New Raman sensor for precision agriculture
Stressed basil plant

Sometimes stress can just creep up on you. Lack of sleep, lack of nutritious food and not drinking enough water have a cumulative effect. Yes, it’s a tough life being a basil plant. Happily precision agriculture techniques can now give early indication that crops are getting stressed. A new portable Raman sensor may help farmers to spot when plants need additional care and attention.

Researchers from the Disruptive & Sustainable Technologies for Agricultural Precision group, part of the Singapore-MIT Alliance for Research and Technology, recently reported an intriguing new method to measure plant nutrients non-invasively. Working at the Temasek Life Science Laboratory in Singapore in collaboration with colleagues at MIT in Massachusetts, they developed and tested a handheld Raman spectroscopy probe for plants. Their innovation was capable of measuring phytonutrients such as beta-carotene and lycopene and nitrate ions (a macronutrient).

Raman spectroscopy is a chemical analysis technique that produces a kind of fingerprint of the vibrations of molecules. Laser light of a single colour is focused onto the sample of interest and light scattered by the sample is collected with a lens or mirror. Virtually all of the light is scattered with the same energy as the laser but something like 1 in a billion of the laser photons is scattered with less energy. The energy difference between the laser and the scattered light corresponds to the energy of a vibration of the molecule. Heavy molecules vibrate slowly, light ones more quickly and large molecules have many different vibrations while simple molecules like oxygen O2 have just one. When the scattered light is dispersed by a prism or grating, the scattered light is separated into a spectrum. Different vibrations of molecules can be identified by the ‘fingerprint’ spots of light in the spectrum.

Raman spectra of five plant leaves, simple nitrate nutrients and four carotenoid phytonutrients (courtesy Nature Scientific Reports)

As the figure above shows, small nitrate molecules have simple spectra with few vibrations but bigger molecules like the carotenoids lycopene, neoxanthin and beta-carotene have many more vibrational Raman peaks. The Singapore-MIT team recorded Raman spectra from the leaves of six different leaf vegetables: arabidopsis, spinach, Pak Choi, Choy Sum, lettuce and Kailan. The spectra of the leafy vegetables look very similar because the leaves are made from similar components (typically lignin, cellulose, pectin and carotenoid) giving the plant cell walls structure and internal colour.

Raman plant probe comprising: an optical fibre probe for laser excitation and Raman collection and magnetic leaf holder (courtesy Nature Scientific Reports)

Surprisingly, the probe was sufficiently sensitive to detect nitrate ions in the water in the leaves. Using a prototype Raman leaf probe, the team compared hydroponic plant growth with sufficient nitrate and with insufficient nitrate. Signal levels were not as good as those of a laboratory Raman instrument but taking a ratio of the 1046 cm-1 nitrate peak and a nearby 1067 cm-1 peak as a comparison, consistently lower ratios of nitrate were seen in the stressed, nitrogen deficient plants. The figure below compares nitrate starved arabidopsis plants with unstressed controls.

Clear differentiation between nitrate starved arabidopsis plants (-N) and control plants (+N) (courtesy Nature Scientific Reports)

Using a different salad plant, Choy Sum, the handheld Raman probe was able to categorise temperature and drought stress. This time a more sophisticated chemometric technique was used to reveal differences between the three groups of plants.

Raman probe measurement of stressed Choy Sum and control plants, showing small but clear spectral differences (courtesy Nature Scientific Reports)

The photographs show that the plants are starting to wilt due to the lack of water and raised temperature. Less obviously, principal component analysis (PCA) of the Raman spectra was able to categorise the plants into three clusters corresponding to control, drought stressed and temperature stressed conditions.

Chemometric classification of stressed Choy Sum plants (courtesy Nature Scientific Reports)

The Singapore-MIT team have reported a fascinating new method of detecting stress in a number of salad crop plants. They developed an innovative Raman leaf probe instrument and combined with detailed analysis of the spectral data, have an encouraging proof of principle. Further development is required to demonstrate earlier detection of crop stress. Basil plants can look forward to a more relaxed, less stressed future.

Read the recent Nature Scientific Report here. The Singapore-MIT have published additional work on measuring N, P, K nutrients in plants and a convincing genotype study of nitrate uptake in Arabidopsis.