Postharvest Chemical Treatment of Physiologically Induced Stem End Blockage Improves Vase Life and Water Relation of Cut Flowers

Abstract

Wound-induced xylem occlusion significantly affects the vase life of cut flowers, as oxidative stress and the polymerization of phenolic compounds lead to the deposition of phenolic compounds/secondary metabolites in the stem ends of cut flowers to heal open tissues of freshly cut stems and prevent microbial invasion. However, this deposition causes blockage of vessels, reduced water uptake, and shortened vase life. The physiological plugging of vessels is linked with various oxidative enzymes’ (PAL, PPOs, LACs, and COs) actions taken to increase the synthesis of different compounds, e.g., lignin, suberin, tyloses, gel, and latex, in wounded areas. The use of chemical preservatives/enzyme inhibitors is one of the safest and most efficient techniques employed to minimize vascular blockage and inhibit phenolic compounds deposition and exudation. This review mainly discusses the types of oxidative enzymes, their pathways and biochemistry along with production of secondary metabolites, their biosynthesis, and their modes of action involved in vascular blockage. It also summarizes the different types of preservatives used in postharvest treatments to improve relative water uptake, flower fresh weight, petal protein content, and hydraulic conductance and prolong the vase life of cut flowers during storage. It is hoped that this elaborate study will help researchers in designing new studies concerning occlusion caused by the accumulation of phenolic compounds in vessels.

1. Introduction

The plant vascular system is composed of xylem and phloem vessels. Xylem consists of tracheid elements and dead vessels that transport nutrients and water from roots to photosynthetic organs [1]. An uninterrupted supply of water through these cells provides hydraulic support to the stems of cut flowers [2]. Three important components determine water balance in cut flowers: water uptake from a vase solution, loss of water through transpiration, and the ability of a cut stem/flower to retain water [3]. The end of vase life is usually attributed to wilting, which occurs when there is an imbalance between water uptake by cut stems and water loss through transpiration despite being placed in vase water [4]. This imbalance is usually linked with a reduction in water transport through xylem because of vascular occlusion or due to increased cell membrane permeability, linked to senescence or stress [2]. When xylem vessels are blocked, there is a reduction in water uptake because of high hydraulic resistance; still, the respiration process is continuous, which leads to water imbalance and results in flower wilting due to a loss of turgidity [5].

Vascular occlusion occurs mainly because of three factors: (1) air embolism, (2) stem end blockage by bacteria and inorganic or organic debris, and (3) physiological blockage due to mechanical wounds (stem end cuts). All these different types of blockages reduce the stem hydraulic conductance of cut flowers, and among them, wound-induced blockage is difficult to repair and damaging in terms of the vase life and postharvest quality of cut flowers [3,4]. Despite being an important factor related to the postharvest vase life of cut flowers, wound-related vascular blockage is the least studied. Therefore, this study will provide a detailed study on wound-induced xylem plugging at the stem ends of cut flowers with an aim to identify various types of enzymes and phenolic compounds involved in vascular blockage and their inhibition through chemicals in the form of preservative solutions.

2. Methodology

We could not identify any previous review articles focusing solely on physiological blockage due to the wounding of cut flower stems. Our questions for the review were related to “How wounding induce vascular occlusion?”, “What type of enzyme leads to blockage?”, “What are the characteristics of deposited compounds cause blockage” etc. Research literature was collected from Elsevier, Google Scholar, Science Direct, NCBI, and PubMed. As it is the first-ever comprehensive study on wound-induced blockage, this review covers the literature (book chapters, previous review articles, conference papers, and research papers) published in the last 30 years (1995–2024). Furthermore, the data presented in tabulated form were also obtained from research studies carried out during this period. Different keywords such as “physiologically induced blockage”, “phenolic compounds deposition”, “lignin”, “oxidative enzymes inhibitors”, “phenylalanine ammonia lyase”, and “chemical preservatives” were used for the literature search. Overall, 157 research studies were analyzed to write a broad-perspective review paper, but after screening the research studies (not specifically related to the topic) and sorting them according to different oxidative enzymes and types of phenolic compounds, a total of 98 publications were included in this review.

3. Wounding

Normally, cutting is the first step in the postharvest management of cut flowers. However, when cut from their parent plants and placed in a vase solution, flowers experience both wounding stress and water stress followed by increased activity of antioxidant enzymes for neutralizing the damaging effects of reactive oxygen species [1]. Wounding stress leads to various physiological and biochemical defense mechanisms that involve a series of reactions, i.e., the synthesis of membrane-degrading proteins and free radicals, hormone signal transduction, structural changes in the cell wall, changes in the level of phyto-hormones and antioxidants, the accumulation and deposition of secondary metabolic compounds [6], an outburst of reactive oxygen species, and controlled cell death. All these reactions help plants to quickly heal wounds while protecting the stem ends from pathogen entry/infection [7]. Among the different defense mechanisms, the deposition of secondary metabolic compounds/phenolic compounds is the foremost response of plants that is triggered to protect freshly wounded tissues by compartmentalizing or isolating damaged tissues [8]. These compounds, according to their compositions, are callose, alkaloids, tannin, lignin, starch, gum (pectin) suberin, resin, tyloses, latex, and mucilage [5,9,10].

They are normally divided into three classes. The first concerns the exudation of material such, as latex (Papaveraceae, Asteraceae, and Euphorbiaceace), gums/resins (Rosaceae), and mucilage (monocots, i.e., Canna, Heliconia, Malvaceae, and Tiliaceae), from the cut surface. The second involves material (tannin and lignin) deposition in the lumen of xylem vessels by living cells, and it occurs in the genera Amelanchier, Acacia, and Prunus. The last one consists of tylose formation, and this response is produced by plants belonging to the genera Prunus and Eucalyptus [11].

Despite this physiological response of plants triggered to prevent microbial entry, xylem occlusion by material deposition at the wound site decreases the water absorption capacity of the cut stem, which leads to a reduction in vase solution/water uptake that ultimately affect cut flower freshness and vase life [7,12]. Proteins, polysaccharides, and lignin are considered the main components of vascular blockage in cut flowers [13], and these materials are usually deposited within 5 cm of the stem base [14].

The phenyl-propanoid pathway is an important pathway in plant secondary metabolites that leads to the production of phenolic compounds [15]. The initiation of secondary metabolite synthesis is positively correlated with enzyme activity in the phenyl-propanoid pathway [6]. Accumulation of these compounds starts with the deamination of phenylalanine to produce trans-cinnamic acid through the action of L-phenylalanine ammonia-lyase (PAL) [16]. PAL is an important enzyme involved in a plant’s reaction to a wound and functions as a catalyst in the phenylpropanoid pathway [6]. After the synthesis of trans-cinnamic acid, different catalytic reactions lead to the generation of different phenolic compounds, such as sinapic acid, ferulic acid, caffeic acid, and p-coumarate, which are converted into their respective monomers (sinapyl, coniferyl, feruloyl, and p-coumaryl alcohols) by esterification, methoxylation, hydroxylation, and reduction reactions [17]. These monolignols are then further polymerized by peroxidase enzyme (POD) into lignin and suberin [18].

Moreover, reactive oxygen species (ROS) also act as a major signaling molecule in response to wounding. They play important roles in the deposition of lignin xylem vessels soon after wounding and in the induction of genes responsible of the synthesis of PAL. Hydrogen peroxide produced in response to wounding can be detected at the cutting site, reaching the highest level after 4–6 h. Along with ROS production, various other molecules also work in positive correlation with ROS to enhance signal initiation after the wounding of the stem. These molecules include abscisic acid (ABA), ethylene, and jasmonic acid (JA) [19]. During oxidative stress, phenolic compounds are bound to phospholipids by hydrogen bonds, causing an accumulation of cinnamic-acid-rich compounds outside and inside the membrane, preventing the damage initiated by ROS [20,21].

Wound-induced blockage usually occurs in a 2 cm section of the basal end after cutting [6]. Re-cutting the stem at a 2.5–5 cm length helps in removing wound-induced deposition of phenolic compounds, but this blockage moves upward with the passage of time [22]. Moreover, additional wounding of tissues leads to the further deposition of phenolic compounds. Besides this, cut flower stem length is also an important quality criterion [23]. Thus, to prevent wound-induced xylem occlusion, the use of chemical inhibitors as a vase solution plays a vital role in negating the activities of oxidative enzymes and maintaining the vase life of cut flowers.

4. Effect of Chemical Inhibitors on Enzyme Activity

An enzyme inhibitor is a specific ion or small molecule that binds to an enzyme, reducing its activity. Inhibition of enzyme activity is known to be a major mechanism for controlling enzyme action [1] (

Table 1. Chemical inhibitors that prevent/reduce enzymatic activities in response to wound-induced xylem occlusion.

Enzyme	Chemical Inhibitor	Reference
Phenylalanine ammonia lyase (PAL)	Aminoxyacetic acid (AOA) l-α-aminooxy-phenylpropionic acid (AOPP)	[24]
	S-carvone	[25]
	2-aminoindan-2-phosphonic acid (AIP)	[26]
Polyphenol oxidase (PPO)	p-chlorophenol p-Nitrocatechol, p-Nitrophenol, Sodium metabisulfate	[22]
	Phenylhydrazine	[27]
	Pepsin Bromelain Papain	[28]
	Cysteine Citric acid	[10]
Laccase (LAC)	Cetyltrimethylammonium bromide (CTAB)	[29]
	Thioglycolic acid (TGA) Dithiothreitol (DTT) Diethyldithiocarbamate (DDC)	[1]
	Sodium azide	[30]
Catechol oxidase (CO)	Calcium chloride	[22]
	2-Mercapto-ethanol	[31]
	2,3-Dihydroxynaphthalene	[32]
	Tropolone	[33]
Peroxidase (PRX)	Cycloheximide (CHI)	[34]
	Phloroglucinol Propylgallate Butylated hydroxytoluene	[20]
	3-Amino-1,2,4, triazole (3-AT)	[22]
	Copper sulphate	[31]
	Hydroquinone	[32]
	Sodium metabisulphite	[35]
	L-cysteine 1,4-dithiothreitol 2-mercaptoethanol	[36]
	p-phenylenediamine (PPD) Catechol	[37]

). As blockage/occlusion involves oxidative reactions, it can be inhibited through antioxidant treatments [22].

4.1. Phenylalanine Ammonia Lyase (PAL)

The mechanical wounding of a cut stem leads to increased expression of many oxidative enzymes, such as phenoloxidase, peroxidase, PAL [1], etc. PAL catalyzes the first and most important step in the phenylpropanoid pathway that produces lignin, flavonoids, and anthocyanin [16], and it is considered one of the main regulatory enzymes in wound reaction [6]. PAL activity is usually initiated by low temperatures and light intensity [16]. The activity of this enzyme is also positively correlated with oxidative stress [21].

PAL activity significantly increases after tissue wounding [38]. Its activity started to increase after 12 h in the basal end of a 2 cm carnation (Dianthus caryophyllus) cut stem and reached the highest level after 48 h. Moreover, an increase in the expression level of two PAL genes, DcPAL2 and DcPAL3, also increases during wound-induced reactions [6]. In chrysanthemum (Dendranthema grandiflora), PAL activity increases up to 5–15-fold after the first day of cutting. However, its activity depends on plant genotypes, as the chrysanthemum var “Vikyng” (0.12 µmol h⁻¹g⁻¹ FW) exhibited higher activity after wounding compared to another variety, namely, “Cassa” (0.45 µmol h⁻¹g⁻¹ FW) [38]. In lily (Lilium lancifolium) cut stems, the activity of PAL was upregulated 6 h after wounding [7].

S-carvone, a monoterpene, reduced the biosynthesis of wound-healing compounds in Geraldton waxflower (Chamelaucium uncinatum) by inhibiting PAL activity and enhanced vase life by increasing solution uptake and the fresh weight of the flowers [25]. Salicylic acid (200 mg/L), due to its antioxidant role, downregulated PAL enzyme activity in rose (Rosa grandiflora), thus improving vase life up to 8 days [39]. The PAL inhibitor α-aminooxi-β-phenyl propionic acid (AOPP), at a concentration of 3 mM, significantly improved flower longevity by up to 70% as compared to a control by inhibiting polysaccharide formation in the cut stem of tuberose (Polianthus tuberosa) [40].

4.2. Polyphenol Oxidases

Lignin is formed through the oxidation and polymerization of monolignols, catalyzed by a group of enzymes such as phenol oxidase (PPOs), peroxidases (PRXs), and laccase (LACs), collectively known as phenoloxidases [1,41]. After wounding, plant tissues initiate defense mechanisms—such as different enzymes, i.e., methyl jasmonates, and salicylic-acid-enhanced PPO transcripts—that lead to increased expression of PPO genes [42].

4.2.1. Polyphenol Oxidase (PPOs)

Polyphenol oxidase, a key enzyme in physiological blockage, functions by oxidizing different alcohols, such as coniferyl, sinapyl, and p-coumaryl, known as precursors of lignin [5]. PPOs are copper-containing enzymes that are usually present in all life forms, including animals, plants, fungi, and bacteria. PPO genes normally regulate in response to biotic and abiotic stresses [41]. PPOs are usually localized in chloroplasts and encounter their phenolic substrates (present in vacuoles) in response to wounding due to a loss of cell membrane integrity [42,43]. These enzymes function in two steps. In the first step, the hydroxylation of monophenols to o-diphenols occurs, while in the second step, there is an oxidation of o-diphenols to quinones and semi-quinones [44]. As a result of these two steps, class III peroxidase leads to lignification, suberization, and the cross-linking of glycoproteins and polysaccharides [45]. Increased activity of PPO significantly reduced relative water uptake (RWU) in gerbera cut flowers [46].

Salicylic acid treatment, as a vase solution, decreased PPO activity by up to 41% on the second day of treatment and increased the vase life of gerbera (Gerbera jamesonii) by improving water uptake [46]. Furthermore, the pulse treatment of spermine and γ-Aminobutyric acid also had a role in reducing the activity of PPO in cut stems of gerbera [21]. Similarly, γ-Aminobutyric acid (GABA) at a 1 mM concentration functions as a PPO inhibitor by enhancing narcissus cut flower (Narcissus tazetta) fresh weight and maintaining membrane stability [45], whereas cerium nitrate and salicylic acid induce the lowest PPO activity along with an improvement in vase solution absorption and petal protein content of lisianthus (Eustoma grandiflorum) flowers [47]. Silver nanoparticles (SNPs), in 125 μM proportions, act as antioxidants by decreasing PPO activity, leading to an improvement in cell membrane stability, vase life, and total monomeric anthocyanin in gladiolus (Gladiolus grandiflorus) flower petals [48].

4.2.2. Laccase (LACs)

Lignin bases are formed through the polymerization of monolignols, a process catalyzed by two main enzymes: laccase (O₂-dependent) and peroxidase (H₂O₂-dependent) [1]. Laccases (EC 1.10.3.2 p-diphenol oxidoreductase), monomeric glycoproteins, are members of the blue copper family of oxidases [49]. They are found in plants, bacteria, fungi, arthropods, and archaea and are highly stable at different temperatures [50]. LAC enzymes are present in the walls of vascular cells and have a role in the polymerization of monolignols through the oxidation of lignin-forming compounds [51]. LACs oxidize various organic substrates, such as o- and p-monophenols and diphenols, using molecular oxygen as a terminal electron acceptor [50]. Their catalytic sites consist of four histidine-rich copper-binding proteins [52]. Laccase oxidizes lignin monomers to lignin by coupling the phenoxy radicals produced from the oxidation of lignin phenolic groups [41,50].

A total of 0.2 mM of dithiothreitol, a chemical inhibitor, minimizes laccase activity, controls lignin deposition, and delays anthocyanin degradation in gerbera (Gerbera jamesonii) cut flowers [1]. Sodium fluoride, a laccase inhibitor, in combination with sucrose and 8-HQS increased the vase life of gerbera by 12 days [53]. Another inhibitor, cetyltrimethylammonium bromide (CM), improved the fresh weight, water relations, and vase life of cut foliage of silver wattle (Acacia holosericea) [29].

4.2.3. Catechol Oxidase (COs)

PAL, along with catechol oxidase, is involved in wound-induced reactions in cut flowers [4]. Catechol oxidases (COs) are also known as PPOs due to their structure and substrate specificity [54]. Catechol oxidase is a copper-binding protein using molecular oxygen as a co-factor [52]. This enzyme (diphenoloxidase or tyrosinase) catalyzes two reactions in response to wounds: (i) the oxidation of phenolic compounds with OH groups (o-diphenol oxidase activity -O₂: o-diphenol oxidoreductase, EC 1.10.3.1) and (ii) the conversion of monophenols to o-dihydroxy phenols (monophenol monooxygenase activity (EC 1.14.18.1)) [22]. COs are membrane-bound or cytosolic enzymes, while their substrates are present in vacuoles. After a wound is inflicted, the membrane is lysed, causing both the enzyme and its substrate to come into contact, producing quinones for polymerization [55]. Vascular blockage due to COs is pH-dependent, as its activity is optimal at pH 5–6, whereas no blockage in chrysanthemum cut flowers was observed below pH 4 [22,56].

The CO inhibitor 4-hexylresorcinol (HR) at a 10 mM concentration significantly delayed leaf wilting by up to 30 days in Bouvardia (Bouvardia × domestica) cut stems by oxidizing phenolic compounds [31]. Pulsing treatment of chrysanthemum cut stems with p-phenylene diamide (10 mM) for 5 h significantly delayed wilting by up to 6 days [22]. The use of 4-HR as a catechol oxidase inhibitor increases relative fresh weight and vase solution uptake and delayed decline in hydraulic conductance in stems of spider flowers (Grevillea) [4].

4.2.4. Peroxidase Enzymes (PRXs)

The primary function of peroxidase, a haem-containing enzyme, is to oxidize phenolic compounds using H₂O₂ as a co-substrate [36]. PRXs are glycoproteins and are mainly localized in cell walls and vacuoles. Peroxidase has a role in many growth-related processes, such as cell wall extension, auxin catabolism, lignin synthesis, the production of reactive nitrogen species (RNS) and reactive oxygen species (ROS), and phytoalexin synthesis [57]. Peroxidase, in response to wounding, is released from the cell surface into the apoplast, where it exhibits both peroxidative and oxidative activities [29]. Moreover, PRXs are involved in the final polymerization step of lignin biosynthesis and play an important role in suberin synthesis [22]. Precursors of lignin are enzymatically dehydrogenated to phenoxy radicals in the cell wall, further polymerizing via peroxidase, producing a complex net of cross-linking cell wall components (polysaccharides, monolignols, and proteins) [58]. PODs exist in two main isoenzymatic forms, namely, anionic (acidic) and cationic (basic), based on their isoelectric points. Both forms are involved in the lignification and suberization process [37]. High temperature reduces the activities of these enzymes as they have protein structures and high temperature destroys those structures [10].

In rose (Rosa hybrida) cut flowers, increased peroxidase activity leads to reduced anthocyanin content, browning, and the de-coloration of petals [51]. POD inhibitors function by blocking the activities of iosenzymatic forms that are involved in lignin and suberin accumulation [37]. The peroxidase inhibitor catechol (1,2-dihydroxybenzene) in combination with 8-HQC significantly reduced xylem blockage in gerbera (Gerbera jamesonii) [14]. Hydroquinone (p-benzenediol) delayed wilting in Astilbe × arendsii flowers and 2-mercaptoethanol in bird of paradise (Strelitzia reginae) by inhibiting peroxidase activity [32,36], while amitrol (3-Amino-1,2,4-triazole) enhanced the vase life of Geraldton waxflower (Chamelaucium uncinatum) by improving water relation [29], and copper sulphate increased the vase life of lisianthus (Eustoma grandiflorum) by up to 4 days by increasing antioxidant enzyme (SOD and CAT) activities [59].

4.3. pH

Enzyme activities leading to lignification or suberization can be inhibited at low pH [60]. Low pH reduces blockage, and no incidence of material deposition in the vessel lumen is observed below pH 4 [22]. Thus, the use of 8-HQC and solutions with low pH significantly reduced vascular blockage, caused in response to a wound reaction [61]. As physiological plugging is enzymatically mediated, 8-HQC, by acidifying a solution, alters enzymatic activities [62].

5. Effect of Chemical Inhibitors on Secondary Metabolite Deposition

The occlusion/blockage of xylem vessels because of wounding occurs due to plant physiological responses. After a few hours of wounding, as a defense mechanism, lignin and suberin biosynthesis gene expression increase with the deposition of these compounds in the wounded area [8,51]. Other compounds, such as gel, tyloses, latex, etc., compartmentalize/isolate affected tissues, thus aiding the wound-healing process [8]. The phenomenon of the deposition of these materials in the lumen of water-conducting cells is found in many cut flower species [11]. Due to the blocking of xylem vessels, the rate of water absorption is less than that of water precipitation or transpiration, which leads to the development of negative water pressure [34]. This leads to premature wilting and termination of vase life [9] (Figure 1). Thus, the inclusion of chemical inhibitors as preservatives is one of the safest and most economical methods applied to prolong the vase life of cut flowers (

Table 2. Chemical inhibitors at different concentrations and under different environmental conditions minimize/prevent physiologically induced xylem vessel blockage.

Crop	Chemical Inhibitor	Concentration	Type of Preservative	Experimental Conditions	Results	Reference
Gerbera (Gerbera jamesonii)	Thioglycolic acid (TGA)	0.5 mM	20 h Pulsing	70 ± 5% Relative humidity 15 µmol m−2 s−1 Light intensity 20 ± 2 °C Temperature	Minimized lignin content in vessels	[1]
	Sodium azide	0.2 mM	10 h Pulsing	60 ± 5% Relative humidity 75 µmol m−2 s−1 Light intensity 10 °C Temperature	Improved healing of wounds and minimization of lignin deposition	[63]
	Catechol 8-HQC	1.0 mM 0.45 mM	Holding solution	35–55% Relative humidity 15 µmol m−2 s−1 Light intensity 18–22 °C Temperature	Decreased vascular occlusion by gums	[14]
Leather flower (Clematis spp)	8-HQC+ Sugar	200 mg L−1 + 20 mg L−1	Holding solution	60% Relative humidity 35 µmol m−2 s−1 Light intensity 20 °C Temperature	Minimized vessel occlusion was achieved due to tylose deposition	[64]
Silver wattle (Acacia holosericea)	Cu2+	2.2 mM	5 h Pulsing	60 ± 5% Relative humidity 18 µmol m−2 s−1 Light intensity 19 ± 2 °C Temperature	Prevented gel secretion from parenchyma cells	[65]
	S-carvone	0.636 mM	Holding solution	60 ± 10% Relative humidity 14 µmol m−2 s−1 Light intensity 20 ± 1 °C Temperature	Improved fresh weight and water uptake rate	[25]
Lotus (Nelumbo nucifera)	Citric acid	150 mg L−1	Holding solution	65–75% Relative humidity 15 µmol m−2 s−1 Light intensity 25 °C Temperature	Reduced latex flow from lactifiers	[27]
Geraldton waxflower (Chamelaucium uncinatum)	Phenyl hydrazine	10 mM	5 h Pulsing	60 % Relative humidity 15 µmol m−2 s−1 Light intensity 20 °C Temperature	Prevented phenolic compound deposition and improved water uptake	[29]
Lilac (Syringa vulgaris L.)	Chrysal professional		Holding solution	70 ± 5% Relative humidity 25 µmol m−2 s−1 Light intensity 18–20 °C Temperature	Reduced vessel blockage by tyloses by up to 9.6%	[66]
Spider flower (Grevillea spp)	S-carvone 4-hexylresorcinol	0.636 mM 2.5 mM	Holding solution	60 ± 10% Relative humidity 12 µmol m−2 s−1 Light intensity 20 ± 1 °C Temperature	Prevented stem end blockage by inhibiting suberin deposition	[4]
Astilbe (Astilbe x arendsi)	Copper sulphate	0.25 mM	Holding solution	60 ± 5% Relative humidity 15 µmol m−2 s−1 Light intensity 20 °C Temperature	Improved water uptake by inhibiting phenolic compound secretion	[32]
Chrysanthemum (Chrysanthemum grandiflorum)	p-phenylene diamide	10 mM	5 h Pulsing	60% Relative humidity 15 µmol m−2 s−1 Light intensity 20 °C Temperature	Delayed leaf wilting	[22]

).

5.1. Lignin

Lignin is a natural three-dimensional aromatic phenolic heteropolymer with a complex structure mainly deposited in secondary thickened cell walls [67]. It is used to defend plant cells and helps in maintaining cell wall rigidity [15,68]. Lignin can be categorized as guaiacyl (G), hydroxyphenyl (H), or syringal (S) lignin [69]. Moreover, it is resistant to biodegradation and serves as a structural barrier against insect attacks/pathogen colonization [70]. In flowers, the initiation of lignin biosynthesis is an important defense response of cut stems to wounding and microbial invasion [71]. Lignin is formed through the oxidative polymerization of three p-hydroxycinnamyls (p-coumaryl, coniferyl, and sinapyl) in the phenylpropanoid pathway with the help of the enzymes peroxidase, laccase, cinnamyl alcohol dehydrogenase (CAD), and cinnamoyl-CoA reductase (CCR) [68,72]. Deposition of lignin in wounded cells prevents the spread of toxins and enzymes from microbial invasion into the host plant. However, lignification at wounding sites also causes blockage of xylem conduits, thus reducing water uptake [7,67]. Physiological occlusion of xylem vessels due to lignin deposition was observed in chrysanthemum (Dendranthema grandiflora) [20], Bouvardia (Bouvardia × domestica) [31], Lisianthus (Eustoma grandiflorum) [37], and rose (Rosa grandiflora) [10].

In tulip (Tulipa gesnerana), an increase in lignin biosynthesis gene expression was observed 6–48 h after cutting [7]. Sodium azide (0.1 mM) and catechol (10 mM) influenced the maintenance of rose (Rosa hybrida) petal anthocyanin and leaf chlorophyll content by delaying senescence and decreasing lignin content [10]. Similarly, sodium azide and sodium fluoride enhanced water absorption and prevented occlusion at the cut ends of gerbera (Gerbera jamesonii) [63]. Chlorine dioxide (CIO₂) at 0.100 g/L improves the vase life of cut peony (Paeonia suffruticosa) by inhibiting the deposition of lignin and related enzyme activities [18]. An increase in cationic charge (K⁺) induced by using KCL in a vase solution also leads to the shrinkage of lignin compounds in pit membranes, thus improving stem hydraulic conductance of silver wattle stems (Acacia holosericea) [73].

5.2. Suberin

Suberin is a phenolic, aliphatic heteropolymer located in specialized cells such as exodermal and endodermal cells, bundle sheath cells, seed coats, the periderm of roots and shoots, etc. [74]. It is deposited in between the plasma membrane and the cell wall, thus forming a hydrophobic protective barrier against microbial invasion. Suberin not only provides strength to cells but also prevents microbial entry and minimizes water loss by sealing off a layer of suberized cells [70].

One of the important compound deposits in response to cell injury at the cut ends of stems is suberin (a lipid–phenolic complex) [4]. Generally, the first response of plant tissues to mechanical wounds such as those inflicted by pathogen invasion or injury is suberization. Suberin is a hydrophobic material attached to the cell wall. It helps in compartmentalization after wounding, and its formation starts within 24 h after wounding [75]. Xylem vessels at maturity are dead cells (lacking protoplasts), but parenchyma cells and xylem fibers are living cells. Hence, parenchyma cells are considered important sites for the synthesis of lipids (precursors to suberin). These lipid components are first synthesized in living cells and then moved towards the wounded area. In another study, it was observed that suberin was first produced in intercellular spaces between parenchyma cells close to wounded xylem cells and further moved into pit membranes and to vessels’ secondary walls. It only formed a thin layer on the xylem lumen during the early development stage; however, this small amount was enough to disturb water flow in cut flowers [9].

S-carvone has a role in inhibiting suberin formation, thus improving the hydraulic conductance of spider flower (Grevillea) stems at the basal end, therefore improving vase solution uptake and the fresh weight of cut flowers [4]. Similarly, the use of S-carvone as a vase solution also reduced stem blockage in the cut stems of weeping coast myrtle (Baeckea frutescens) [25].

5.3. Tyloses

Tyloses are present in many plant species. They are common defense responses of plants towards biotic (bacterial and vascular fungal infection) and abiotic stresses (mechanical injury, freezing, and flooding) [66]. The formation of tyloses starts with the accumulation of dead lumens of tracheary elements by living parenchyma cells. Tylose production is the foremost response of plant cells to wounding and pathogen infection [76].

Tyloses is described as a ballon-shaped uncontrolled cell growth that deposits in conducting vessels (xylem lumen) around the wounding area. However, it does not accumulate in large amounts to cause blockage; instead, other products of high molecular weight also produced with tyloses affect the fluidity of water channels [5]. Tyloses/gel formation reduces water transport efficiency in the xylem. Tyloses are among the most common types of vascular occlusions in sapwood xylem. Deposition of tyloses starts from as early as 12 h after a wound is inflicted in maize root xylem to 2 days in common reed around the wounded surface and slowly moves towards other areas [3]. During the formation of true tyloses, the pit membrane ruptures and degrades as the protective layer that forms on the inner side of parenchyma cell walls passes through it into the vessel lumens and expands to form tyloses which contain cytoplasm [77]. The degradation of the pit membrane (cellulose) is essential for tyloses formation [34]. Tyloses are usually composed of lignins and polycarbohydrates [78]. Formation of tyloses may protect plant vascular system from microorganisms as tyloses contain anti-bacterial compounds like coumarins, procatechuic acid, catechol, and flavonoids that prevent microbes from spreading. The most sensitive region to microorganism attack is the basal part (0–10 cm), and tyloses formation is usually concentrated around this area [66].

During a postharvest study of leather flower (Clematis), reduction in water uptake was mainly attributed to tyloses formation in parenchyma cells [13]. In an experiment, 40% of vessels of common lilac cut stems were covered with tyloses when placed in distilled water, whereas the use of a professional perseverative reduced this percentage to 10–15% [66]. Similarly, in snapdragon (Antirrhinum majus), the placement of cut stems in tap water increased blockage by tyloses up to 4.5-fold [78]. Preservatives normally reduce tylose formation in cut flowers, but it is not completely inhibited, as the whole lumen vessel is never occluded when a cut stem is placed in a preservative solution [76]. Moreover, a preservative solution of 8-HQC+ 2% sugar prevents tylose formation in xylem vessels of leather flower (Clematis L.) cut stems [64]. Nano silver at 1 mg/L in combination with 2% sucrose improved the vase life of snap dragon (Antirrhinum majus) by reducing buildup of tyloses in the vessel lumen of cut stems [78]. Furthermore, 8-HQC in combination with sucrose significantly dropped the production of tyloses in cut stem of lilac (Syringa vulgaris) [79].

5.4. Gel

Gel is a non-lignified pectinaceous material consisting of acidic polysaccharides. During their maturation process, gels are cross-linked by proteins and undergo polymerization, becoming hard [8]. Gels appear to be fibrillar, thin networks of varying electron density that fill and block vessels’ lumens [70]. Gel is composed of different components, such as phenolics, tannins, lipids, pectins, and carbohydrates [3]. Gel deposition in the vessel lumen usually occurs due to the digestion of cell walls between parenchyma cells and vessels [80]. All vessels along with their surroundings are in contact with paratracheal axial parenchyma cells that can exude gel [65]. Gels can protrude into conducting cells from neighboring cells through pits in the cell wall. Mature or hard gels are usually non-hydrated materials as they become resistant to solubilization. Their properties will change over time after interaction with other bio-chemicals such as proteins, phenolics, and mineral ions in cell sap. Pectins can be cross-linked by Ca²⁺ and can change from being basophilic to acidophilic as they mature [73]. Species having a minimum pit aperture of less than 3 mm secrete gels, while having an aperture of more than 3 mm will produce tyloses. Occlusion/blockage by gel narrows the vessel lumen area available for water transport, thus reducing stem hydraulic conductance [65]. Gel also has the potential to re-induce embolism in vessels when they are enzymatically degraded or partially hydrated due to a low solute and matric potential [2]. These occluding gels significantly reduced the stem hydraulic conductance of the cut stems of silver wattle (Acacia holosericea) [73]. Ratnayake et al. [8] observed that the secretion of gel into vessel lumens of silver wattle started 1–2 days after a wound was inflicted and was detected in xylem on day 3. The gels in silver wattle stems were originally colorless, but they adopted a red to brown color as they matured [81].

Cu²⁺, known as a wound reaction enzyme inhibitor, helps in preventing gel formation in vessels by disrupting the metabolic functions of the parenchyma cells contacting vessels and by inactivating the gel secretory process [65]. Moreover, Cu²⁺ inactivates antioxidant enzymes activities involved in the wound-healing process (the deposition of phenolic compounds) [82]; thus, the placement of cut silver wattle (Acacia holosericea) stems in a Cu²⁺ vase solution prevents gel formation in vascular cells, improves the rate of water uptake, and enhances vase life up to 3.8-fold [8,65].

5.5. Latex

Latex is a milky fluid consisting of tiny droplets of organic matter dispersed in an aqueous medium; it is better known as a milky sap stored in plant tissues, “lactifiers’, and is exuded or mobilized immediately to the site/point of damage after a wound is inflicted [83]. Latex contains a variety of secondary metabolites and defense chemicals such as chitin-bonding proteins, alkaloids, terpenoids, furanocoumarins, sugar, rubber, starch, phenolics, oils, enzymes (glucosidases, proteases, and chitinases), cardenolides, etc. It has a role in defense against pathogens and herbivores, covering damaged tissues and the excretion of waste metabolites [84]. After exudation, latex acts as a physical barrier because it rapidly coagulates on exposure to oxygen [85]. Latex has a significant role in vascular blockage in cut flowers, as latex becomes hardened when it encounters air and hardens more rapidly when placed in water, thus preventing vase solution uptake [86].

After cutting, lotus cut stems immediately exude a white milky sap produced in laticifers, linked with the xylem and phloem, which coagulates the cut surface and reduces or prevents water absorption [87]. Latex rapidly covers xylem vessel conduits freshly opened by cutting and entering these conduits. When latex-exuding stems were placed in vase water, latex became suspended in it and was taken along with water into the stem, thus blocking vessels [27]. The dipping of latex-containing cut stems into boiling water (scalding) prevents xylem occlusion by preventing latex exudation [88]. The pulsing of lotus (Nelumbo nucifera) stems with citric acid (150 mg L⁻¹) reduced latex flow from the cut stems [27]. Pretreatment of lotus stems with 90% isopropyl alcohol for 10 min completely stops latex flow due to its coagulation in lactifiers [87].

5.6. Tannins

Leaf blackening is one of the main postharvest disorders in protea (Protea cynaroides). It occurs due to the polymerization and oxidation of hydroxyphenols and tannins when a disruption in cell compartmentalization occurs due to the wounding of a cut stem [89]. When protea stems were placed in water, leuco-anthocyanidins, a condensed tannins, were leached form protea stems into the vase/preservative solutions. Thus, the uptake of these high-molecular-weight condensed tannins leads to vascular occlusion, which causes leaf blackening [90]. Tannins are usually water-insoluble and can be found as precipitate complexes [91]. Therefore, leaf blackening is usually minimized by using chemicals such as lead acetate and phenyl mercuric acetate, which have been used to precipitate these phenolic compounds from a vase solution [92]. Apart from these processes, microbial growth promoted by the leaching of these compounds also results in vascular blockage [90].

6. Ethylene Synthesis

Along with physiological plugging, mechanical wounding also induced the biosynthesis of endogenous ethylene and initiated abscission and senescence [88,93]. It has been observed that a significant amount of ethylene was produced from the cut stems of rose [61]. Ethylene induction usually occurs within minutes, hours, or days after wounding and then declines afterward [93]. In a study, significant amount of ethylene production after 4h of wounding were observed in styles and ovaries of carnation [94]. Furthermore, the metabolism of phenolic compounds is linked with ethylene signaling [29], as it initiates lignin formation in snapdragon stem by activating PAL enzyme activity [95], thus increasing lignin biosynthesis gene expression (PAL and 4CL) [96] and promoting tylosis formation in various cut flowers [97]. However, inhibitor of ethylene (STS) significantly reduced tyloses deposition in cut lilac stems (Syringa vulgaris) [98].

7. Conclusions

Physiological blockage is a wound-healing mechanism, and it occurs in cut flowers in response to mechanical damage (cutting). This process leads to xylem dysfunction and the deposition of phenolic compounds such as tannins, lignin, gel, suberin, and tyloses. But this deposition limits water uptake, induces water imbalance, and promotes flower senescence. Postharvest technology using chemical preservatives not only minimizes oxidative enzyme activity involved in the phenyl propanoid pathway but also inhibits the polymerization of phenolic compounds (precursor of lignin, suberin) to improve cut flower vase life.