light curve of chlorophyll fluorescence in I. rubescens leaves (average).

PAR	Y(II)	Y(NPQ)	Y(NO)	NPQ	qN	qP	qL	ETR
0	0.7900	0.000	0.210	0.000	0.000	1.0000	1.0000	0.00
6	0.6763	0.018	0.306	0.059	0.067	0.8670	0.5890	1.70
31	0.4633	0.206	0.330	0.644	0.458	0.6663	0.3803	6.03
101	0.2837	0.421	0.295	1.456	0.683	0.4687	0.2593	12.03
198	0.2007	0.531	0.268	1.997	0.763	0.3597	0.2000	16.70
363	0.1363	0.601	0.263	2.299	0.795	0.2550	0.1377	20.77
619	0.0920	0.646	0.262	2.465	0.822	0.1877	0.1047	23.93
981	0.0677	0.669	0.263	2.546	0.828	0.1393	0.0767	27.87
1386	0.0540	0.684	0.262	2.617	0.833	0.1120	0.0613	31.43
2015	0.0397	0.699	0.261	2.682	0.837	0.0837	0.0457	33.60
2970	0.0320	0.708	0.260	2.734	0.842	0.0690	0.0380	40.17
3588	0.033	0.716	0.251	2.849	0.834	0.065	0.033	49.9
4292	0.027	0.725	0.248	2.922	0.838	0.054	0.027	49.4
a	−0.00000359906
b	0.033474211
c	5.362466019
Fv’/Fm’ x ETR factor/2		0.33167 electrons/photons
Alpha		0.08 electrons/photons
ETRmax		52.333μmol electrons/(m2·s)
Ik		705.3μmol electrons/(m2·s)
Fv’/Fm’		0.78988
R2		0.99038

Note: Fv’/Fm’ x ETR factor/2 is the maximum quantum yield of PSII with a saturated pulse after dark adaptation. Alpha is the initial slope. ETRmax is the maximum electron transport rate. Ik is the minimum saturation of the light intensity.

Figure 4.

Rapid light curve of chlorophyll fluorescence in I. rubescens leaves.

The rapid light curve of chlorophyll fluorescence in I. rubescens leaves was automatically fitted with a PAM-2500 portable chlorophyll fluorescence apparatus according to the model of Eilers and Peeters [5]. The fitted results are shown in Table 5.

The rapid light curve of chlorophyll fluorescence in I. rubescens leaves indicates that the maximum quantum yield of PSII with a saturated pulse after dark adaptation (Fv’/Fm’ x ETR factor/2) was higher than the effective quantum yield of PSII (Y(II)). The initial slope (alpha) signifying the maximum photosynthetic efficiency was higher than the apparent quantum yield fitted in the light response curve of I. rubescens leaves.

Discussion and Conclusion

The modified rectangular hyperbola model is suitable for fitting light response curves and CO₂ response curves. We compared the fit of the light response curve and CO₂ response curve of Paeonia lactiflora created with different models. It was found that the fit results based on the modified rectangular hyperbola model were more similar than the results from other models to the observed values⁶.

I. rubescens is a heliophyte plant, which can tolerate intense light. There are very few reports about photosynthesis of I. Rubescens. There was no obvious midday depression of photosynthesis in I. rubescens leaves in terms of this study. The midday photosynthetic depression occurred in most of plants. The factors such as intense light, high air temperature, low soil moisture, low air humidity and so on can cause midday photosynthetic depression^7–10. There is no midday photosynthetic depression in some other plants, such as C₄ plants (Characterized by the Hatch-Slack photosynthetic pathway), CAM plants (plants with crassulacean acid metabolism) and aquatic plant^11,12. Some plants perform midday photosynthetic depression in a certain environment but express no midday photosynthetic depression in another environment. Their performances are affected by environment or some chemicals^13–16. The environment of I. rubescens studied in this paper was consistent with that of yield I. rubescens. It was sunny day and the light intensity was highest in a year in the locality when the data were determined. I. rubescens performed no midday photosynthetic depression in the severe environment, which indicated that it would similarly perform in suitable environment. Therefore, I. rubescens can tolerate intense light.

There was no obvious difference between the net photosynthetic rate of light saturation point and that of light intensities of 2000 µmol·m⁻²·s⁻¹ although the light saturation point of I. rubescens leaves was 802.262 µmol·m⁻²·s⁻¹. Therefore, there was no obvious effect of intense light above the light saturation point on the photosynthesis of I. rubescens leaves. The net photosynthetic rate of the light intensities of 1484.135 µmol·m⁻²·s⁻¹ was the highest in diurnal variation of photosynthesis because the temperature was suitable for it at that time. I. rubescens can also tolerate low light. Leaves of I. rubescens can utilize low light (i.e., at an intensity of 20 µmol·m⁻²·s⁻¹). Therefore, I. rubescens can grow on shady slopes. The most important factor affecting the photosynthetic efficiency in I. rubescens leaves is the concentration of CO₂ in the air. Photosynthesis in I. rubescens leaves was not obviously affected by high concentrations of CO₂ alone.

The maximum electron transport rate (ETRmax) in I. rubescens leaves was far higher than the observed electron transport rate (ETR). The chlorophyll fluorescence characteristics of I. rubescens leaves showed that there was very large potential for photosynthesis in I. rubescens leaves. The fraction of energy dissipated as heat via the regulated photoprotective NPQ mechanism (Y(NPQ)) was much more than that passively dissipated in the form of heat and fluorescence (Y(NO)). The minimum saturation light intensity (Ik) was far less than the light saturation point (LSP). Therefore, I. rubescens leaves can tolerate intense light.

I. rubescens performs no midday photosynthetic depression and can tolerate intense light. It can utilize low light and possesses high value of Fv/ Fm (the maximal photochemical efficiency of photosystem II). This indicated that I. rubescens leaves have a strong photoprotective capacity. However, the growth and cultivation of I. rubescens are affected by many factors such as light, air temperature, rainfall, soil, and so on^17,18. This study is aimed at the photosynthetic and chlorophyll fluorescence characteristics of I. rubescens. The suitable environment for the growth and cultivation of I. rubescens still needs to study.

Materials and Methods

Instruments

Li-6400 Photosynthesis system (LI-6400 Inc., Lincoln, NE, USA). PAM-2500 portable chlorophyll fluorescence apparatus (PAM-2500, Walz, Germany).

Materials

Approximately 60 I. rubescens plants were dug up from Taihang Mountain and evenly planted in 12 flowerpots (30 cm in diameter and 35 cm in depth) in March 2018. Then, the plants were irrigated to ensure that they grew well.

Determination of photosynthetic characteristics

The photosynthetic characteristics of mature leaves on the I. rubescens plants were determined on June 5–7 (sunny day, the light intensity is highest in a year), 2019. The concentration of CO₂ in the air was approximately 370 µmol·mol⁻¹ when the diurnal variation of photosynthesis was determined. The temperature of the leaf chamber was set at 30 °C, and the concentration of CO₂ in the leaf chamber was set at 400 µmol·mol⁻¹ when the light response curve was determined. The light intensity in the leaf chamber was set at 1200 µmol·m⁻²·s⁻¹, and the temperature of the leaf chamber was set at 30 °C when the CO₂ response curve was determined. These photosynthetic characteristics were determined with the Li-6400 Photosynthesis system. Each determination was repeated three times.

Determination of chlorophyll fluorescence characteristics

The fluorescence characteristics of mature leaves on the I. rubescens plants were determined on June 7–8, 2019. The leaves were under dark adaptation for 30 min before the determination of the chlorophyll fluorescence characteristics. The slow kinetics of chlorophyll fluorescence were determined before determining the light curve of chlorophyll fluorescence. The tests were repeated three times.

Data analysis

The light response curve and CO₂ response curves were analysed with SPSS (Statistical Product and Service Solutions, International Business Machines Corporation, USA). The light response curve and CO₂ response curve were all fitted with a modified rectangular hyperbola model⁴.

Modified rectangular hyperbola model:

$$\mathrm{Photo}=\mathrm{E}·\left(1-\mathrm{M}·\mathrm{PAR}\right)·\left(\mathrm{PAR}-\mathrm{LCP}\right)/(1+\mathrm{N}·\mathrm{PAR})$$

PAR is the value of light intensity in the light response curve (or the value of concentration of CO₂ in CO₂ response curve). Photo is the net photosynthetic rate. LCP is the light compensation point (or CO₂ compensation point). E, M and N are parameters. E is also the apparent quantum yield. The dark respiration rate under the light compensation point = E·LCP. The light saturation point is calculated as follows:

$$\left(\mathrm{LSP}\right)=\left(\right((\mathrm{M}+\mathrm{N})·(1+\mathrm{N}·\mathrm{LCP})/{\mathrm{M})}^{1/2})/-1)/\mathrm{N}.$$

The net photosynthetic rate under the light saturation point (LSP) or CO₂ saturation point (CSP) can be calculated according to the model.

The data related to the determination of the light curve of chlorophyll fluorescence were automatically fitted according to the model of Eilers and Peeters⁵.

The model of Eilers and Peeters is as follows:

$$\mathrm{ETR}=\mathrm{PAR}/(\mathrm{a}·{\mathrm{PAR}}^2+\mathrm{b}·\mathrm{PAR}+\mathrm{c})$$

ETR is the electron transport rate of photosynthetic system II. PAR is the fluorescence intensity. The letters a, b and c are parameters.

Author contributions

JIAN Zaiyou. Designed the study, implemented the experiment and wrote the manuscript. ZHOU Xiuren. Participate in the experiment. TIAN Jing. Participate in the data analysis.

Data availability

Data have been permanently archived: 10.5061/dryad.bg79cnp7h.

Competing interests

The authors declare no competing interests.