The spookiness is at an all time high and a seasonal force disrupted the Root Lab's game plan this week. After a few minor explosions and other mishaps, we finally got our stress tests running on Thursday. The late start pushed our NSC analysis back a week, but don't you worry, I'm fulfilling my promise and explaining the process in full, no more cliff-hangers! Keep reading to find out more about how we measure our woody friends' energy reserves and a special Halloween treat exploring the complex history of "witchcraft" and spell-binding plant potions.
If you can remember all the way back to the blog update on August 5th when I first explained how plants store and use nonstructural carbohydrates (NSCs), then I applaud you for your great recall and dedicated readership. If not, then please begin reviewing this material every night before you go to bed. Someday a random stranger will surely ask you about NSCs, and your preparedness for such a question might determine the fate of the universe. Okay, back to the carbohydrates.
NSCs play a central role in many plant metabolic processes. They are the main product of photosynthesis, like McDonald's French fries or Levi's jeans. They provide the energy for plant biosynthesis and are therefore involved in most all physiological processes. Scientists have just begun to explore the influence of NSCs on plant response to environmental change, specifically regarding drought. They also happen to be a reoccurring guest on this blog, so let's get into their molecular structure and how we can measure them in plant tissue.
Plants biosynthesize numerous compounds that we classify as NSCs. All are molecularly light weight mono- and oligosaccharides (simple sugars), polysaccharides (starch), and/or fructans. Below are some examples of carbohydrate molecules (Image source). There are many methods to measure NSCs, some are better at extracting and quantifying specific compounds, so deciding on which protocols to use depends on which compounds you want to target in your analysis.
The three main methods of NSC analysis are ion chromatography (IC), enzymatic conversion, and oxidation via concentrated acids. IC separates and quantifies a wide range of mono- and oligosaccharides, as well as any present sugar alcohols in mixtures (sugar alcohols are another NSC compound synthesized by some plant taxa). Mass spectrometers can then be used to identify the separated compounds. Enzymatic methods quantify the simple sugars (sucrose, fructose, and glucose) in mixtures. They do this by converting the sugars into molecules that can be monitored at a certain wavelength using a Ultraviolet-Visible spectrometer (UV-Vis spectrometer). We chose to go with the acid method for our study because acids oxidize all water-soluble sugars and storage NSCs. The concentration of the oxidized products are measured by adding color-producing reagents and running the samples through a spectrophotometer, just like we did with our stress tests last week. We will be using phenol-sulfuric acid for the reagent because that method has shown the best results in previous studies. Speaking of results, what are we expecting to see in our trees?
Eons ago, in a blog far-far-away, I first introduced the different 'root strategies' that some of our trees may employ to tolerate the stress of waterlogging. Whatever story our NSC analysis reveals will surely be an interesting one, but for now it remains somewhat of a mystery. We are particularly interested in observing how the stem and coarse root NSC pools shift after the recovery period for both magnolia species. Remember, we harvested stem and root tissue on the day we took the treated trees out of the kiddie-pools, and then again after a three-week recovery period. If rapid die-back of the root system occurs during sustained waterlogging, followed by a new flush of root growth once the environment is drained of water and reoxygenated, then the mechanisms of this strategy may be partially explained through our NSC results. Higher levels of NSCs in the stems and coarse roots during the earlier harvest date, and lower NSCs in the latter would provide evidence supporting this strategy of carbohydrate allocation.
Biosynthesis: simply put, biosynthesis includes all the processes in an organism's body that turns simple structures or molecules into more complex ones. The conveyer belts driving this synthesis are the metabolic pathways, and the factory workers are enzymes. Products can be modified, converted into completely different molecules, or joined to synthesize macromolecules.
Physiological processes: linked to biosynthesis, physiology involves all the inner-workings of an organism's cells, tissues, organs, and organ systems, and how they function to sustain life. Yes, plants have organs too (leaves, stems, roots, and reproductive parts), but they may be more viewer friendly than our own. Displays of delicately curated human organ bouquets on your dining room table typically leads to confused and worried house guests.
Monosaccharides: saccharides are the 'building units' that make up carbohydrates. They contain carbon, hydrogen, and oxygen atoms. Monosaccharides are the simplest of these saccharides, meaning they cannot be broken down, or hydrolyzed, to any simpler sugar. Examples include glucose, fructose, and galactose.
Oligosaccharides: carbohydrate chains containing three to ten simple sugars. Examples include sucrose, lactose, and maltose. The Greek words oligos and oligoi mean "small" and "few."
Polysaccharides: loooooong carbohydrate chains, the most abundant form of carbohydrate found in nature and food. These chains can be linear or branched and are linked together by glycosidic bonds. Polysaccharides can serve both energy and structural purposes. For plants and humans, starch is an important polysaccharide energy source. Glycogen is another you may be familiar with, as it is the main source of fuel for your brain and muscles (sorry plants, you miss out on this one). Structural polysaccharides include cellulose and chitin.
Fructans: long chains of fructose molecules, that (according to wikipedia) are produced in over 12% of angiosperms. Not every species of plant or tree synthesizes fructans as an NSC compound.
Ion chromatography (IC): a form of liquid chromatography that separates the molecules in mixtures based on their charge. An ion, or charged molecule, can be positively charged (cation), or negatively charged (anion). The resin, which is the solid matrix with attached ions to filter your sample, can be either positive or negative. Your sample's ions will attach to to the oppositely charged ions in the resin, leaving you with the remaining cations/anions to quantify.
Mass spectrometers: used to identify unknown compounds, quantify known compounds, and determine chemical properties of molecules. Mass specs do this by first converting molecules to gas-phase ions and then measuring the mass-to-charge ratio of those ions.
UV-Vis spectrometer: like the above, UV-Vis is a type of organic spectrometry that uses a machine called a spectrometer to beam a form of energy at molecules and then measures the response of said molecules. Mass specs use high energy electrons, while UV-Vis specs beam ultraviolet or visible light at whatever molecules you're analyzing.
I unknowingly bit off more than I could chew when selecting witches and botanical elixirs for a holiday themed post. As the costumed kiddos were running around my neighborhood, having also just bitten off more than they could reasonably masticate in the form of caramel filled chocolate bars instead of preliminary research, I sat on my porch reading horrifying details about witch-hunts for nearly six hours. I'm not sure what I was expecting - a couple of articles covering the Salem witch trials and a listicle of witchy herbal remedies might have been a more digestible read, but that's not what I found. Where does the myth of the broomstick wielding magician stem from?
Accounts of magic, sorcery, and witchcraft predate the witch trials by many centuries. Homer's Odyssey, published by Harper Collins in 800 BC, describes the character Circe as a witch. Archeological remains of Greek curse tablets have been found near fountains and grave sites. These spells were often used in sporting events and legal battles. Next time you see a discus thrower hauling around a huge stone tablet at the Olympics, you'll know what it's for. The familiar image of the old, malevolent witch, popularized by folk tales and Disney, wasn't first described until centuries later in the cold and desolate Alps.
The Celtic peoples and other medieval inhabitants of the largest and most extensive mountain range covering south-central Europe are said to have started the legacy of Bertha (also known as Perchta or Befuna). Bertha was depicted as an old, haggardly woman, who was cold in demeanor and possessed magical powers. She rewarded those who did good, and punished those who didn't fulfill their responsibilities. Her focus seemed to be on children and servants, particularly regarding their household and wool spinning duties. I don't want to be presumptuous here, but it kinda sounds like Bertha was a behavioral tool invented by distressed Celtic parents. Now that the origins of the witch have been explained, allow me to attempt to cram the horror of the witchcraze (15th-18th centuries) into the following paragraphs so that we can swiftly shift the topic to magical plants.
Let me begin by stating that the mass persecution of some 30,000-60,000 women and men during the European and Colonial American witch-hunts is a serious and tragic topic that I most certainly won't do justice. If you're looking for a deep dive on witchcraft and the socio-political factors that initiated the hysteria during this era, check out this article (and this)! When researching, I also found this podcast to be extremely informative. To give a brief recount of what I learned, just before the end of the middle ages in Europe (1450's), religious tension, political unrest, and the culturally accepted suspicion and violence towards women led to an increase in witch-hunts across Germany, France, Italy, Switzerland, and the Low Countries. Theological theories of "demonology" fueled these hunts, and those accused were allowed no defensive counsel. This same moral panic infected the lives of Colonial Americans, most notably during the late 17th century witch trials in Salem, Massachusetts.
Surprisingly, of the 144 citizens accused in Salem, none proved to possess any demonic powers. Why then, were they indicted in the first place? When societal unrest reaches its peaks, humanity in turn reaches for control and condemnation to combat the fear and instability of the times. The accused weren't mid-wives or natural healers in their villages, as commonly misinterpreted. Neither were they gathering at witches' Sabbath, or participating in any other mystical rituals. They were normal people, who just so happened to rub someone else in town the wrong way. With that being said, association with herbal remedies and potions were particular past-times that could secure one a spot on the "maybe a witch?" list.
During this period, the general population had a reservoir of plant knowledge that is lost on us today. All medicines and most materials were still derived from plants at the time. Everyone had a grandmother, sister, or cousin, who could prepare a herbal remedy at their bedside if they came down with the sniffles. Plant potions were widely used and normalized ... except when they didn't work. Unfortunately, poor treatment outcomes could lead to herbal practitioners being accused of witchcraft. Imagine telling your doctor "and just so ya know, this rash better clear up soon," with a wink and sly grin as you leave their office. Let's explore some of the plants and remedies used to treat rashes and expel the evil spirits back in the day.
Vervain, Verbena Genus
Being a cosmopolitan genus found in the Americas, Europe, Africa, and Asia, these plants have a deep history in many ancient cultures and traditions. Egyptians believed the plant originated from the tears of the goddess Isis. Others used them to make protective boundaries from evil spirits. Vervain has anti-inflammatory properties which added to its use as a medicinal herb.
Henbane (Hyoscyamus niger)
The Hyosciamus genus contains 17 species, one of which is the black henbane pictured above. Henbane belongs to the family Solanaceae, the nightshades, as do a couple of the other plants I will cover. Historically used for its psychoactive compounds and other healing properties, henbane is now taken as a treatment for intestinal and respiratory issues, tremors, and nausea.
Wolf's bane, Aconitum genus
Native to the mountain ranges of western and central Europe, this lethal plant got its name from its use as a poison frequently added to animal bait. The toxic chemical, aconitine, was also used to coat swords and arrows during wartimes. It would later become associated with witchcraft and murderous potions.
Jimsonweed (Datura stromonium), Datura genus
I present to you another proud Solanaceae member, native to the Americas and now found across the globe. Jimsonweed is known for both its psychoactive and healing properties. During the Spanish Inquisition, use of the plant could lead to immediate prosecution. Currently, Jimsonweed is used to treat asthma, influenza, and nerve diseases.
Mandrake (Mandigora officinarum)
Here's our third representative of the Solanaceae family, the mandrake. This plant belongs to the genus Mandragora, which contains three species native to central Asia and the Mediterranean. Many of you may notice this humanoid looking veg from Harry Potter. If that's not a claim to fame, then I don't know what is.
Rosemary (Rosmarinus officinalis)
Rosemary was sacred to ancient Egyptians, Greeks, and Romans. The Virgin Mary was said to have laid her cloak over a bush while resting, turning the flowers blue and legally changing the plant's government name to "Rose of Mary." It was a tedious process for the plant, but a very relaxing process for Mary.
Mugworts/wormwood, Artemisia genus
The Artemisia genus is quite large, containing 500 species found mostly in North America, Europe, and Asia. Used as a key ingredient in absinthe, a remedy to expel intestinal parasites, and a magical potion conjuring love and protection, this plant served many functions.
Blackthorn (Prunus spinosa)
Native to northern Africa, western Asia, and Europe, this prickly shrub of the rose family has a deep history in Celtic folk-lore. Fairies stood guard at blackthorn thickets and witches used the blackthorn twigs as wands.
catnip/catswort (Nepeta Cataria)
Native to Europe, the Middle East, Central Asia, and parts of China, catnip is used in teas among many cultures for its sedative and healing properties. Also consumed as a juice, tincture, or inhaled smoke, catnip remedied intestinal issues, indigestion, fever, hives, and nervous disorders. The plant's essential oils can be extracted and used as a natural insect repellent. Catnip's tendency to attract cats and pollinators is another plus (unless you're not a cat person).
Belladonna/Deadly nightshade (Atropa belladonna)
The belladonna belongs to the genus Atropa which contains three species native to Europe, Asia, and north Africa. Like the other members of Solanaceae, the plant's psychoactive chemicals were absorbed through ointments rubbed on the skin. In addition to the hallucinogenic and euphoric effects of the belladonna, it (along with all the other nightshades covered earlier) can cause seizures, comas, and even death - so maybe just stick to cocoa butter for your skin hydration needs.