Chapter 4. Apple (Malus × domestica)

Introduction

The apple Malus x domestica Borkh. is a widely distributed, temperate zone fruit crop (Figure 4.1) that has been cultivated for millennia.

Figure 4.1. Apple fruit and seed

Source: Maks Narodenko/Shutterstock.com.

The apple is a member of the Rosaceae family, Amygdaloideae subfamily, Maleae tribe, Malinae subtribe and Malus genus (Potter et al., 2007). The Rosaceae family is distributed worldwide and includes a range of economically important fruit crop species such as the pome fruit species (for example: apple, pear and quince), the stone fruit species (for example: sweet and sour cherry, plum, prune, apricot and peach) and the berry fruit species (for example: strawberry, blackberry and raspberry). The genus Malus consists of six  sections with 27 primary species (Forsline et al., 2003). Most of the species belonging to the Malus genus are cross-compatible, hence natural and artificial hybridisation techniques have resulted in numerous interspecific hybrids and secondary species.

The domestication of apple took place around 4 000 to 10 000 years ago in the Tien Shan Mountains of Central Asia. The origin of the Malus genus is said to be southeast of the People’s Republic of China and the species of the genus were distributed from there in all directions.

The presumed main ancestor of the cultivated apple is Malus sieversii (Ledeb.) M. Roem., which grew wild in the forests of Central Asia from Tajikistan to Western People’s Republic of China (Luby, 2003; Hancock et al., 2008). Based on genomic studies performed in the last century, other species belonging to the Malus genus have contributed to the genome of the cultivated apple. These were mainly Malus sylvestris (L.) Mill. distributed from Western Asia to Europe, and Malus orientalis Uglitzk which grew in the forests of the Caucasus region (Hancock et al., 2008).

The cultivated apple belongs to the genus Malus. The binomial denomination of the cultivated apple Malus × domestica Borkh. reflects its interspecific origin and replaces the former name M. pumila Mill. as well as other names like Pyrus malus L. Malus × domestica is allopolyploid (2n = 2x = 34), gametophytic incompatible, mostly self-unfruitful and requiring pollination. Most of the cultivars are diploid; however, a number of tri- and tetraploid cultivars also exist.

Further description on the apple taxonomy, geographic distribution, centres of origin and diversity, reproductive biology, genetics, hybridisation and introgression, interaction with other organisms (ecology), common pests and pathogens, and biotechnological developments can be found in the OECD Consensus Document on the Biology of Apple (OECD, 2019).

Production of apples

Cultivars

It is estimated that 40 cultivars account for the bulk of commercial production worldwide. Some cultivars have given rise to many mutants, which have been selected for growth habit, fruit colour (Figure 4.2), ripening time and other characteristics. Economically important mutants are especially known among the cultivars “Delicious”, “Jonagold” and “Gala”.

Figure 4.2. Fruit colour/shape diversity of some apple cultivars

Source: cynoclub/Shutterstock.com.

Regional differences in the relative importance of apple cultivars are evident and the choice of cultivars varies from country to country. For instance, southern Europe produces many “Golden Delicious” whereas “Elstar” and “Jonagold” are popular in northern Europe. Australia and New Zealand are major apple exporters based on “Gala”, “Granny Smith” and “Braeburn”. The leading cultivar in the People’s Republic of China is “Fuji”. In many regions of North America, “McIntosh” and “Delicious” are important cultivars; however, many others such as “Fuji”, “Pink Lady”, “Gala”, “Braeburn” and “Jonagold” are also popular.

Uses and processing

Appropriate comparators for testing new cultivars

This document suggests parameters that apple breeders should measure when developing new cultivars.

The data obtained in the analysis of a new apple cultivar should ideally be compared to those obtained from an appropriate near-isogenic non-modified variety, grown and harvested under the same conditions.1^,2 The comparison can also be made between values obtained from new varieties and data available in the literature or chemical analytical data generated from commercial apple cultivars.

Components to be analysed include key nutrients, anti-nutrients, toxicants and allergens. Key nutrients are those which have a substantial impact on the overall diet of humans (food) and animals (feed). These may be major (fats, proteins, and structural and non-structural carbohydrates) or minor constituents (vitamins and minerals). Similarly, the levels of known metabolites and allergens should be considered. Key toxicants are those toxicologically significant compounds known to be inherently present in the species, whose toxic potency and levels may impact human and animal health. Standardised analytical methods and appropriate types of material should be used, adequately adapted to each product and by-product. The key components analysed are used as indicators of whether unintended effects of the genetic modification influencing plant metabolism have occurred or not.

Breeding characteristics screened by developers

Prior to 1900, apple improvement was based on finding chance seedlings with good fruit quality. Scientific breeding work began in the early 20^th century as leading horticultural/agricultural experiment stations and institutes were just being established at that time worldwide. Apple breeding started to be based on controlled crosses combining the best characteristics of cultivars chosen as mother or pollen parents.

Apple cultivars have been developed by selection of desired fruit phenotypes (appearance, uniformity, size, firmness, juiciness, crispiness, taste), as well as for agronomic characteristics (yield, stability of yield, tree growth, pruning effort), resistance to diseases and tolerance to abiotic stress. Apple cultivars are generally propagated vegetatively on rootstocks. The rootstock can impact characteristics that are important for commercial production, for example, vigour of vegetative growth and fruit size. The choice of the rootstock is also important for the purpose of the cultivar, e.g. commercial fruit production or landscape growing.

The traits of major interest for modern breeding programmes include better quality or increased marketability of the fruit, improved storability, reduced production costs, as well as improved disease and pest resistance. Molecular techniques have been developed to facilitate and accelerate apple breeding. Molecular markers have been developed for several disease resistance genes, as well as for some quality traits (Costa et al., 2005; Peil et al., 2011). Marker-assisted seedling selection is applied already by some breeders (Baumgartner et al., 2015). The genome of the apple has been published (Velasco et al., 2010), which will aid in developing more markers. Genomic selection, a statistical approach for estimating breeding potential, has been demonstrated as a tool that could be useful for selecting fruit quality traits (Kumar et al., 2012). Furthermore, there are continuing efforts to address some of the breeding bottlenecks using innovative breeding technologies, like cisgenesis and fast-track breeding approaches (Flachowsky et al., 2007; CFIA, 2014).

Constituents of apple fruits

The composition of apples varies greatly among cultivars. Environmental factors such as climate, soil condition, site of cultivation and storage conditions after harvest have an influence on the overall composition of the fruit. Sugars, organic acids and polyphenol compounds are responsible for the apple’s main sensory attributes of sweetness, acidity and bitterness. The ripening process alters apple composition, which affects the consistency as well as the taste.

Constituents of products and by-products from apple processing

Changes in chemical composition during storage

Apples are still living organisms after harvest and have an active metabolism. Respiration and metabolic activity lead to degradation and transformation of apple metabolites like sugars, acids and vitamins. For example, malic acid is the major substrate for respiration and, therefore, the concentration decreases during storage (Vandendriessche et al., 2013).

To reduce the loss of nutrients, it is important to store apples at temperatures from 0-2°C, if the cultivars are not sensitive to chilling injuries. Low temperatures decelerate respiration and metabolic activity, resulting in slower ripening and senescence of the apples; however, it is difficult to maintain acceptable fruit quality beyond 6 to 8 months of storage. A combination of low temperature and controlled atmosphere (CA, defined as oxygen concentration held at 1%-3% and carbon dioxide at 1%-5%, adjusted according to cultivar) can preserve apple quality over longer storage times. Apples stored under CA at 1°C were shown to be firmer and contain higher levels of acid and vitamin C compared to apples stored at the same temperature under air (Schirmer and Trierweiler, 2005). Apple firmness is strongly correlated with the pectin content of the cell wall. The expression of endogenous enzymes that modify pectin is controlled by ethylene. Apples stored under CA produce less ethylene, which results in less pectin modification over time (Gwanpua et al., 2014; Storch et al., 2015). The pectin metabolising enzymes exhibit activity even at a storage temperature of 4°C, so maintaining the appropriate temperature is important (Gwanpua et al., 2014).

Maintenance of vitamin levels, especially vitamin C, during long-term storage under controlled atmosphere also varies with cultivar. Vitamin C concentration stayed more or less stable in the cultivars “Topaz” and “Braeburn” during seven months of CA-storage at 1°C. In contrast, other cultivars like “Jonagold” and “Fuji” lost about 50% of their vitamin C after several months of storage (Trierweiler, Krieg and Tauschen, 2004). CA storage also results in reduced levels of the volatile esters and other compounds that impart “Gala” apples with their aroma, due to the inhibition of precursor biosynthesis at low oxygen concentrations (Both et al., 2014). The combination of lower concentrations of volatiles and ethylene results in slower ripening of the fruit, however, amino acid and polyphenol levels were unchanged with CA storage (Amarowicz et al., 2009; Both et al., 2014).

Cultivars that are highly sensitive to CA storage include “Braeburn” and “Empire”, which are very susceptible to internal browning at elevated carbon dioxide concentrations (Lee et al., 2011; Hatoum et al., 2016). In “Braeburn” apples, this quality degradation is marked by biochemical changes, including higher levels of aspartate, acetaldehyde, ethanol and ethyl esters, decreased levels of glutamate, and at very high carbon dioxide levels, an increase of cellobiose, which might indicate a cell wall breakdown (Hatoum et al., 2016). Flesh browning can potentially be reduced by treating the apples with antioxidants before storage (Lee et al., 2012).

Other investigations to maintain the nutrient quality of apples have been carried out with 1-methylcyclopropene (1-MCP; Smartfresh®). 1-MCP is an ethylene inhibitor which slows down or inhibits the ripening of the apples by binding to the ethylene receptor and therefore influencing the metabolite profile, for example, amino acids such as threonine, glutamate, ethanol, methanol and volatiles (Lee et al., 2011; Hatoum et al., 2016). Treatment with 1-MCP improved storability by slowing down ripening and reduced flesh browning in certain apple cultivars like “Braeburn” and “Empire” (Fawbush, Nock and Watkins, 2008).

In summary, storage of apples at low temperatures and controlled atmosphere can effectively maintain nutrient quality of many apple cultivars over several months; however, other cultivars would require special treatment or conditions. Further investigation would be necessary to determine the optimal storage conditions for sensitive cultivars, both existing and new.

Allergens

Apple oral allergy is one of the most common fruit allergies. The prevalence of sensitisation (assessed by skin prick test) was around 4.2% in the general population (both children and adults) in Germany (Zuberbier et al., 2004) and 0.1% in children (2-14 years) in France (Rancé et al., 2005). The prevalence of a perceived allergy to apple varied from 0.9% to 8.5% in European children and was estimated to be 0.5% in adults in a study including patients from Europe, the United States, Australia and New Zealand (Woods et al., 2001).

Four main classes of apple allergens have been identified so far. Two major allergens, Mal d 1 and Mal d 3, are responsible for most apple allergies in the general population. The Mal d 1 protein is the main apple allergen observed in northern Europe. It cross-reacts with Bet v1, the main allergen in birch pollen, due to its similar structure. The Mal d 3 protein is the main allergen in the Mediterranean area (Schmitz-Eiberger and Matthes, 2011). Symptoms of an allergy caused by Mal d 3 can be more severe (for example, generalised urticaria, vomiting and abdominal pain) and in rare cases are even life-threatening. The Mal d 3 protein cross-reacts with the peach allergen Pru p 3. The Mal d 1 protein is unstable so that people with an allergy often tolerate processed apple products like stewed fruit, cakes and pasteurised juices. The process of pasteurisation most likely eliminates allergenicity. In contrast, Mal d 3 is very stable and resistant to heating. In this case, even processed apple products can cause an allergic reaction. The last two proteins (Mal d 2 and Mal d 4) are considered minor allergens and are also involved in the birch-apple syndrome. Even if apple allergens seem to be mainly restricted to these four families of proteins, several additional proteins located in fruit tissues have been reported as potential allergens (Savazzini, Ricci and Tartarini, 2015).

Different apple cultivars cause a difference in intensity of symptoms. Scientific studies showed that apple cultivars differ in the content of internal factors involved in apple allergy (Matthes et al., 2009).

Toxicants

Apple seeds contain small amounts of amygdalin (D-mandelonitrile-β-D-gentiobioside), a cyanogenic glycoside. Cyanogenic glycosides are naturally occurring plant toxins and are stored in the vacuoles within plant cells to serve as important chemical defence compounds against herbivores (Bolarinwa, Orfila and Morgan, 2015). Total cyanogen content was determined to be 1.08 mg CN⁻/g apple seeds (Surleva and Drochioiu, 2013). The amygdalin levels were measured in desiccated apple seeds for 15 apple varieties and ranged from 0.95 ± 0.22 to 3.91 ± 0.49 mg/g (Bolarinwa, Orfila and Morgan, 2015). The lethal dose for cyanide is reported to be 0.5-3.5 mg/kg body weight (bw). An acute reference dose (ARfD) was estimated to be 20 µg/kg bw (EFSA CONTAM Panel, 2016).

Degradation of amygdalin by enzymes can lead to the production of cyanide (prussic acid) when the seeds are macerated or crushed. Apple seeds are generally not consumed by humans but apple juice is produced from whole apples including the seeds. Among the commercially-available apple products, one variety of pressed apple juice had an amygdalin content of 0.09 mg/g, apple purée had 0.02 mg/g and a fruit smoothie had 0.01 mg/g. Amygdalin was not detected in the cider of two brands (Bolarinwa, Orfila and Morgan, 2014). The amygdalin content of 10 commercially-available apple juices ranged from 0.010 to 0.039 mg/mL (Bolarinwa, Orfila and Morgan, 2015). Although the level of amygdalin is considered low in juice in relation to the toxic level, the concentration might be higher in pomace and closer to a level with toxic effect in animals. However, due to the fact that a high level of amygdalin would only occur when the seeds are crushed, and this is not expected to be the normal situation in pomace, the level of amygdalin is expected to be well below any toxic effect level. For humans, the small amounts of amygdalin present in apple seeds and, in turn, apple juice are unlikely to present any health problems to consumers.

Other metabolites: Organic acids, phenolic compounds

Apples are a source of phenolic compounds because of their widespread consumption in many countries and their year-round availability. Phenolic compounds and organic acids are responsible for the characteristic acidic taste and astringency of the fruit (Campo et al., 2006). The most abundant organic acid in apples is malic acid (up to 90%) (Kyzlink, 1990) while citric and quinic acids are also present in substantial quantities (Fuleki, Pelayo and Palabay, 1995).

During ripening, there is a general tendency for a decrease in acidity (Campo et al., 2006). However, there is some disagreement concerning changes in phenolics during maturation and storage. Whether there is an increase or decrease in the amount of phenolic compounds seems to depend on the apple cultivar (Burda, Oleszek and Lee, 1990).

In apples, the content of phenolic compounds is represented mainly by epicatechin and procyanidin (Burda, Oleszek and Lee, 1990; Wojdylo, Oszmiański and Laskowski, 2008). Chlorogenic acid is also found in considerable amounts. It is important to note that phenolic compounds are present in the skin at a relatively higher level than in the flesh of the apples. This is especially true for the apple anthocyanins; apple cultivars with red or partially darkred peels are generally the richest sources of anthocyanins. It is also noteworthy that quercetin glycosides are essentially only located in the apple skin (Burda, Oleszek and Lee, 1990; Gorinstein et al., 2001; Veberic et al., 2005). Table 4.9 provides a list of “other metabolite” levels present in apple fruits. Additional information regarding concentration levels of phenolic compounds in apples based on their fresh weight can be found in Ceyman et al. (2012), Jakobek and Barron (2016) and Stracke et al. (2009).

Key products consumed by humans

The majority of apples are consumed as fresh apples for their flavour and nutritional qualities. Apples are a source of potassium and soluble fibre, including pectin and other complex carbohydrates, and phenolic antioxidants.

Suggested analysis for food use of new cultivars

The suggested key nutritional parameters to be analysed in apples for human food use are shown in Table 4.10. Demonstration that composition of a novel apple variety is as expected, i.e. similar to control and/or within reference ranges would be sufficient to extrapolate to juice and other processed apple products for food.

Key products consumed by animals

As reported above, most of the use of apple processing waste for animal feed is apple pomace, and most of the apple pomace is fed to ruminants. The nutrients of major concern are crude protein, crude fat, carbohydrate and ash, and, for ruminants, ADF and NDF. While calcium and phosphorus are very important minerals in animal feeds, measuring these nutrients in apple is not warranted, due to their very low concentrations relative to the dietary requirements for livestock. Total phenolics may be of importance due to their effects on protein digestibility. They are present mainly in the skin of the apple, which is concentrated in apple by-products fed to livestock.

Suggested analysis for feed use of new cultivars

Table 4.11 below shows the suggested nutritional and compositional parameters to be analysed in unprocessed apples and apple pomace for feed use. For comparative purpose, it is suggested that analysing either the apple fruit (unprocessed) or apple pomace would suffice. The nutrient content of the pomace would not be expected to change if the nutrient content of the apple fruit does not change.